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Biochemistry, the chemistry of biological molecules, explains the complex processes that sustain all living organisms. This college-level course taught by Professor Raffi Hovasapian is perfect for Pre-Medical students as well as Biology and Chemistry majors. Each lesson contains fundamental definitions, numerous worked-out examples, and many step-by-step explanations of complex processes. Professor Hovasapian carefully goes through how Proteins, Carbohydrates, Nucleic Acids and other biomolecules make life on Earth possible. Combining his triple degrees in Mathematics, Chemistry, and Classics, along with 10+ years of teaching experience, Professor Hovasapian expertly helps students understand difficult biochemical concepts. Raffi also teaches AP Chemistry, Multivariable Calculus, and Linear Algebra on Educator.

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I. Preliminaries on Aqueous Chemistry
  Aqueous Solutions & Concentration 39:57
   Intro 0:00 
   Aqueous Solutions and Concentration 0:46 
    Definition of Solution 1:28 
    Example: Sugar Dissolved in Water 2:19 
    Example: Salt Dissolved in Water 3:04 
    A Solute Does Not Have to Be a Solid 3:37 
    A Solvent Does Not Have to Be a Liquid 5:02 
    Covalent Compounds 6:55 
    Ionic Compounds 7:39 
    Example: Table Sugar 9:12 
    Example: MgCl₂ 10:40 
    Expressing Concentration: Molarity 13:42 
   Example 1 14:47 
    Example 1: Question 14:50 
    Example 1: Solution 15:40 
    Another Way to Express Concentration 22:01 
   Example 2 24:00 
    Example 2: Question 24:01 
    Example 2: Solution 24:49 
    Some Other Ways of Expressing Concentration 27:52 
   Example 3 29:30 
    Example 3: Question 29:31 
    Example 3: Solution 31:02 
  Dilution & Osmotic Pressure 38:53
   Intro 0:00 
   Dilution 0:45 
    Definition of Dilution 0:46 
    Example 1: Question 2:08 
    Example 1: Basic Dilution Equation 4:20 
    Example 1: Solution 5:31 
    Example 2: Alternative Approach 12:05 
   Osmotic Pressure 14:34 
    Colligative Properties 15:02 
    Recall: Covalent Compounds and Soluble Ionic Compounds 17:24 
    Properties of Pure Water 19:42 
    Addition of a Solute 21:56 
    Osmotic Pressure: Conceptual Example 24:00 
    Equation for Osmotic Pressure 29:30 
    Example of 'i' 31:38 
    Example 3 32:50 
  More on Osmosis 29:01
   Intro 0:00 
   More on Osmosis 1:25 
    Osmotic Pressure 1:26 
    Example 1: Molar Mass of Protein 5:25 
    Definition, Equation, and Unit of Osmolarity 13:13 
    Example 2: Osmolarity 15:19 
    Isotonic, Hypertonic, and Hypotonic 20:20 
    Example 3 22:20 
    More on Isotonic, Hypertonic, and Hypotonic 26:14 
    Osmosis vs. Osmotic Pressure 27:56 
  Acids & Bases 39:11
   Intro 0:00 
   Acids and Bases 1:16 
    Let's Begin With H₂O 1:17 
    P-Scale 4:22 
    Example 1 6:39 
    pH 9:43 
    Strong Acids 11:10 
    Strong Bases 13:52 
    Weak Acids & Bases Overview 14:32 
    Weak Acids 15:49 
    Example 2: Phosphoric Acid 19:30 
    Weak Bases 24:50 
    Weak Base Produces Hydroxide Indirectly 25:41 
    Example 3: Pyridine 29:07 
    Acid Form and Base Form 32:02 
    Acid Reaction 35:50 
    Base Reaction 36:27 
    Ka, Kb, and Kw 37:14 
  Titrations and Buffers 41:33
   Intro 0:00 
   Titrations 0:27 
    Weak Acid 0:28 
    Rearranging the Ka Equation 1:45 
    Henderson-Hasselbalch Equation 3:52 
    Fundamental Reaction of Acids and Bases 5:36 
    The Idea Behind a Titration 6:27 
    Let's Look at an Acetic Acid Solution 8:44 
    Titration Curve 17:00 
    Acetate 23:57 
   Buffers 26:57 
    Introduction to Buffers 26:58 
    What is a Buffer? 29:40 
    Titration Curve & Buffer Region 31:44 
    How a Buffer Works: Adding OH⁻ 34:44 
    How a Buffer Works: Adding H⁺ 35:58 
    Phosphate Buffer System 38:02 
  Example Problems with Acids, Bases & Buffers 44:19
   Intro 0:00 
   Example 1 1:21 
    Example 1: Properties of Glycine 1:22 
    Example 1: Part A 3:40 
    Example 1: Part B 4:40 
   Example 2 9:02 
    Example 2: Question 9:03 
    Example 2: Total Phosphate Concentration 12:23 
    Example 2: Final Solution 17:10 
   Example 3 19:34 
    Example 3: Question 19:35 
    Example 3: pH Before 22:18 
    Example 3: pH After 24:24 
    Example 3: New pH 27:54 
   Example 4 30:00 
    Example 4: Question 30:01 
    Example 4: Equilibria 32:52 
    Example 4: 1st Reaction 38:04 
    Example 4: 2nd Reaction 39:53 
    Example 4: Final Solution 41:33 
  Hydrolysis & Condensation Reactions 18:45
   Intro 0:00 
   Hydrolysis and Condensation Reactions 0:50 
    Hydrolysis 0:51 
    Condensation 2:42 
    Example 1: Hydrolysis of Ethyl Acetate 4:52 
    Example 2: Condensation of Acetic Acid with Ethanol 8:42 
    Example 3 11:18 
    Example 4: Formation & Hydrolysis of a Peptide Bond Between the Amino Acids Alanine & Serine 14:56 
II. Amino Acids & Proteins: Primary Structure
  Amino Acids 38:19
   Intro 0:00 
   Amino Acids 0:17 
    Proteins & Amino Acids 0:18 
    Difference Between Amino Acids 4:20 
    α-Carbon 7:08 
    Configuration in Biochemistry 10:43 
    L-Glyceraldehyde & Fischer Projection 12:32 
    D-Glyceraldehyde & Fischer Projection 15:31 
    Amino Acids in Biological Proteins are the L Enantiomer 16:50 
    L-Amino Acid 18:04 
    L-Amino Acids Correspond to S-Enantiomers in the RS System 20:10 
    Classification of Amino Acids 22:53 
   Amino Acids With Non-Polar R Groups 26:45 
    Glycine 27:00 
    Alanine 27:48 
    Valine 28:15 
    Leucine 28:58 
    Proline 31:08 
    Isoleucine 32:42 
    Methionine 33:43 
   Amino Acids With Aromatic R Groups 34:33 
    Phenylalanine 35:26 
    Tyrosine 36:02 
    Tryptophan 36:32 
  Amino Acids, Continued 27:14
   Intro 0:00 
   Amino Acids With Positively Charged R Groups 0:16 
    Lysine 0:52 
    Arginine 1:55 
    Histidine 3:15 
   Amino Acids With Negatively Charged R Groups 6:28 
    Aspartate 6:58 
    Glutamate 8:11 
   Amino Acids With Uncharged, but Polar R Groups 8:50 
    Serine 8:51 
    Threonine 10:21 
    Cysteine 11:06 
    Asparagine 11:35 
    Glutamine 12:44 
   More on Amino Acids 14:18 
    Cysteine Dimerizes to Form Cystine 14:53 
    Tryptophan, Tyrosine, and Phenylalanine 19:07 
    Other Amino Acids 20:53 
    Other Amino Acids: Hydroxy Lysine 22:34 
    Other Amino Acids: r-Carboxy Glutamate 25:37 
  Acid/Base Behavior of Amino Acids 48:28
   Intro 0:00 
   Acid/Base Behavior of Amino Acids 0:27 
    Acid/Base Behavior of Amino Acids 0:28 
    Let's Look at Alanine 1:57 
    Titration of Acidic Solution of Alanine with a Strong Base 2:51 
    Amphoteric Amino Acids 13:24 
    Zwitterion & Isoelectric Point 16:42 
    Some Amino Acids Have 3 Ionizable Groups 20:35 
    Example: Aspartate 24:44 
    Example: Tyrosine 28:50 
    Rule of Thumb 33:04 
    Basis for the Rule 35:59 
    Example: Describe the Degree of Protonation for Each Ionizable Group 38:46 
    Histidine is Special 44:58 
  Peptides & Proteins 45:18
   Intro 0:00 
   Peptides and Proteins 0:15 
    Introduction to Peptides and Proteins 0:16 
    Formation of a Peptide Bond: The Bond Between 2 Amino Acids 1:44 
    Equilibrium 7:53 
    Example 1: Build the Following Tripeptide Ala-Tyr-Ile 9:48 
    Example 1: Shape Structure 15:43 
    Example 1: Line Structure 17:11 
    Peptides Bonds 20:08 
    Terms We'll Be Using Interchangeably 23:14 
    Biological Activity & Size of a Peptide 24:58 
    Multi-Subunit Proteins 30:08 
    Proteins and Prosthetic Groups 32:13 
    Carbonic Anhydrase 37:35 
    Primary, Secondary, Tertiary, and Quaternary Structure of Proteins 40:26 
  Amino Acid Sequencing of a Peptide Chain 42:47
   Intro 0:00 
   Amino Acid Sequencing of a Peptide Chain 0:30 
    Amino Acid Sequence and Its Structure 0:31 
    Edman Degradation: Overview 2:57 
    Edman Degradation: Reaction - Part 1 4:58 
    Edman Degradation: Reaction - Part 2 10:28 
    Edman Degradation: Reaction - Part 3 13:51 
    Mechanism Step 1: PTC (Phenylthiocarbamyl) Formation 19:01 
    Mechanism Step 2: Ring Formation & Peptide Bond Cleavage 23:03 
   Example: Write Out the Edman Degradation for the Tripeptide Ala-Tyr-Ser 30:29 
    Step 1 30:30 
    Step 2 34:21 
    Step 3 36:56 
    Step 4 38:28 
    Step 5 39:24 
    Step 6 40:44 
  Sequencing Larger Peptides & Proteins 62:33
   Intro 0:00 
   Sequencing Larger Peptides and Proteins 0:28 
    Identifying the N-Terminal Amino Acids With the Reagent Fluorodinitrobenzene (FDNB) 0:29 
    Sequencing Longer Peptides & Proteins Overview 5:54 
    Breaking Peptide Bond: Proteases and Chemicals 8:16 
    Some Enzymes/Chemicals Used for Fragmentation: Trypsin 11:14 
    Some Enzymes/Chemicals Used for Fragmentation: Chymotrypsin 13:02 
    Some Enzymes/Chemicals Used for Fragmentation: Cyanogen Bromide 13:28 
    Some Enzymes/Chemicals Used for Fragmentation: Pepsin 13:44 
    Cleavage Location 14:04 
    Example: Chymotrypsin 16:44 
    Example: Pepsin 18:17 
    More on Sequencing Larger Peptides and Proteins 19:29 
    Breaking Disulfide Bonds: Performic Acid 26:08 
    Breaking Disulfide Bonds: Dithiothreitol Followed by Iodoacetate 31:04 
   Example: Sequencing Larger Peptides and Proteins 37:03 
    Part 1 - Breaking Disulfide Bonds, Hydrolysis and Separation 37:04 
    Part 2 - N-Terminal Identification 44:16 
    Part 3 - Sequencing Using Pepsin 46:43 
    Part 4 - Sequencing Using Cyanogen Bromide 52:02 
    Part 5 - Final Sequence 56:48 
  Peptide Synthesis (Merrifield Process) 49:12
   Intro 0:00 
   Peptide Synthesis (Merrifield Process) 0:31 
    Introduction to Synthesizing Peptides 0:32 
    Merrifield Peptide Synthesis: General Scheme 3:03 
    So What Do We Do? 6:07 
    Synthesis of Protein in the Body Vs. The Merrifield Process 7:40 
   Example: Synthesis of Ala-Gly-Ser 9:21 
    Synthesis of Ala-Gly-Ser: Reactions Overview 11:41 
    Synthesis of Ala-Gly-Ser: Reaction 1 19:34 
    Synthesis of Ala-Gly-Ser: Reaction 2 24:34 
    Synthesis of Ala-Gly-Ser: Reaction 3 27:34 
    Synthesis of Ala-Gly-Ser: Reaction 4 & 4a 28:48 
    Synthesis of Ala-Gly-Ser: Reaction 5 33:38 
    Synthesis of Ala-Gly-Ser: Reaction 6 36:45 
    Synthesis of Ala-Gly-Ser: Reaction 7 & 7a 37:44 
    Synthesis of Ala-Gly-Ser: Reaction 8 39:47 
    Synthesis of Ala-Gly-Ser: Reaction 9 & 10 43:23 
   Chromatography: Eluent, Stationary Phase, and Eluate 45:55 
  More Examples with Amino Acids & Peptides 54:31
   Intro 0:00 
   Example 1 0:22 
    Data 0:23 
    Part A: What is the pI of Serine & Draw the Correct Structure 2:11 
    Part B: How Many mL of NaOH Solution Have Been Added at This Point (pI)? 5:27 
    Part C: At What pH is the Average Charge on Serine 10:50 
    Part D: Draw the Titration Curve for This Situation 14:50 
    Part E: The 10 mL of NaOH Added to the Solution at the pI is How Many Equivalents? 17:35 
    Part F: Serine Buffer Solution 20:22 
   Example 2 23:04 
    Data 23:05 
    Part A: Calculate the Minimum Molar Mass of the Protein 25:12 
    Part B: How Many Tyr Residues in this Protein? 28:34 
   Example 3 30:08 
    Question 30:09 
    Solution 34:30 
   Example 4 48:46 
    Question 48:47 
    Solution 49:50 
III. Proteins: Secondary, Tertiary, and Quaternary Structure
  Alpha Helix & Beta Conformation 50:52
   Intro 0:00 
   Alpha Helix and Beta Conformation 0:28 
    Protein Structure Overview 0:29 
    Weak interactions Among the Amino Acid in the Peptide Chain 2:11 
    Two Principals of Folding Patterns 4:56 
    Peptide Bond 7:00 
    Peptide Bond: Resonance 9:46 
    Peptide Bond: φ Bond & ψ Bond 11:22 
    Secondary Structure 15:08 
    α-Helix Folding Pattern 17:28 
    Illustration 1: α-Helix Folding Pattern 19:22 
    Illustration 2: α-Helix Folding Pattern 21:39 
    β-Sheet 25:16 
    β-Conformation 26:04 
    Parallel & Anti-parallel 28:44 
    Parallel β-Conformation Arrangement of the Peptide Chain 30:12 
    Putting Together a Parallel Peptide Chain 35:16 
    Anti-Parallel β-Conformation Arrangement 37:42 
    Tertiary Structure 45:03 
    Quaternary Structure 45:52 
    Illustration 3: Myoglobin Tertiary Structure & Hemoglobin Quaternary Structure 47:13 
    Final Words on Alpha Helix and Beta Conformation 48:34 
IV. Proteins: Function
  Protein Function I: Ligand Binding & Myoglobin 51:36
   Intro 0:00 
   Protein Function I: Ligand Binding & Myoglobin 0:30 
    Ligand 1:02 
    Binding Site 2:06 
    Proteins are Not Static or Fixed 3:36 
    Multi-Subunit Proteins 5:46 
    O₂ as a Ligand 7:21 
    Myoglobin, Protoporphyrin IX, Fe ²⁺, and O₂ 12:54 
    Protoporphyrin Illustration 14:25 
    Myoglobin With a Heme Group Illustration 17:02 
    Fe²⁺ has 6 Coordination Sites & Binds O₂ 18:10 
    Heme 19:44 
    Myoglobin Overview 22:40 
    Myoglobin and O₂ Interaction 23:34 
    Keq or Ka & The Measure of Protein's Affinity for Its Ligand 26:46 
    Defining α: Fraction of Binding Sites Occupied 32:52 
    Graph: α vs. [L] 37:33 
    For The Special Case of α = 0.5 39:01 
    Association Constant & Dissociation Constant 43:54 
    α & Kd 45:15 
    Myoglobin's Binding of O₂ 48:20 
  Protein Function II: Hemoglobin 63:36
   Intro 0:00 
   Protein Function II: Hemoglobin 0:14 
    Hemoglobin Overview 0:15 
    Hemoglobin & Its 4 Subunits 1:22 
    α and β Interactions 5:18 
    Two Major Conformations of Hb: T State (Tense) & R State (Relaxed) 8:06 
    Transition From The T State to R State 12:03 
    Binding of Hemoglobins & O₂ 14:02 
    Binding Curve 18:32 
    Hemoglobin in the Lung 27:28 
    Signoid Curve 30:13 
    Cooperative Binding 32:25 
    Hemoglobin is an Allosteric Protein 34:26 
    Homotropic Allostery 36:18 
    Describing Cooperative Binding Quantitatively 38:06 
    Deriving The Hill Equation 41:52 
    Graphing the Hill Equation 44:43 
    The Slope and Degree of Cooperation 46:25 
    The Hill Coefficient 49:48 
    Hill Coefficient = 1 51:08 
    Hill Coefficient < 1 55:55 
    Where the Graph Hits the x-axis 56:11 
    Graph for Hemoglobin 58:02 
  Protein Function III: More on Hemoglobin 67:16
   Intro 0:00 
   Protein Function III: More on Hemoglobin 0:11 
    Two Models for Cooperative Binding: MWC & Sequential Model 0:12 
    MWC Model 1:31 
    Hemoglobin Subunits 3:32 
    Sequential Model 8:00 
    Hemoglobin Transports H⁺ & CO₂ 17:23 
    Binding Sites of H⁺ and CO₂ 19:36 
    CO₂ is Converted to Bicarbonate 23:28 
    Production of H⁺ & CO₂ in Tissues 27:28 
    H⁺ & CO₂ Binding are Inversely Related to O₂ Binding 28:31 
    The H⁺ Bohr Effect: His¹⁴⁶ Residue on the β Subunits 33:31 
    Heterotropic Allosteric Regulation of O₂ Binding by 2,3-Biphosphoglycerate (2,3 BPG) 39:53 
    Binding Curve for 2,3 BPG 56:21 
V. Enzymes
  Enzymes I 41:38
   Intro 0:00 
   Enzymes I 0:38 
    Enzymes Overview 0:39 
    Cofactor 4:38 
    Holoenzyme 5:52 
    Apoenzyme 6:40 
    Riboflavin, FAD, Pyridoxine, Pyridoxal Phosphate Structures 7:28 
    Carbonic Anhydrase 8:45 
    Classification of Enzymes 9:55 
    Example: EC 1.1.1.1 13:04 
    Reaction of Oxidoreductases 16:23 
    Enzymes: Catalysts, Active Site, and Substrate 18:28 
    Illustration of Enzymes, Substrate, and Active Site 27:22 
    Catalysts & Activation Energies 29:57 
    Intermediates 36:00 
  Enzymes II 44:02
   Intro 0:00 
   Enzymes II: Transitions State, Binding Energy, & Induced Fit 0:18 
    Enzymes 'Fitting' Well With The Transition State 0:20 
    Example Reaction: Breaking of a Stick 3:40 
    Another Energy Diagram 8:20 
    Binding Energy 9:48 
    Enzymes Specificity 11:03 
    Key Point: Optimal Interactions Between Substrate & Enzymes 15:15 
    Induced Fit 16:25 
    Illustrations: Induced Fit 20:58 
   Enzymes II: Catalytic Mechanisms 22:17 
    General Acid/Base Catalysis 23:56 
    Acid Form & Base Form of Amino Acid: Glu &Asp 25:26 
    Acid Form & Base Form of Amino Acid: Lys & Arg 26:30 
    Acid Form & Base Form of Amino Acid: Cys 26:51 
    Acid Form & Base Form of Amino Acid: His 27:30 
    Acid Form & Base Form of Amino Acid: Ser 28:16 
    Acid Form & Base Form of Amino Acid: Tyr 28:30 
    Example: Phosphohexose Isomerase 29:20 
    Covalent Catalysis 34:19 
    Example: Glyceraldehyde 3-Phosphate Dehydrogenase 35:34 
    Metal Ion Catalysis: Isocitrate Dehydrogenase 38:45 
    Function of Mn²⁺ 42:15 
  Enzymes III: Kinetics 56:40
   Intro 0:00 
   Enzymes III: Kinetics 1:40 
    Rate of an Enzyme-Catalyzed Reaction & Substrate Concentration 1:41 
    Graph: Substrate Concentration vs. Reaction Rate 10:43 
    Rate At Low and High Substrate Concentration 14:26 
    Michaelis & Menten Kinetics 20:16 
    More On Rate & Concentration of Substrate 22:46 
    Steady-State Assumption 26:02 
    Rate is Determined by How Fast ES Breaks Down to Product 31:36 
    Total Enzyme Concentration: [Et] = [E] + [ES] 35:35 
    Rate of ES Formation 36:44 
    Rate of ES Breakdown 38:40 
    Measuring Concentration of Enzyme-Substrate Complex 41:19 
    Measuring Initial & Maximum Velocity 43:43 
    Michaelis & Menten Equation 46:44 
    What Happens When V₀ = (1/2) Vmax? 49:12 
    When [S] << Km 53:32 
    When [S] >> Km 54:44 
  Enzymes IV: Lineweaver-Burk Plots 20:37
   Intro 0:00 
   Enzymes IV: Lineweaver-Burk Plots 0:45 
    Deriving The Lineweaver-Burk Equation 0:46 
    Lineweaver-Burk Plots 3:55 
    Example 1: Carboxypeptidase A 8:00 
    More on Km, Vmax, and Enzyme-catalyzed Reaction 15:54 
  Enzymes V: Enzyme Inhibition 51:37
   Intro 0:00 
   Enzymes V: Enzyme Inhibition Overview 0:42 
    Enzyme Inhibitors Overview 0:43 
    Classes of Inhibitors 2:32 
   Competitive Inhibition 3:08 
    Competitive Inhibition 3:09 
    Michaelis & Menten Equation in the Presence of a Competitive Inhibitor 7:40 
    Double-Reciprocal Version of the Michaelis & Menten Equation 14:48 
    Competitive Inhibition Graph 16:37 
   Uncompetitive Inhibition 19:23 
    Uncompetitive Inhibitor 19:24 
    Michaelis & Menten Equation for Uncompetitive Inhibition 22:10 
    The Lineweaver-Burk Equation for Uncompetitive Inhibition 26:04 
    Uncompetitive Inhibition Graph 27:42 
   Mixed Inhibition 30:30 
    Mixed Inhibitor 30:31 
    Double-Reciprocal Version of the Equation 33:34 
    The Lineweaver-Burk Plots for Mixed Inhibition 35:02 
   Summary of Reversible Inhibitor Behavior 38:00 
    Summary of Reversible Inhibitor Behavior 38:01 
    Note: Non-Competitive Inhibition 42:22 
   Irreversible Inhibition 45:15 
    Irreversible Inhibition 45:16 
    Penicillin & Transpeptidase Enzyme 46:50 
  Enzymes VI: Regulatory Enzymes 51:23
   Intro 0:00 
   Enzymes VI: Regulatory Enzymes 0:45 
    Regulatory Enzymes Overview 0:46 
    Example: Glycolysis 2:27 
    Allosteric Regulatory Enzyme 9:19 
    Covalent Modification 13:08 
    Two Other Regulatory Processes 16:28 
    Allosteric Regulation 20:58 
    Feedback Inhibition 25:12 
    Feedback Inhibition Example: L-Threonine → L-Isoleucine 26:03 
    Covalent Modification 27:26 
    Covalent Modulators: -PO₃²⁻ 29:30 
    Protein Kinases 31:59 
    Protein Phosphatases 32:47 
    Addition/Removal of -PO₃²⁻ and the Effect on Regulatory Enzyme 33:36 
    Phosphorylation Sites of a Regulatory Enzyme 38:38 
    Proteolytic Cleavage 41:48 
    Zymogens: Chymotrypsin & Trypsin 43:58 
    Enzymes That Use More Than One Regulatory Process: Bacterial Glutamine Synthetase 48:59 
    Why The Complexity? 50:27 
  Enzymes VII: Km & Kcat 54:49
   Intro 0:00 
   Km 1:48 
    Recall the Michaelis–Menten Equation 1:49 
    Km & Enzyme's Affinity 6:18 
    Rate Forward, Rate Backward, and Equilibrium Constant 11:08 
    When an Enzyme's Affinity for Its Substrate is High 14:17 
    More on Km & Enzyme Affinity 17:29 
    The Measure of Km Under Michaelis–Menten kinetic 23:19 
   Kcat (First-order Rate Constant or Catalytic Rate Constant) 24:10 
    Kcat: Definition 24:11 
    Kcat & The Michaelis–Menten Postulate 25:18 
    Finding Vmax and [Et} 27:27 
    Units for Vmax and Kcat 28:26 
    Kcat: Turnover Number 28:55 
    Michaelis–Menten Equation 32:12 
   Km & Kcat 36:37 
    Second Order Rate Equation 36:38 
    (Kcat)/(Km): Overview 39:22 
    High (Kcat)/(Km) 40:20 
    Low (Kcat)/(Km) 43:16 
    Practical Big Picture 46:28 
    Upper Limit to (Kcat)/(Km) 48:56 
    More On Kcat and Km 49:26 
VI. Carbohydrates
  Monosaccharides 77:46
   Intro 0:00 
   Monosaccharides 1:49 
    Carbohydrates Overview 1:50 
    Three Major Classes of Carbohydrates 4:48 
    Definition of Monosaccharides 5:46 
   Examples of Monosaccharides: Aldoses 7:06 
    D-Glyceraldehyde 7:39 
    D-Erythrose 9:00 
    D-Ribose 10:10 
    D-Glucose 11:20 
    Observation: Aldehyde Group 11:54 
    Observation: Carbonyl 'C' 12:30 
    Observation: D & L Naming System 12:54 
   Examples of Monosaccharides: Ketose 16:54 
    Dihydroxy Acetone 17:28 
    D-Erythrulose 18:30 
    D-Ribulose 19:49 
    D-Fructose 21:10 
    D-Glucose Comparison 23:18 
    More information of Ketoses 24:50 
    Let's Look Closer at D-Glucoses 25:50 
   Let's Look At All the D-Hexose Stereoisomers 31:22 
    D-Allose 32:20 
    D-Altrose 33:01 
    D-Glucose 33:39 
    D-Gulose 35:00 
    D-Mannose 35:40 
    D-Idose 36:42 
    D-Galactose 37:14 
    D-Talose 37:42 
   Epimer 40:05 
    Definition of Epimer 40:06 
    Example of Epimer: D-Glucose, D-Mannose, and D-Galactose 40:57 
   Hemiacetal or Hemiketal 44:36 
    Hemiacetal/Hemiketal Overview 45:00 
    Ring Formation of the α and β Configurations of D-Glucose 50:52 
    Ring Formation of the α and β Configurations of Fructose 61:39 
   Haworth Projection 67:34 
    Pyranose & Furanose Overview 67:38 
    Haworth Projection: Pyranoses 69:30 
    Haworth Projection: Furanose 74:56 
  Hexose Derivatives & Reducing Sugars 37:06
   Intro 0:00 
   Hexose Derivatives 0:15 
    Point of Clarification: Forming a Cyclic Sugar From a Linear Sugar 0:16 
    Let's Recall the α and β Anomers of Glucose 8:42 
    α-Glucose 10:54 
   Hexose Derivatives that Play Key Roles in Physiology Progression 17:38 
    β-Glucose 18:24 
    β-Glucosamine 18:48 
    N-Acetyl-β-Glucosamine 20:14 
    β-Glucose-6-Phosphate 22:22 
    D-Gluconate 24:10 
    Glucono-δ-Lactone 26:33 
   Reducing Sugars 29:50 
    Reducing Sugars Overview 29:51 
    Reducing Sugars Example: β-Galactose 32:36 
  Disaccharides 43:32
   Intro 0:00 
   Disaccharides 0:15 
    Disaccharides Overview 0:19 
    Examples of Disaccharides & How to Name Them 2:49 
    Disaccharides Trehalose Overview 15:46 
    Disaccharides Trehalose: Flip 20:52 
    Disaccharides Trehalose: Spin 28:36 
    Example: Draw the Structure 33:12 
  Polysaccharides 39:25
   Intro 0:00 
   Recap Example: Draw the Structure of Gal(α1↔β1)Man 0:38 
   Polysaccharides 9:46 
    Polysaccharides Overview 9:50 
    Homopolysaccharide 13:12 
    Heteropolysaccharide 13:47 
    Homopolysaccharide as Fuel Storage 16:23 
    Starch Has Two Types of Glucose Polymer: Amylose 17:10 
    Starch Has Two Types of Glucose Polymer: Amylopectin 18:04 
    Polysaccharides: Reducing End & Non-Reducing End 19:30 
    Glycogen 20:06 
    Examples: Structures of Polysaccharides 21:42 
    Let's Draw an (α1→4) & (α1→6) of Amylopectin by Hand. 28:14 
    More on Glycogen 31:17 
    Glycogen, Concentration, & The Concept of Osmolarity 35:16 
  Polysaccharides, Part 2 44:15
   Intro 0:00 
   Polysaccharides 0:17 
    Example: Cellulose 0:34 
    Glycoside Bond 7:25 
    Example Illustrations 12:30 
    Glycosaminoglycans Part 1 15:55 
    Glycosaminoglycans Part 2 18:34 
    Glycosaminoglycans & Sulfate Attachments 22:42 
    β-D-N-Acetylglucosamine 24:49 
    β-D-N-AcetylGalactosamine 25:42 
    β-D-Glucuronate 26:44 
    β-L-Iduronate 27:54 
    More on Sulfate Attachments 29:49 
    Hylarunic Acid 32:00 
    Hyaluronates 39:32 
    Other Glycosaminoglycans 40:46 
  Glycoconjugates 44:23
   Intro 0:00 
   Glycoconjugates 0:24 
    Overview 0:25 
    Proteoglycan 2:53 
    Glycoprotein 5:20 
    Glycolipid 7:25 
    Proteoglycan vs. Glycoprotein 8:15 
    Cell Surface Diagram 11:17 
    Proteoglycan Common Structure 14:24 
    Example: Chondroitin-4-Sulfate 15:06 
    Glycoproteins 19:50 
    The Monomers that Commonly Show Up in The Oligo Portions of Glycoproteins 28:02 
    N-Acetylneuraminic Acid 31:17 
    L-Furose 32:37 
    Example of an N-Linked Oligosaccharide 33:21 
    Cell Membrane Structure 36:35 
    Glycolipids & Lipopolysaccharide 37:22 
    Structure Example 41:28 
  More Example Problems with Carbohydrates 40:22
   Intro 0:00 
   Example 1 1:09 
   Example 2 2:34 
   Example 3 5:12 
   Example 4 16:19 
    Question 16:20 
    Solution 17:25 
   Example 5 24:18 
    Question 24:19 
    Structure of 2,3-Di-O-Methylglucose 26:47 
    Part A 28:11 
    Part B 33:46 
VII. Lipids
  Fatty Acids & Triacylglycerols 54:55
   Intro 0:00 
   Fatty Acids 0:32 
    Lipids Overview 0:34 
    Introduction to Fatty Acid 3:18 
    Saturated Fatty Acid 6:13 
    Unsaturated or Polyunsaturated Fatty Acid 7:07 
    Saturated Fatty Acid Example 7:46 
    Unsaturated Fatty Acid Example 9:06 
    Notation Example: Chain Length, Degree of Unsaturation, & Double Bonds Location of Fatty Acid 11:56 
    Example 1: Draw the Structure 16:18 
    Example 2: Give the Shorthand for cis,cis-5,8-Hexadecadienoic Acid 20:12 
    Example 3 23:12 
    Solubility of Fatty Acids 25:45 
    Melting Points of Fatty Acids 29:40 
   Triacylglycerols 34:13 
    Definition of Triacylglycerols 34:14 
    Structure of Triacylglycerols 35:08 
    Example: Triacylglycerols 40:23 
    Recall Ester Formation 43:57 
    The Body's Primary Fuel-Reserves 47:22 
    Two Primary Advantages to Storing Energy as Triacylglycerols Instead of Glycogen: Number 1 49:24 
    Two Primary Advantages to Storing Energy as Triacylglycerols Instead of Glycogen: Number 2 51:54 
  Membrane Lipids 38:51
   Intro 0:00 
   Membrane Lipids 0:26 
    Definition of Membrane Lipids 0:27 
    Five Major Classes of Membrane Lipids 2:38 
   Glycerophospholipids 5:04 
    Glycerophospholipids Overview 5:05 
    The X Group 8:05 
    Example: Phosphatidyl Ethanolamine 10:51 
    Example: Phosphatidyl Choline 13:34 
    Phosphatidyl Serine 15:16 
    Head Groups 16:50 
    Ether Linkages Instead of Ester Linkages 20:05 
   Galactolipids 23:39 
    Galactolipids Overview 23:40 
    Monogalactosyldiacylglycerol: MGDG 25:17 
    Digalactosyldiacylglycerol: DGDG 28:13 
    Structure Examples 1: Lipid Bilayer 31:35 
    Structure Examples 2: Cross Section of a Cell 34:56 
    Structure Examples 3: MGDG & DGDG 36:28 
  Membrane Lipids, Part 2 38:20
   Intro 0:00 
   Sphingolipids 0:11 
    Sphingolipid Overview 0:12 
    Sphingosine Structure 1:42 
    Ceramide 3:56 
    Subclasses of Sphingolipids Overview 6:00 
   Subclasses of Sphingolipids: Sphingomyelins 7:53 
    Sphingomyelins 7:54 
   Subclasses of Sphingolipids: Glycosphingolipid 12:47 
    Glycosphingolipid Overview 12:48 
    Cerebrosides & Globosides Overview 14:33 
    Example: Cerebrosides 15:43 
    Example: Globosides 17:14 
   Subclasses of Sphingolipids: Gangliosides 19:07 
    Gangliosides 19:08 
    Medical Application: Tay-Sachs Disease 23:34 
   Sterols 30:45 
    Sterols: Basic Structure 30:46 
    Important Example: Cholesterol 32:01 
    Structures Example 34:13 
  The Biologically Active Lipids 48:36
   Intro 0:00 
   The Biologically Active Lipids 0:44 
    Phosphatidyl Inositol Structure 0:45 
    Phosphatidyl Inositol Reaction 3:24 
    Image Example 12:49 
    Eicosanoids 14:12 
    Arachidonic Acid & Membrane Lipid Containing Arachidonic Acid 18:41 
   Three Classes of Eicosanoids 20:42 
    Overall Structures 21:38 
    Prostagladins 22:56 
    Thromboxane 27:19 
    Leukotrienes 30:19 
   More On The Biologically Active Lipids 33:34 
    Steroid Hormones 33:35 
    Fat Soluble Vitamins 38:25 
    Vitamin D₃ 40:40 
    Vitamin A 43:17 
    Vitamin E 45:12 
    Vitamin K 47:17 
VIII. Energy & Biological Systems (Bioenergetics)
  Thermodynamics, Free Energy & Equilibrium 45:51
   Intro 0:00 
   Thermodynamics, Free Energy and Equilibrium 1:03 
    Reaction: Glucose + Pi → Glucose 6-Phosphate 1:50 
    Thermodynamics & Spontaneous Processes 3:31 
    In Going From Reactants → Product, a Reaction Wants to Release Heat 6:30 
    A Reaction Wants to Become More Disordered 9:10 
    ∆H < 0 10:30 
    ∆H > 0 10:57 
    ∆S > 0 11:23 
    ∆S <0 11:56 
    ∆G = ∆H - T∆S at Constant Pressure 12:15 
    Gibbs Free Energy 15:00 
    ∆G < 0 16:49 
    ∆G > 0 17:07 
    Reference Frame For Thermodynamics Measurements 17:57 
    More On BioChemistry Standard 22:36 
    Spontaneity 25:36 
    Keq 31:45 
    Example: Glucose + Pi → Glucose 6-Phosphate 34:14 
   Example Problem 1 40:25 
    Question 40:26 
    Solution 41:12 
  More on Thermodynamics & Free Energy 37:06
   Intro 0:00 
   More on Thermodynamics & Free Energy 0:16 
    Calculating ∆G Under Standard Conditions 0:17 
    Calculating ∆G Under Physiological Conditions 2:05 
    ∆G < 0 5:39 
    ∆G = 0 7:03 
    Reaction Moving Forward Spontaneously 8:00 
    ∆G & The Maximum Theoretical Amount of Free Energy Available 10:36 
    Example Problem 1 13:11 
    Reactions That Have Species in Common 17:48 
    Example Problem 2: Part 1 20:10 
    Example Problem 2: Part 2- Enzyme Hexokinase & Coupling 25:08 
    Example Problem 2: Part 3 30:34 
    Recap 34:45 
  ATP & Other High-Energy Compounds 44:32
   Intro 0:00 
   ATP & Other High-Energy Compounds 0:10 
    Endergonic Reaction Coupled With Exergonic Reaction 0:11 
    Major Theme In Metabolism 6:56 
   Why the ∆G°' for ATP Hydrolysis is Large & Negative 12:24 
    ∆G°' for ATP Hydrolysis 12:25 
    Reason 1: Electrostatic Repulsion 14:24 
    Reason 2: Pi & Resonance Forms 15:33 
    Reason 3: Concentrations of ADP & Pi 17:32 
   ATP & Other High-Energy Compounds Cont'd 18:48 
    More On ∆G°' & Hydrolysis 18:49 
    Other Compounds That Have Large Negative ∆G°' of Hydrolysis: Phosphoenol Pyruvate (PEP) 25:14 
    Enzyme Pyruvate Kinase 30:36 
    Another High Energy Molecule: 1,3 Biphosphoglycerate 36:17 
    Another High Energy Molecule: Phophocreatine 39:41 
  Phosphoryl Group Transfers 30:08
   Intro 0:00 
   Phosphoryl Group Transfer 0:27 
    Phosphoryl Group Transfer Overview 0:28 
    Example: Glutamate → Glutamine Part 1 7:11 
    Example: Glutamate → Glutamine Part 2 13:29 
    ATP Not Only Transfers Phosphoryl, But Also Pyrophosphoryl & Adenylyl Groups 17:03 
    Attack At The γ Phosphorous Transfers a Phosphoryl 19:02 
    Attack At The β Phosphorous Gives Pyrophosphoryl 22:44 
  Oxidation-Reduction Reactions 49:46
   Intro 0:00 
   Oxidation-Reduction Reactions 1:32 
    Redox Reactions 1:33 
    Example 1: Mg + Al³⁺ → Mg²⁺ + Al 3:49 
    Reduction Potential Definition 10:47 
    Reduction Potential Example 13:38 
    Organic Example 22:23 
    Review: How To Find The Oxidation States For Carbon 24:15 
   Examples: Oxidation States For Carbon 27:45 
    Example 1: Oxidation States For Carbon 27:46 
    Example 2: Oxidation States For Carbon 28:36 
    Example 3: Oxidation States For Carbon 29:18 
    Example 4: Oxidation States For Carbon 29:44 
    Example 5: Oxidation States For Carbon 30:10 
    Example 6: Oxidation States For Carbon 30:40 
    Example 7: Oxidation States For Carbon 31:20 
    Example 8: Oxidation States For Carbon 32:10 
    Example 9: Oxidation States For Carbon 32:52 
   Oxidation-Reduction Reactions, cont'd 35:22 
    More On Reduction Potential 35:28 
    Lets' Start With ∆G = ∆G°' + RTlnQ 38:29 
    Example: Oxidation Reduction Reactions 41:42 
  More On Oxidation-Reduction Reactions 56:34
   Intro 0:00 
   More On Oxidation-Reduction Reactions 0:10 
    Example 1: What If the Concentrations Are Not Standard? 0:11 
    Alternate Procedure That Uses The 1/2 Reactions Individually 8:57 
    Universal Electron Carriers in Aqueous Medium: NAD+ & NADH 15:12 
    The Others Are… 19:22 
    NAD+ & NADP Coenzymes 20:56 
    FMN & FAD 22:03 
    Nicotinamide Adenine Dinucleotide (Phosphate) 23:03 
    Reduction 1/2 Reactions 36:10 
    Ratio of NAD+ : NADH 36:52 
    Ratio of NADPH : NADP+ 38:02 
    Specialized Roles of NAD+ & NADPH 38:48 
    Oxidoreductase Enzyme Overview 40:26 
    Examples of Oxidoreductase 43:32 
    The Flavin Nucleotides 46:46 
  Example Problems For Bioenergetics 42:12
   Intro 0:00 
   Example 1: Calculate the ∆G°' For The Following Reaction 1:04 
    Example 1: Question 1:05 
    Example 1: Solution 2:20 
   Example 2: Calculate the Keq For the Following 4:20 
    Example 2: Question 4:21 
    Example 2: Solution 5:54 
   Example 3: Calculate the ∆G°' For The Hydrolysis of ATP At 25°C 8:52 
    Example 3: Question 8:53 
    Example 3: Solution 10:30 
    Example 3: Alternate Procedure 13:48 
   Example 4: Problems For Bioenergetics 16:46 
    Example 4: Questions 16:47 
    Example 4: Part A Solution 21:19 
    Example 4: Part B Solution 23:26 
    Example 4: Part C Solution 26:12 
   Example 5: Problems For Bioenergetics 29:27 
    Example 5: Questions 29:35 
    Example 5: Solution - Part 1 32:16 
    Example 5: Solution - Part 2 34:39 
IX. Glycolysis and Gluconeogenesis
  Overview of Glycolysis I 43:32
   Intro 0:00 
   Overview of Glycolysis 0:48 
    Three Primary Paths For Glucose 1:04 
    Preparatory Phase of Glycolysis 4:40 
    Payoff Phase of Glycolysis 6:40 
    Glycolysis Reactions Diagram 7:58 
    Enzymes of Glycolysis 12:41 
   Glycolysis Reactions 16:02 
    Step 1 16:03 
    Step 2 18:03 
    Step 3 18:52 
    Step 4 20:08 
    Step 5 21:42 
    Step 6 22:44 
    Step 7 24:22 
    Step 8 25:11 
    Step 9 26:00 
    Step 10 26:51 
   Overview of Glycolysis Cont. 27:28 
    The Overall Reaction for Glycolysis 27:29 
    Recall The High-Energy Phosphorylated Compounds Discusses In The Bioenergetics Unit 33:10 
    What Happens To The Pyruvate That Is Formed? 37:58 
  Glycolysis II 61:47
   Intro 0:00 
   Glycolysis Step 1: The Phosphorylation of Glucose 0:27 
    Glycolysis Step 1: Reaction 0:28 
    Hexokinase 2:28 
    Glycolysis Step 1: Mechanism-Simple Nucleophilic Substitution 6:34 
   Glycolysis Step 2: Conversion of Glucose 6-Phosphate → Fructose 6-Phosphate 11:33 
    Glycolysis Step 2: Reaction 11:34 
    Glycolysis Step 2: Mechanism, Part 1 14:40 
    Glycolysis Step 2: Mechanism, Part 2 18:16 
    Glycolysis Step 2: Mechanism, Part 3 19:56 
    Glycolysis Step 2: Mechanism, Part 4 (Ring Closing & Dissociation) 21:54 
   Glycolysis Step 3: Conversion of Fructose 6-Phosphate to Fructose 1,6-Biphosphate 24:16 
    Glycolysis Step 3: Reaction 24:17 
    Glycolysis Step 3: Mechanism 26:40 
   Glycolysis Step 4: Cleavage of Fructose 1,6-Biphosphate 31:10 
    Glycolysis Step 4: Reaction 31:11 
    Glycolysis Step 4: Mechanism, Part 1 (Binding & Ring Opening) 35:26 
    Glycolysis Step 4: Mechanism, Part 2 37:40 
    Glycolysis Step 4: Mechanism, Part 3 39:30 
    Glycolysis Step 4: Mechanism, Part 4 44:00 
    Glycolysis Step 4: Mechanism, Part 5 46:34 
    Glycolysis Step 4: Mechanism, Part 6 49:00 
    Glycolysis Step 4: Mechanism, Part 7 50:12 
    Hydrolysis of The Imine 52:33 
   Glycolysis Step 5: Conversion of Dihydroxyaceton Phosphate to Glyceraldehyde 3-Phosphate 55:38 
    Glycolysis Step 5: Reaction 55:39 
   Breakdown and Numbering of Sugar 57:40 
  Glycolysis III 59:17
   Intro 0:00 
   Glycolysis Step 5: Conversion of Dihydroxyaceton Phosphate to Glyceraldehyde 3-Phosphate 0:44 
    Glycolysis Step 5: Mechanism, Part 1 0:45 
    Glycolysis Step 5: Mechanism, Part 2 3:53 
   Glycolysis Step 6: Oxidation of Glyceraldehyde 3-Phosphate to 1,3-Biphosphoglycerate 5:14 
    Glycolysis Step 6: Reaction 5:15 
    Glycolysis Step 6: Mechanism, Part 1 8:52 
    Glycolysis Step 6: Mechanism, Part 2 12:58 
    Glycolysis Step 6: Mechanism, Part 3 14:26 
    Glycolysis Step 6: Mechanism, Part 4 16:23 
   Glycolysis Step 7: Phosphoryl Transfer From 1,3-Biphosphoglycerate to ADP to Form ATP 19:08 
    Glycolysis Step 7: Reaction 19:09 
    Substrate-Level Phosphorylation 23:18 
    Glycolysis Step 7: Mechanism (Nucleophilic Substitution) 26:57 
   Glycolysis Step 8: Conversion of 3-Phosphoglycerate to 2-Phosphoglycerate 28:44 
    Glycolysis Step 8: Reaction 28:45 
    Glycolysis Step 8: Mechanism, Part 1 30:08 
    Glycolysis Step 8: Mechanism, Part 2 32:24 
    Glycolysis Step 8: Mechanism, Part 3 34:02 
    Catalytic Cycle 35:42 
   Glycolysis Step 9: Dehydration of 2-Phosphoglycerate to Phosphoenol Pyruvate 37:20 
    Glycolysis Step 9: Reaction 37:21 
    Glycolysis Step 9: Mechanism, Part 1 40:12 
    Glycolysis Step 9: Mechanism, Part 2 42:01 
    Glycolysis Step 9: Mechanism, Part 3 43:58 
   Glycolysis Step 10: Transfer of a Phosphoryl Group From Phosphoenol Pyruvate To ADP To Form ATP 45:16 
    Glycolysis Step 10: Reaction 45:17 
    Substrate-Level Phosphorylation 48:32 
    Energy Coupling Reaction 51:24 
   Glycolysis Balance Sheet 54:15 
    Glycolysis Balance Sheet 54:16 
    What Happens to The 6 Carbons of Glucose? 56:22 
    What Happens to 2 ADP & 2 Pi? 57:04 
    What Happens to The 4e⁻ ? 57:15 
  Glycolysis IV 39:47
   Intro 0:00 
   Feeder Pathways 0:42 
    Feeder Pathways Overview 0:43 
    Starch, Glycogen 2:25 
    Lactose 4:38 
    Galactose 4:58 
    Manose 5:22 
    Trehalose 5:45 
    Sucrose 5:56 
    Fructose 6:07 
   Fates of Pyruvate: Aerobic & Anaerobic Conditions 7:39 
    Aerobic Conditions & Pyruvate 7:40 
    Anaerobic Fates of Pyruvate 11:18 
   Fates of Pyruvate: Lactate Acid Fermentation 14:10 
    Lactate Acid Fermentation 14:11 
   Fates of Pyruvate: Ethanol Fermentation 19:01 
    Ethanol Fermentation Reaction 19:02 
    TPP: Thiamine Pyrophosphate (Functions and Structure) 23:10 
    Ethanol Fermentation Mechanism, Part 1 27:53 
    Ethanol Fermentation Mechanism, Part 2 29:06 
    Ethanol Fermentation Mechanism, Part 3 31:15 
    Ethanol Fermentation Mechanism, Part 4 32:44 
    Ethanol Fermentation Mechanism, Part 5 34:33 
    Ethanol Fermentation Mechanism, Part 6 35:48 
  Gluconeogenesis I 41:34
   Intro 0:00 
   Gluconeogenesis, Part 1 1:02 
    Gluconeogenesis Overview 1:03 
    3 Glycolytic Reactions That Are Irreversible Under Physiological Conditions 2:29 
    Gluconeogenesis Reactions Overview 6:17 
    Reaction: Pyruvate to Oxaloacetate 11:07 
    Reaction: Oxaloacetate to Phosphoenolpyruvate (PEP) 13:29 
    First Pathway That Pyruvate Can Take to Become Phosphoenolpyruvate 15:24 
    Second Pathway That Pyruvate Can Take to Become Phosphoenolpyruvate 21:00 
    Transportation of Pyruvate From The Cytosol to The Mitochondria 24:15 
    Transportation Mechanism, Part 1 26:41 
    Transportation Mechanism, Part 2 30:43 
    Transportation Mechanism, Part 3 34:04 
    Transportation Mechanism, Part 4 38:14 
  Gluconeogenesis II 34:18
   Intro 0:00 
   Oxaloacetate → Phosphoenolpyruvate (PEP) 0:35 
    Mitochondrial Membrane Does Not Have a Transporter for Oxaloactate 0:36 
    Reaction: Oxaloacetate to Phosphoenolpyruvate (PEP) 3:36 
    Mechanism: Oxaloacetate to Phosphoenolpyruvate (PEP) 4:48 
    Overall Reaction: Pyruvate to Phosphoenolpyruvate 7:01 
    Recall The Two Pathways That Pyruvate Can Take to Become Phosphoenolpyruvate 10:16 
    NADH in Gluconeogenesis 12:29 
   Second Pathway: Lactate → Pyruvate 18:22 
    Cytosolic PEP Carboxykinase, Mitochondrial PEP Carboxykinase, & Isozymes 18:23 
    2nd Bypass Reaction 23:04 
    3rd Bypass Reaction 24:01 
    Overall Process 25:17 
   Other Feeder Pathways For Gluconeogenesis 26:35 
    Carbon Intermediates of The Citric Acid Cycle 26:36 
    Amino Acids & The Gluconeogenic Pathway 29:45 
    Glycolysis & Gluconeogenesis Are Reciprocally Regulated 32:00 
  The Pentose Phosphate Pathway 42:52
   Intro 0:00 
   The Pentose Phosphate Pathway Overview 0:17 
    The Major Fate of Glucose-6-Phosphate 0:18 
    The Pentose Phosphate Pathway (PPP) Overview 1:00 
   Oxidative Phase of The Pentose Phosphate Pathway 4:33 
    Oxidative Phase of The Pentose Phosphate Pathway: Reaction Overview 4:34 
    Ribose-5-Phosphate: Glutathione & Reductive Biosynthesis 9:02 
    Glucose-6-Phosphate to 6-Phosphogluconate 12:48 
    6-Phosphogluconate to Ribulose-5-Phosphate 15:39 
    Ribulose-5-Phosphate to Ribose-5-Phosphate 17:05 
   Non-Oxidative Phase of The Pentose Phosphate Pathway 19:55 
    Non-Oxidative Phase of The Pentose Phosphate Pathway: Overview 19:56 
    General Transketolase Reaction 29:03 
    Transaldolase Reaction 35:10 
    Final Transketolase Reaction 39:10 
X. The Citric Acid Cycle (Krebs Cycle)
  Citric Acid Cycle I 36:10
   Intro 0:00 
   Stages of Cellular Respiration 0:23 
    Stages of Cellular Respiration 0:24 
   From Pyruvate to Acetyl-CoA 6:56 
    From Pyruvate to Acetyl-CoA: Pyruvate Dehydrogenase Complex 6:57 
    Overall Reaction 8:42 
    Oxidative Decarboxylation 11:54 
    Pyruvate Dehydrogenase (PDH) & Enzymes 15:30 
    Pyruvate Dehydrogenase (PDH) Requires 5 Coenzymes 17:15 
    Molecule of CoEnzyme A 18:52 
    Thioesters 20:56 
    Lipoic Acid 22:31 
    Lipoate Is Attached To a Lysine Residue On E₂ 24:42 
    Pyruvate Dehydrogenase Complex: Reactions 26:36 
    E1: Reaction 1 & 2 30:38 
    E2: Reaction 3 31:58 
    E3: Reaction 4 & 5 32:44 
    Substrate Channeling 34:17 
  Citric Acid Cycle II 49:20
   Intro 0:00 
   Citric Acid Cycle Reactions Overview 0:26 
    Citric Acid Cycle Reactions Overview: Part 1 0:27 
    Citric Acid Cycle Reactions Overview: Part 2 7:03 
    Things to Note 10:58 
   Citric Acid Cycle Reactions & Mechanism 13:57 
    Reaction 1: Formation of Citrate 13:58 
    Reaction 1: Mechanism 19:01 
    Reaction 2: Citrate to Cis Aconistate to Isocitrate 28:50 
    Reaction 3: Isocitrate to α-Ketoglutarate 32:35 
    Reaction 3: Two Isocitrate Dehydrogenase Enzymes 36:24 
    Reaction 3: Mechanism 37:33 
    Reaction 4: Oxidation of α-Ketoglutarate to Succinyl-CoA 41:38 
    Reaction 4: Notes 46:34 
  Citric Acid Cycle III 44:11
   Intro 0:00 
   Citric Acid Cycle Reactions & Mechanism 0:21 
    Reaction 5: Succinyl-CoA to Succinate 0:24 
    Reaction 5: Reaction Sequence 2:35 
    Reaction 6: Oxidation of Succinate to Fumarate 8:28 
    Reaction 7: Fumarate to Malate 10:17 
    Reaction 8: Oxidation of L-Malate to Oxaloacetate 14:15 
   More On The Citric Acid Cycle 17:17 
    Energy from Oxidation 17:18 
    How Can We Transfer This NADH Into the Mitochondria 27:10 
    Citric Cycle is Amphibolic - Works In Both Anabolic & Catabolic Pathways 32:06 
    Biosynthetic Processes 34:29 
    Anaplerotic Reactions Overview 37:26 
    Anaplerotic: Reaction 1 41:42 
XI. Catabolism of Fatty Acids
  Fatty Acid Catabolism I 48:11
   Intro 0:00 
   Introduction to Fatty Acid Catabolism 0:21 
    Introduction to Fatty Acid Catabolism 0:22 
   Vertebrate Cells Obtain Fatty Acids for Catabolism From 3 Sources 2:16 
    Diet: Part 1 4:00 
    Diet: Part 2 5:35 
    Diet: Part 3 6:20 
    Diet: Part 4 6:47 
    Diet: Part 5 10:18 
    Diet: Part 6 10:54 
    Diet: Part 7 12:04 
    Diet: Part 8 12:26 
    Fats Stored in Adipocytes Overview 13:54 
    Fats Stored in Adipocytes (Fat Cells): Part 1 16:13 
    Fats Stored in Adipocytes (Fat Cells): Part 2 17:16 
    Fats Stored in Adipocytes (Fat Cells): Part 3 19:42 
    Fats Stored in Adipocytes (Fat Cells): Part 4 20:52 
    Fats Stored in Adipocytes (Fat Cells): Part 5 22:56 
   Mobilization of TAGs Stored in Fat Cells 24:35 
   Fatty Acid Oxidation 28:29 
    Fatty Acid Oxidation 28:48 
    3 Reactions of the Carnitine Shuttle 30:42 
    Carnitine Shuttle & The Mitochondrial Matrix 36:25 
    CAT I 43:58 
    Carnitine Shuttle is the Rate-Limiting Steps 46:24 
  Fatty Acid Catabolism II 45:58
   Intro 0:00 
   Fatty Acid Catabolism 0:15 
    Fatty Acid Oxidation Takes Place in 3 Stages 0:16 
   β-Oxidation 2:05 
    β-Oxidation Overview 2:06 
    Reaction 1 4:20 
    Reaction 2 7:35 
    Reaction 3 8:52 
    Reaction 4 10:16 
    β-Oxidation Reactions Discussion 11:34 
   Notes On β-Oxidation 15:14 
    Double Bond After The First Reaction 15:15 
    Reaction 1 is Catalyzed by 3 Isozymes of Acyl-CoA Dehydrogenase 16:04 
    Reaction 2 & The Addition of H₂O 18:38 
    After Reaction 4 19:24 
    Production of ATP 20:04 
   β-Oxidation of Unsaturated Fatty Acid 21:25 
    β-Oxidation of Unsaturated Fatty Acid 22:36 
   β-Oxidation of Mono-Unsaturates 24:49 
    β-Oxidation of Mono-Unsaturates: Reaction 1 24:50 
    β-Oxidation of Mono-Unsaturates: Reaction 2 28:43 
    β-Oxidation of Mono-Unsaturates: Reaction 3 30:50 
    β-Oxidation of Mono-Unsaturates: Reaction 4 31:06 
   β-Oxidation of Polyunsaturates 32:29 
    β-Oxidation of Polyunsaturates: Part 1 32:30 
    β-Oxidation of Polyunsaturates: Part 2 37:08 
    β-Oxidation of Polyunsaturates: Part 3 40:25 
  Fatty Acid Catabolism III 33:18
   Intro 0:00 
   Fatty Acid Catabolism 0:43 
    Oxidation of Fatty Acids With an Odd Number of Carbons 0:44 
    β-oxidation in the Mitochondrion & Two Other Pathways 9:08 
    ω-oxidation 10:37 
    α-oxidation 17:22 
   Ketone Bodies 19:08 
    Two Fates of Acetyl-CoA Formed by β-Oxidation Overview 19:09 
    Ketone Bodies: Acetone 20:42 
    Ketone Bodies: Acetoacetate 20:57 
    Ketone Bodies: D-β-hydroxybutyrate 21:25 
    Two Fates of Acetyl-CoA Formed by β-Oxidation: Part 1 22:05 
    Two Fates of Acetyl-CoA Formed by β-Oxidation: Part 2 26:59 
    Two Fates of Acetyl-CoA Formed by β-Oxidation: Part 3 30:52 
XII. Catabolism of Amino Acids and the Urea Cycle
  Overview & The Aminotransferase Reaction 40:59
   Intro 0:00 
   Overview of The Aminotransferase Reaction 0:25 
    Overview of The Aminotransferase Reaction 0:26 
    The Aminotransferase Reaction: Process 1 3:06 
    The Aminotransferase Reaction: Process 2 6:46 
    Alanine From Muscle Tissue 10:54 
    Bigger Picture of the Aminotransferase Reaction 14:52 
    Looking Closely at Process 1 19:04 
    Pyridoxal Phosphate (PLP) 24:32 
    Pyridoxamine Phosphate 25:29 
    Pyridoxine (B6) 26:38 
    The Function of PLP 27:12 
    Mechanism Examples 28:46 
    Reverse Reaction: Glutamate to α-Ketoglutarate 35:34 
  Glutamine & Alanine: The Urea Cycle I 39:18
   Intro 0:00 
   Glutamine & Alanine: The Urea Cycle I 0:45 
    Excess Ammonia, Glutamate, and Glutamine 0:46 
    Glucose-Alanine Cycle 9:54 
    Introduction to the Urea Cycle 20:56 
    The Urea Cycle: Production of the Carbamoyl Phosphate 22:59 
    The Urea Cycle: Reaction & Mechanism Involving the Carbamoyl Phosphate Synthetase 33:36 
  Glutamine & Alanine: The Urea Cycle II 36:21
   Intro 0:00 
   Glutamine & Alanine: The Urea Cycle II 0:14 
    The Urea Cycle Overview 0:34 
    Reaction 1: Ornithine → Citrulline 7:30 
    Reaction 2: Citrulline → Citrullyl-AMP 11:15 
    Reaction 2': Citrullyl-AMP → Argininosuccinate 15:25 
    Reaction 3: Argininosuccinate → Arginine 20:42 
    Reaction 4: Arginine → Orthinine 24:00 
    Links Between the Citric Acid Cycle & the Urea Cycle 27:47 
    Aspartate-argininosuccinate Shunt 32:36 
  Amino Acid Catabolism 47:58
   Intro 0:00 
   Amino Acid Catabolism 0:10 
    Common Amino Acids and 6 Major Products 0:11 
    Ketogenic Amino Acid 1:52 
    Glucogenic Amino Acid 2:51 
    Amino Acid Catabolism Diagram 4:18 
    Cofactors That Play a Role in Amino Acid Catabolism 7:00 
    Biotin 8:42 
    Tetrahydrofolate 10:44 
    S-Adenosylmethionine (AdoMet) 12:46 
    Tetrahydrobiopterin 13:53 
    S-Adenosylmethionine & Tetrahydrobiopterin Molecules 14:41 
   Catabolism of Phenylalanine 18:30 
    Reaction 1: Phenylalanine to Tyrosine 18:31 
    Reaction 2: Tyrosine to p-Hydroxyphenylpyruvate 21:36 
    Reaction 3: p-Hydroxyphenylpyruvate to Homogentisate 23:50 
    Reaction 4: Homogentisate to Maleylacetoacetate 25:42 
    Reaction 5: Maleylacetoacetate to Fumarylacetoacetate 28:20 
    Reaction 6: Fumarylacetoacetate to Fumarate & Succinyl-CoA 29:51 
    Reaction 7: Fate of Fumarate & Succinyl-CoA 31:14 
   Phenylalanine Hydroxylase 33:33 
    The Phenylalanine Hydroxylase Reaction 33:34 
    Mixed-Function Oxidases 40:26 
    When Phenylalanine Hydoxylase is Defective: Phenylketonuria (PKU) 44:13 
XIII. Oxidative Phosphorylation and ATP Synthesis
  Oxidative Phosphorylation I 41:11
   Intro 0:00 
   Oxidative Phosphorylation 0:54 
    Oxidative Phosphorylation Overview 0:55 
    Mitochondrial Electron Transport Chain Diagram 7:15 
    Enzyme Complex I of the Electron Transport Chain 12:27 
    Enzyme Complex II of the Electron Transport Chain 14:02 
    Enzyme Complex III of the Electron Transport Chain 14:34 
    Enzyme Complex IV of the Electron Transport Chain 15:30 
    Complexes Diagram 16:25 
   Complex I 18:25 
    Complex I Overview 18:26 
    What is Ubiquinone or Coenzyme Q? 20:02 
    Coenzyme Q Transformation 22:37 
    Complex I Diagram 24:47 
    Fe-S Proteins 26:42 
    Transfer of H⁺ 29:42 
   Complex II 31:06 
    Succinate Dehydrogenase 31:07 
    Complex II Diagram & Process 32:54 
    Other Substrates Pass Their e⁻ to Q: Glycerol 3-Phosphate 37:31 
    Other Substrates Pass Their e⁻ to Q: Fatty Acyl-CoA 39:02 
  Oxidative Phosphorylation II 36:27
   Intro 0:00 
   Complex III 0:19 
    Complex III Overview 0:20 
    Complex III: Step 1 1:56 
    Complex III: Step 2 6:14 
   Complex IV 8:42 
    Complex IV: Cytochrome Oxidase 8:43 
   Oxidative Phosphorylation, cont'd 17:18 
    Oxidative Phosphorylation: Summary 17:19 
    Equation 1 19:13 
    How Exergonic is the Reaction? 21:03 
    Potential Energy Represented by Transported H⁺ 27:24 
    Free Energy Change for the Production of an Electrochemical Gradient Via an Ion Pump 28:48 
    Free Energy Change in Active Mitochondria 32:02 

Hello and welcome to Educator.com and welcome to the first lesson of Biochemistry.0000

Biochemistry is absolutely an extraordinary, extraordinary class.0006

There is a lot of information and all of the information is absolutely exciting.0012

Before we actually jump into the biochemistry with proteins and lipids and carbohydrates and metabolism, what we want to do is do just a little bit of a general chemistry review all of the things that are going to be very, very important.0018

Now, you've seen most of these things before but it may have been a while since you've actually worked with them.0031

For the first couple of lessons, we're just going to do a nice chemistry review just to get everything going again and then we'll jump into the biochemistry.0035

Let's get started and welcome again.0043

OK. We're going to start off by discussing aqueous solutions and the notion of concentrations.0048

An aqueous solution, the reason we're discussing this is because the body of chemistry that takes place in the body, takes place in an aqueous solution - in water; we're mostly just water.0055

And it's just a whole bunch of molecules that are dissolved in that water running into each other and doing the things that they do.0070

Because this is biochemistry, the chemistry of biological systems, the chemistry of biological molecules, all of the chemistry that you learned in general chemistry regarding aqueous solutions, all of it absolutely applies here.0073

Let's go ahead and start with our definition of solution and we'll work our way forward.0089

Solution is just a solute dissolved in a solvent.0099

There are two things, you have the solvent and then you have the solute, the thing that's actually dissolved in it.0113

For biochemistry, it works out really, really great because the only solvent that we're concerned with is water, thus, aqueous.0119

If you remember from organic chemistry, you're going to have all kinds of solvents.0124

You can have hexane, you can have ethyl acetate, you can have all kinds of things alcohol, but for biochemistry, its water, so it makes our life that much simpler.0130

Some examples of solutions... Let's see what we've got.0140

We have sugar dissolved in water. That is a sugar solution, sugar dissolved in water.0145

The sugar is the solute and sure enough, the water is the solvent.0155

H2O is the solvent.0164

In this particular case, you have a solid, the sugar crystals, and the solvent happens to be a liquid.0167

Now, it doesn't have to be this way.0172

A solute does not have to be a solid and a solvent doesn't have to be a liquid.0174

It just turns out that way most of the time and certainly in our case.0179

Let's see. How about another example? Let's take salt dissolved in water so a salt solution.0185

When a salt is dissolved in water, again, the salt is your solute and H2O is again, the solvent.0194

Oops, not solute. What we have is solvent. OK.0209

Now, as I said before, a solute does not have to be a solid.0218

A solute does not have to be a solid.0223

It just happens to be most of our experience from general chemistry, organic chemistry and just normal day to day stuff.0233

Most solutes happen to be solid because we dissolve them in something.0240

We see the crystals just sort of disappear as the solution is created but it doesn’t have to be that way and in fact we have a daily example of that- carbonated water.0244

Carbonated water or soda is actually just CO2 gas dissolved in water so a solute is a gas.0255

Let me write this out... carbonated water.0265

In this particular case, the CO2 is the solute.0271

It is a gas and H2O is the solvent. H2O is the solvent.0277

Now, in this particular case, in order to make sure that the CO2 actually stays dissolved in the water to create the carbonic acid solution - the carbon dioxide solution, we have to put it under pressure.0284

Which is why when you pop the can, the carbon dioxide escapes and that's the bubbles rising. Ok.0295

Now the solvent itself... the solvent does not have to be a liquid.0304

Does not have to be a liquid...0316

This notion of a solution is actually a very, very broad definition.0321

It's when something is dissolved in another.0325

In other words, when you get what looks like a completely homogeneous thing where you can't see the individual particles of the solute to the solvent, it just looks like one thing.0327

The solvent does not have to be a liquid, but for us, we don't have to worry about that, our solvent is water.0337

OK. So let's see what we've got here. OK.0344

Sugar and salt, they both dissolve in water but do they dissolve the same way?0350

Sugar crystals, salt crystals, when you're a kid, it looks like they behave the same way.0355

It isn't until you actually get to chemistry that you discover that the dissolving process is actually completely different and what is going on inside the solution, the chemistry is entirely different.0359

Let's write that out.0372

Sugar and salt both dissolve the same way, actually, they don't dissolve the same way, they dissolve.0375

Question is "Do they dissolve the same way?" Sorry about that.0394

Both dissolve but do they do so the same way, and the answer is no, they do not.0397

Now, covalent compounds...0415

Covalent compounds are basically compounds that are made of non-metal non-metal bonds sharing of electrons.0420

Covalent compounds...they dissolve.0429

When they do so, we say that they dissolve. The example of that is sugar.0433

Sugar is a covalent compound even though it has some hydroxy groups where some of the Hs can actually be removed.0437

It is actually is considered a covalent compound because you have carbons bonded to other carbons.0444

You have carbons bonded to oxygens. You have oxygens bonded to hydrogens.0448

These are single bonds, single and double bonds.0453

When these dissolve, they just dissolve.0457

Now, salts, or ionic compounds...I'll write them as ionic compounds and of course the word "salt" is a generic term for any ionic compound.0459

I'll put salt in parenthesis.0470

Now, when salts dissolve... When these dissolve, because not all salts dissolve...Do you remember when you were doing solubility product?0473

If you take sodium chloride which is normal table salt, put it in the water, yes it'll dissolve up to a certain point.0483

If you put silver chloride into water, it'll just sink to the bottom.0489

Remember precipitation? Precipitation is salts that don't dissolve in water.0493

Now, salts when they do dissolve, they dissociate.0497

This is very, very important...they dissociate.0505

In other words, dissociation means they separate into individual ions, into individual free ions.0509

This is very, very important.0522

When a covalent compound...you can put it in there and you are not creating an electrically conductive solution.0524

But, when salt dissolves, like sodium chloride, it breaks up into Na+ and Cl- ions floating around.0532

Well, now, this solution actually will conduct electricity because you have positive and negative charges floating around.0539

The chemistry, the behaviour of the solution, is entirely different even though they actually look the same.0545

That is what's important. OK.0550

Let's just take a table sugar. Let's just sort of see what this looks like.0552

C12H22O11 - this is sucrose as a solid.0557

When we dissolve it in the water, what we end up with is C12H22O11 aqueous.0565

This aq tells us that it is dissolved.0573

This is solid, drop it in water, it is dissolved. OK.0576

That is what this aq means.0580

Let me go ahead and write that.0582

aq means dissolved. It means that is surrounded.0591

Each individual molecule of sucrose has separated from the crystal and is now surrounded by a bunch of water molecules which is why you can't see the individual crystals of the water anymore.0594

It is now a sugar solution, not a sugar crystal.0604

OK. Notice. One molecule of sugar produces one molecule of aqueous sugar.0608

1mol of the sugar crystals will produce 1mol of free individual particles.0616

These particles right here... this aqueous. That means these individual sugar molecules are floating around freely as molecules.0622

Nothing has come apart. The carbon hydrogen oxygen bonds have not broken.0630

It's just a whole molecule just floating around freely whereas here, each molecule is arranged in a crystal.0634

Now, let's go ahead and take something like magnesium chloride, a salt.0642

Magnesium chloride, an ionic compound...This is a solid.0647

When I drop this in water, what happens is it dissociates.0650

It completely comes apart into its free ions.0655

It separates into a Mg2+ ion floating around and you have two chloride ions floating around so what happens is, one unit of these...0658

We don't speak about ionic compounds as molecules because this is not really a covalent bond.0668

This is a positive charge and a negative charge that are stuck together.0674

It's a very strong bond but it's not covalent so we don’t talk about it as a molecule.0677

You can say, you can call it a unit.0682

I mean, it's not going to be the end of the world if you call it a molecule but just to let you know.0684

So 1 unit of magnesium chloride produces three particles: one magnesium ion particle and two chloride ion particles.0688

This is very, very important as you'll see in a minute.0695

Let's go ahead and put aq and aq.0701

In general, when you have ions on the right side of the arrow on an equation, the presumption is that they are aqueous, that they're dissolved,0705

unless you are specifically speaking about a gaseous phase, but we're not.0712

Everything is aqueous chemistry for us so we don't need to put the aq but I'll put them here however, in the future, I will not.0715

OK. So notice.0723

One unit of MgCl2, of the magnesium chloride, it produces three free particles.0726

That's it. That's what's going on here. Three free particles floating around in a solution.0741

Floating around in a solution...0747

Covalent compounds dissolve salts. When they do dissolve they're actually dissociating.0753

OK. Now let's talk about the notion of concentration.0760

If I take 1g of salt and I drop it into 100mL of water versus if I take 20g of salt and I drop it into 100mL of water, clearly, there is going to be a hell of a lot more salt.0765

The concentration of the solution is going to be larger in the second one. There's more salt in it.0779

The volume of the solution is the same. It still stays 100mL but I have 1g and 120g in the other.0785

I need a numerical method for differentiating between the two.0791

We call that concentration.0796

Concentration, there's a whole bunch of ways to express concentration. We're going to be concerned with two of them: molarity and percent by mass.0798

Those are the ones that I'm actually going to introduce and do examples with.0806

However, for the most part, we're really only going to be concerned with molarity - moles of solute per liters of total solution.0808

OK. So expressing concentration... Let me go ahead and put a little line here.0816

Expressing concentration... OK.0829

The first way and the primary way is something called molarity and it is expressed with a capital M.0834

Molarity is defined as the moles of solute divided by the liters of solution.0840

This is the final volume0855

Now, you remember I said that the solute doesn't have to be a solid.0857

If I take a liquid solute like liquid glucose and I drop it into liquid water, well, if I take 10mL of the liquid glucose and put it into 100mL of liquid water, now the total volume of my solution is 110mL.0860

It's not 100mL. So molecules take up volume.0875

This is liters of total solution. This is just moles of solute the things that you add.0880

It's very, very important to keep this thing straight.0885

OK. So let's do an example with molarity.0887

Again, it's all about the examples, all about working with these things using your intuition, everything that you already know from previous classes.0893

We have 7mL of lactic acid dissolved in 130mL of H2O.0902

What is the molarity of the lactic acid?0923

What is the molarity of this lactic acid solution?0928

OK. Well we'll try to write our definition.0941

The molarity is going to equal the moles of lactic acid divided by the liters of solution.0944

That is what we need.0954

We need this number, the moles of lactic acid.0956

We need the liters of solution and then we’re going to do the division and that will give us our concentration in mol/L.0958

That's the unit- mol/L.0965

Let me write that here: moles per liter and it is symbolized with a capital M.0967

I actually prefer to see my entire unit, this M thing is always...hasn't confused me but I like to work with my entire unit. I don't like anything to be hidden but that’s just a personal preference.0975

OK. So, before I do that, this is biochemistry and you know our examples.0986

We're going to try our best to use as many biological molecules as possible.0990

As we do that, I'm going to just draw out the structures of these things just so you get accustomed to seeing them and that's how we develop a sense of familiarity with these biomolecules and they are going to get larger and larger and larger.0996

So, lactic acid looks like this.1009

I'm going to do a straight carbon structure.1012

H and CH3 and we have an OH there...1020

We have three carbons: We have a carbonyl group, this is a carboxylic acid group, this is an alpha-hydroxy acid, actually, and it's an alpha-hydroxy acid.1026

Remember, this carbonyl carbon, this is the carbonyl carbon right here, the one that is attached to the double bond.1034

Let me do this in red.1039

That is the carbonyl carbon. This is called the alpha carbon and this is called the hydroxy group.1041

This is called an alpha hydroxy acid, an alpha hydroxy carboxylic acid - three carbons long.1047

This is lactic acid.1053

This is what develops in your muscles when you start to get sore, when you exercise really, really fast and the body starts to metabolize under anaerobic conditions without oxygen.1055

The by-product is actually lactic acid.1070

That's what you feel when your muscles start to get really, really sore when your exercising really, really fast.1073

OK. Now let me see what were we doing?1079

We want the molarity of this lactic acid solution.1082

OK. I'm going to keep it in red.1084

Let's do moles of... So, we need the moles of lactic acid.1087

Well, we have the milliliter of lactic acid. That is what they give us.1096

We want the moles of lactic acid.1102

How can I go from milliliter to moles?1105

Well, I know that I can go from grams of lactic acid to moles via the molar mass and I can go from milliliters to grams via the density.1107

That's my solution path. From milliliters, I'm going to go to grams and then from grams, I'm going to go to moles.1121

This is density and this is molar mass.1128

OK. Now, I look up the molar mass for lactic acid. I look up the density for lactic acid.1131

If they don't give it to me in the problem, and there is no guarantee that you're going to be given it to the problem, part of the idea is to use your resources whether they be computer resources or book resources to find the things that you need.1138

It is really, really important to be able to do that.1150

There are tables and it's very important that you become adept at utilizing your resources because there is no guarantee in real life.1152

You are just going to be presented with the problem. You have to look up these things.1162

When we look things up, the density of lactic acid is 1.209g/mL and of course I hope you will confirm this for me because I could have read it wrong, myself.1167

And, the molar mass of lactic acid is 90.08g/mol1181

We have 7mL of lactic acid times 1.209g/mL times 1mol happens to be 90.08g.1193

Now, of course, gram cancels gram, milliliter cancels milliliter and what we're left with is 0.0939mol of lactic acid.1211

You know what, I'm going to write this up. I'm just going to note moles of lactic, I'm just going to put lactic.1226

OK. Now we have the number of moles. We have the numerator, we have that.1232

Now we need the liters of solution.1238

Liters of solution...1241

OK. Well, this one is really easy.1244

We had 7mL of lactic acid.1246

We have 130mL of H2O. Both of them are liquid.1251

Remember what we said: liquid solute, liquid solvent, just add the liquids.1256

The total volume of the solution is going to be 137mL which is equivalent to 0.137L.1259

Because again, liters, moles per liter, that's the definition.1271

OK. Now, let's just go ahead and solve the problem.1275

I'll do it on the next page here.1278

So, molarity... The concentration is equal to 0.0939mol of lactic acid divided by 0.137L, 137mL, what you get is 0.686mol/L or 0.686M, molarity. There you go.1281

Either one is absolutely fine.1312

That's our concentration in moles per liter.1314

In this particular situation, in one liter of solution, you have 0.686mol. OK.1316

Now, let's introduce another expression, another way of expressing concentration.1324

This is presented by mass. This is also very, very popular.1329

You'll see this a lot on bottles at the grocery store and stuff.1331

They're expressed in terms of mass.1334

Let me go back to black here.1338

Another way to express concentration is percent by mass. OK.1344

Now, percent by mass, you'll see it this way, %m/m. That's what this symbol is for- percent by mass.1368

The definition is this: It's the mass of the solute.1380

Like any percent, it's always the part over the whole times a hundred. It's a fraction times a hundred.1387

That's what a percent is. A percent is just a fraction that has turned into a number that's a little easier to handle.1392

That's the only reason a percent exists.1398

You actually don't really need that whole multiplying by a hundred.1400

The decimal is just fine. But I guess some people just prefer numbers that are not pure decimals. OK.1404

Mass of solute over total mass of solution...1410

In other words, if I had a solution once I've made it, let's say a sugar solution that weighs 100g and of that 100g, if 5g of sugar are floating around in there, 5g of some solid sugar, what I have is 5 over 100 that is 5% sugar solution.1419

That is what this is. All percentages are just part over the whole, the part over the whole.1436

OK. Let's do an example.1441

This is going to be example 2.1446

What is the percent by mass, the %m/m of the lactic acid solution in the previous example?1451

OK. Well, we need of course the...let's do this in red again.1479

We need the mass of the solute. We need the total mass of the solution. OK.1485

Well, the mass of the lactic acid... so the mass of the lactic acid, that's just going to be the 7mL times its density.1490

We don't want to go all the way to moles so we're going to stop with grams times 1.209g/mL and when we do that we get 8.463g1501

That is the mass of the lactic acid. We have our numerator.1517

Now, total mass...1522

Well, total mass...What is our total mass?1524

Our total mass is the 8.463g of the lactic acid plus the mass of the water.1532

Well, water was 130mL. They gave it to us in volume.1542

Well, the density of water, normally, we just take it as 1g/mL so 130mL of water weighs 130g.1545

So, we have a total of 138.463 and I sure hope that you're confirming my Mathematics. I'm notorious for arithmetic mistakes.1555

OK. Well there we go. We're done.1560

So, the percent by mass of this solution is equal to 8.463g divided by...1574

Oh look at these crazy lines, can't have that. It tends to happen down at the bottom of the page so I think what I am going to do is I'm going to go ahead and move on to the next page because I don't want these crazy lines all over the place.1586

Let's try this again. So, percent by mass is equal to 8.463g of lactic acid divided by 138.463g of solution, and of course whenever we're dealing with the percent, the percent doesn’t have a unit.1598

Gram needs to cancel gram. OK. That's the whole idea...times 100 and when I do the Mathematics, I get 6.11%.1624

Our lactic acid solution was 0.686mol/L. It's 6.11%m/m.1638

That means if I had 100g of this lactic acid solution, 6.11% by mass is made up of lactic acid.1646

That's what this means. So, two ways of expressing concentration, but again, the one that we're going to be concerned with, most of the time, is going to be molarity- moles per liter.1655

And you already know that from chemistry. Most of the time, concentration, molarity is what we use. OK.1664

Well, let’s see what we've got.1673

I'm just going to list a couple of the other ways that concentration is expressed but we’re not going to be doing any samples with them because they're not going to be important for our purposes but I'd like you to know them. Ok1675

I'll go back to black here. OK.1685

Some other ways of expressing concentration had at some point or other, you certainly have heard of these or you will be hearing about them some other time in your career.1693

One of the ways is something called molality, and molality comes up when we talk about colligative properties of boiling point elevation, freezing point depression.1712

We're not going to be concerned with molality.1721

There's something called mole fraction, very, very important and again, a fraction is always the same thing. It's a part of the whole.1724

We did percent by mass. There is also something called percent by volume so it's usually designated as %v/v, I'll just go ahead and write the words out - percent by volume.1732

If I have a 100mL of solution, how many milliliters of that, what volume of that is actually the solute?1748

Percent by mass and percent by volume are two different numbers- they are not the same thing.1755

They might happen to be that same thing coincidentally but they're not the same thing.1760

And, there are other ways, I'm sure.1765

OK. Let's do another example here.1770

See if we can combine our concentration things.1777

A biochemist finds a bottle that a colleague has left at his bench that reads L-Malic 8.66%m/m.1784

OK.1832

What is this solution's molarity?1838

Molarity...1844

A biochemist goes to his colleague's bench and he finds this bottle that says L-Malic acid 8.66%m/m.1849

He knows that it is a Malic acid solution 8.66% by mass.1855

He wants to know what the molarity is, what is this solution’s molarity. OK.1859

In this particular case, we are going to be going from one expression of concentration to another. OK.1864

Malic acid...1869

I told you that we're going to be dealing with biological molecules so let me go ahead and draw the structure of malic acids so you see this one.1872

OK. Let's see. Shall I do it...That's fine, I'll just do it over here. OK.1880

So C,C,C,C, we have hydroxy there and then we have a wedge here.1886

Shall I put...that's fine...I'll go ahead and put the Hs in and then of course we have this.1896

So, this is malic acid. It has four carbons. It is a dicarboxylic acid. There is a carboxylic acid on one end.1902

There is a carboxylic acid on the other end and, it is also an alpha hydroxy acid because on the alpha carbon to one of the ends, there is a hydroxy and this wedge just means that it's actually coming out at us.1910

Biological molecules have a handedness, remember chirality from organic chemistry?1923

There is an L-Malic acid and there is a D-Malic acid.1928

In this particular case, it's L-Malic acid, so if we write it like this, it's actually coming out at us.1933

You will also see just for...you will also see it this way- the regular line structure instead of this.1940

It's up to you how you want to draw, whatever is most comfortable for you.1949

I tend to draw these, I don’t really care for this very much, again, because I like to see my carbons, I like to see what I'm working with.1953

Even after all these years, I still just feel most comfortable doing it this way but of course in the books, you know you're going to see it like this, so you certainly need to be able to recognize it.1960

That is the line structure for malic acid.1974

Malic acid is what gives green apples their tartness.1977

A very, very important biological molecule as you'll find out later in the course when we discuss the citric acid cycle. OK.1981

8.66% by mass means the following.1988

They gave us the number 8.66% - that means this.1997

It means the mass of the malic acid, I'll just call it malic, over the mass of solution times 100 is equal to 8.66.2001

They gave us this. This is the definition, so, I want to start with this.2019

I'm going to work my way back.2023

OK. Now let's ask ourselves what are these that we want and let's do this in red.2027

We want molarity.2030

It's really, really important that you do know what are these that you want.2032

I want molarity, so here's...so, molarity it means I want the moles of malic acid over the total liters of solution.2037

I need those two numbers. I need this and I need this.2043

How can I get that based on the information that I have? OK. Let's take a look at this.2057

The first thing I'm going to do is, I need the moles of malic, liters of solution.2063

The one that's quickest here...what I'm just going to do is I'm going to just...Well, let me write out what the biochemist did.2068

The biochemist measures the volume of the solution.2075

In other words, he just takes it and pours in into a graduated cylinder to see what the volume is in order to get that number.2084

measures the volume of solution to be 17.5mL.2093

In that bottle, he has 17.5mL.2104

He has the volume, he has the denominator, half our problem is done.2108

Now, we just need to find the moles of malic acid. OK.2111

That's the second part.2114

How do we find the moles of malic acid?2116

Well, I need the moles of malic acid and I know that I can get the moles of malic acid from the grams of malic acid, from the mass, from the grams of the malic acid.2119

And my conversion factor is the molar mass. Well molar mass is easy, I just looked it up. OK.2134

So the biochemist measures the mass of the solution. OK.2141

The biochemist measures the mass of the solution.2152

He puts it on the scale and he actually measures it or actually, he takes a glass vile.2156

He takes its mass. He empties the contents into that vile and then he takes that mass.2162

He subtracts the mass of the vile that he knows the mass of and he has the mass of the solution.2167

The biochemist measures the mass of the solution to be 17.75g. OK.2173

Now he has the mass of the solution.2191

Well, we need the mass of the malic acid.2195

Well, we just said earlier, the mass of malic acid divided by the mass of the solution, the total mass which is now 17.75g x 100 = 8.66.2197

We have all these numbers so now we have this equation.2216

We just do a little bit of algebra to find the mass of malic acid and that turns out to be....2218

Actually, let me do it underneath here. Sorry about that.2220

After a little bit of rearranging and a little bit of algebra, the mass of malic acid happens to be 1.537g.2234

There you go. I have 1.537g, that's my mass of malic acid.2246

I have my liters of solution - 17.5mL.2250

Now, I just have to make sure the units are appropriate.2253

Oh, I'm sorry, moles of malic acid.2258

This is my mass of malic acid. I still have to do the conversion to moles.2262

Let's do that.2265

I've got 1.537g of malic times 1mol and again, I'll look it up if the problem doesn't give it to me - 134.09g.2268

This is just basic stoichiometry, then I get 0.0115mol of malic acid.2283

OK. We are done.2293

The molarity equals 0.0115mol of malic acid divided by, and we said we had 17.5mL, right? OK. 17.75mL, the unit has to be liters so move the decimal over 3 times, we get 0.0175L2295

And when we do that, we get 0.066M or 0.66mol/L, my preferred expression for that unit.2329

That's it.2343

We were given a concentration in one expression, percent by mass, we wanted molarity, we wrote down the definition of molarity and we just took a look to see what we needed.2345

We needed moles of solute, we needed liters of solution.2356

Well, the liters of solution is easy, you just measure the volume so that it gives you that.2360

We use the information of percent by mass to recover the mass of the solute, the malic acid, and then from the mass, we use molar mass to get to moles.2363

These are the kinds of things that you want to do. Write it all down.2372

See some sort of a solution path.2376

There is only a handful of definitions. Molarity is moles per liter. Percent by mass is mass of the solute divided by total mass times 100.2379

Everything should come together really, really nicely.2388

OK. Thank you for joining us for our first lesson of biochemistry. We look forward to seeing you again. Take care.2392

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

On the last lesson, we started talking about the amino acids and, we were undergoing the process of classifying those amino acids.0004

We are going to continue with that right now.0011

Let's just jump right on in.0013

OK.0016

We talked about the non-polar R-groups, and we talked about the aromatic R-groups.0017

The next group we are going to do is going to be the positively-charged R-groups.0023

Let's see.0029

Let's go ahead and stick with our blue here.0030

OK.0043

And again, at a pH equal to about 7, which is near the physiological pH.0045

OK.0052

The first one that we have is...let's go ahead and do C, COO-, we have H, we have our amino with a positive charge.0053

And now, CH2, CH2, CH2, CH2, +NH3.0067

OK.0071

This is lysine, and it is Lys, and its single letter designation is a K.0080

Curious isn't it?0093

OK.0094

Our R-group that we're talking about is right here.0097

Lysine has 1,2,3,4 - I always forget how many - 3 or 4.0100

1,2,3,4, methylene groups CH2, and then of course, you notice, you have another amino group and it is protonated.0105

OK.0116

Now, we'll do this one over here.0117

C, COO-, H, +NH3.0123

Now, we have CH2, CH2 - this one is kind of interesting, we have an NH, we have a C, we have +NH2, and we have NH2.0130

It's a bit of a curious one, all kinds of nitrogens in this one.0151

OK.0156

This is called arginine and it is Arg, and its single letter designation is an R.0157

Notice, on the alpha carbon, you've got 2 methylene groups, and then you have a nitrogen with a hydrogen, then you have another carbon; and then you have on this carbon, you have an NH2 group, an amino group, and you have a protonated amino group.0167

That is arginine- positively charged.0185

Here is your positive charge on this R-group.; here is your positive charge on that R-group.0188

OK.0195

Let's see.0197

OK, that's fine; I guess I can do it here.0199

COO-, H, NH3- no changes here - this is an +NH3.0201

Alright.0212

We have CH2, so that's N, that's C, that's N, and that's C, and double bond there.0213

Oops, why do you put a single bond, there is not a triple bond, definitely not a triple bond.0236

That is going to be a double bond.0240

This is going to be an H.0243

This is going to be an H.0245

Let me see if I've forgotten anything.0246

1, 2, 3, oh yes...we have an H here, 4, 3.0248

OK.0253

This is histidine, and we're going to be talking about histidine in just a little bit - very, very important - it is His; it is that.0255

Now, notice, we said positively charged R-groups, but you notice, I haven’t put a positive charge on here.0266

Here is why: that nitrogen, at a pH equal to about 7.0, which is what these are, notice that we've written this particular histidine with this not being protonated.0274

There is no hydrogen attached to it to give it a positive charge, so as it is, this is neutral - OK - pH7.0298

Although we wrote the R-group as neutral, this particular N is actually protonated to a fair amount.0308

It isn't completely protonated.0335

Some of it is deprotonated, but enough of it is protonated to where we can actually group it as in the ones that have a positive charge.0339

You can think of an H being attached there and there being a positive charge.0347

We just haven’t written it that way because it's not completely protonated like these are at pH7.0355

At pH7, some is protonated, most actually, is not, but enough is protonated to where we can consider it as part of this group.0361

OK.0371

As neutral, the N is actually protonated to a fair extent.0372

And again, all of these will make sense when we discuss the acid-base behavior.0381

OK.0387

Now, let's move on to the next group.0388

We have negatively charged R-groups, and I apologize for the tedium of not just putting a picture up and running through these.0391

This is more active; this is important.0402

OK.0407

Alright.0418

Let's do C, COO-, H, +NH3, we have a CH2, we have a C - I can never do that right on the angle, oh, I don’t want to be doing this is - negatively-charged; t7:33 his is a carboxylate.0419

There we go.0453

This is called aspartate, and it is Asp; and single letter designation is D.0454

The group is right here.0461

We have a methylene group, and notice, we have another carboxyl group.0466

This carboxyl group is not this one.0469

It's not attached to the alpha carbon.0472

This is the one that makes it alpha-amino acid, not this.0474

This just happens to be an extra carboxyl group, which is deprotonated at neutral pH.0480

OK, let me see.0488

And, let's do C, COO-, H, +NH3, we've got CH2, we've got CH2.0492

So, we have 2 methylene groups, and then we have our other carboxylate again- this is glutamate.0506

Glutamate, Glu and single letter designation is E; and we have that as our R-group.0516

OK.0529

Let's go to uncharged but polar R-groups- R-group, I should say.0531

OK.0545

We've got C, COO-, H, +NH3, and we have CH2, and we have OH just like glyceraldehyde.0547

This is serine, Ser, S - a very, very, very important amino acid - plays a pivotal role in many, many, many enzymes.0564

Where are we?0580

And again, I encourage you to double check these structures.0581

I am not using a figure and running through it.0584

I'm actually drawing these things out, so please confirm that I have actually drawn them correctly.0588

We are all human; we all make mistakes, so by all means, this is actually a great way for you to actually look at a figure in your book; and make sure that I'm actually drawing it out correctly, and I'm matching the right amino acid, with the right name and the right three-letter symbol and the right single-letter symbol, rather than just taking my word for it.0593

OK.0613

Let's go: C, C uncharged…OK, that's fine0615

Let's do C, COO-, this is H, this is +NH3, and now, we've got a C here, we have a hydroxy, we have that, and we have CH3.0623

OK, this is threonine.0641

So, you notice the C here, you have the OH attached here, there is another C, this is just another CH3 group, threonine, this is Thr and this is a T.0646

Our R-group is right there, and here, our R-group is right there.0657

That is threonine.0664

OK.0666

The third one in this group: COO-, H, +NH3, we have CH2, and we have SH, sulphur- this is cysteine, Cys and C.0668

We’ll talk more about cysteine in just a little bit.0692

Let's do C, COO-, this is an H, this is our+NH3, and then we have our CH2, and then we have C, and double bond O.0696

Now, we have an NH2.0713

So now, instead of a carboxylate here, we have the amide.0716

This is asparagine.0721

It is Asn, and its single letter designation is that.0727

OK.0734

Now, asparagine is the amide of aspartate, the one that we had before in the previous.0740

It's just, instead of a carboxyl right here, we have replaced that O minus, that OH group, where the H is deprotonated.0746

We've replaced it with an NH2.0753

So, whenever you have a carbon double bonded to an oxygen in that same carbon of the carbonyl carbon, is bonded to a nitrogen, that's called an amide or an amide.0755

Again, pronunciation doesn't matter.0763

Now, our last one is COO-, H, +NH3, now, we have a CH2, CH2, C.0767

I should really draw this a little bit... you know what, I’m just going to do it horizontally, How is that...NH2, and this is glutamine.0785

This is glutamine; it is Glm, and it is a Q for a single-letter designation.0802

Asparagine is the amide - I actually prefer to say amide myself - well, let me do it in red.0811

This is your group; these are your groups- it's the R-group.0822

In the case of asparagine and glutamine, they happen to be amide of aspartate, and the amide of glutamate, the 2 other amino acids.0828

OK.0843

There you go.0844

Those are our 20 common amino acids broken down into groups.0845

In this particular case, we happen to choose a group of 5.0851

That's it.0856

Now, let's go on and talk a little bit about these.0858

Some things you should know about the amino acids, there are thousands of things that you should know about the amino acids, but we don't have all the time in world.0861

We'll worry about that as you go on in your biomedical career, but a couple of things that you should keep in mind as we begin our discussion of proteins and amino acids: you should know about the amino acids.0874

OK.0893

Cysteine easily diamerizes, and diamerize means 2 molecules come together to form a single molecule.0896

A dimer made up of 2 pieces diamerizes to form something called interestingly enough, cysteine, Ine.0908

That's why it was different.0922

It's written with an EI instead, so the dimer version is the one with the Ine.0923

Let me go ahead and write out this reaction in structural form.0929

C, COO-, H, +NH3, we have CH2, and we have SH.0935

We are going to add it to another one, so let me just go ahead and write it from left to right this way.0946

COO-, H, CH2 and SH.0952

And, what happens is the following.0958

Now, I'm going to write this vertically.0963

OK.0964

So, let's go ahead and do C, COO-, +NH3, this is going to be CH2, this is going to be S, this is going to be S, this is going to be CH2; this is going to be another alpha carbon - yes, that's right – this is going to be COO-,COO-, this is going to be +NH3, and there is an H, S, S.0965

There you go.1005

OK.1006

I'll go ahead and make the arrows a little bit longer and I'm going to introduce just a little bit of biochemical notation.1009

A little arrow going out, 2H+ + 2 electrons; or if they're coming in this way, 2H+ + 2 electrons.1017

What happens is the following.1029

When these diamerize, what ends up happening is this H plus an electron goes away.1033

This H plus an electron goes away and this S bonds with this S to form a dimer.1038

Here is one of them; here is the other.1047

OK.1052

This right here, this is called the disulfide bond.1053

Profoundly important - we'll be talking about this in a couple of lessons - a disulfide bond.1058

This is how long chains of amino acids, actually they bend and fold, and when a cysteine in one part of the protein chain, and there is a cysteine on another part of the protein chain, when they tend to come together, they end up actually forming a covalent bond, this disulfide bonds.1065

So, what you have is this loop, and there might be other places where it's connected.1084

Disulfide bond is very, very important in the overall structure of a protein.1088

Now, 2H+ + 2 electrons, or you can think about it as 2 hydrogen atoms, either way.1093

This right here.1104

When these are lost, what you have is oxidation- fixed oxidation.1107

You're taking away electrons or another way to think about oxidation is removing hydrogens.1112

By adding electrons, adding hydrogens, you're reducing it.1117

Two cysteine amino acids oxidize to a cysteine dimer.1122

A cysteine dimer reduces to 2 cysteine amino acids.1127

OK.1131

So, our lost, so we call this oxidation.1133

This is oxidation.1137

This is oxidation.1140

OK.1144

Now, tryptophan and tyrosine.1148

I will let you look up their structures- tryptophan and tyrosine.1153

I'll give you a hint: they are in the aromatic group- tryptophan and tyrosine.1157

So again, this is just some basic information on some of the amino acids, just some things to keep in mind that we should know.1167

To some extent phenylalanine, they absorb, because they had aromatic groups, they absorb ultraviolet light strongly.1172

There is a very strong absorption - no, I'm not going to use that symbol for at, I'm just going to go ahead and write AT - at about a wavelength of 280 nanometers.1195

OK. This explains why proteins absorb UV light.1217

It's because of the tryptophan, tyrosine and the phenylalanine content in that particular protein.1231

And of course, this gives us an analytic tool by which to do something, measure things with proteins, analyze proteins.1238

It gives us another thing that we can measure or identify.1246

OK. Now, let's go back to blue.1253

OK, so, there are other amino acids, in fact, a few hundred amino acids floating around on the cell, that are not so common.1258

OK.1277

Some are derived from common amino acids and they do show up in proteins, but are modified after the protein has been synthesized- after protein synthesis.1280

In other words, there are some uncommon amino acids that you do find in proteins, but they're not incorporated while the protein is being built one amino acid at a time, strung together as beads on a string.1311

They use these particular uncommon amino acids.1324

The protein is built using the 20 common amino acids, and then after that, there is a modification.1329

There is something added; there is something subtracted to a particular amino acid residue, to a particular amino acid on the protein chain.1337

I'm just going to list a couple of them, just so you know.1345

And again, you'll find more on your book and I would certainly urge you to take a look at them just structurally.1348

They are very, very interesting1354

So, the first one we're going to take a look at is, well, let me go ahead and draw this structure first, and I'm going to draw it linearly like horizontally.1356

Let's go, C, CO-, H, +NH3, we have CH.1366

OK.1391

I've got CH2, CH2, I've got CH2, I've got a hydroxy there, I've got CH2, and I've got +NH3, 1,2,3,4.1392

OK.1414

This is called 5-hydroxylysine.1416

You'll see it as one word normally, the hydroxylysine or the small L.1425

I tend to separate them, that's just my personal preference.1428

Don't worry, you're not going to get points taken away you shouldn't, unless your teacher specifically says keep it as one word.1432

I know some teachers like that.1438

So, 5-hydroxylysine...this is the 1-carbon, this is the 2-carbon. this is the 3-carbon, this is the 4-carbon, this is the 5-carbon, right?1440

We said the carbonyl carbon is the first carbon. So 5-hydroxy...there is an OH on the 5-hydroxylysine.1450

This is originally lysine, so if I block out this hydroxy, what I’ll have is the normal amino acid lysine.1458

So the protein is synthesized, lysine is put on there, and then after that, the hydroxy is put on there.1463

This is one carbon, this is the alpha carbon.1474

This is the beta carbon, this is the gamma carbon.1478

This is the delta carbon, so it's also called delta-hydroxylysine.1481

And again, number 1 and alpha are not the same.1487

Alpha, beta, gamma, delta, epsilon, and so forth; 1, 2, 3, 4, 5...so, you'll see them both.1491

OK.1499

This is found in collagen, a connective tissue protein- a very, very, very, very important protein.1504

It's what keeps skin soft and supple, elastic.1519

OK.1538

Another one would be something like this.1539

Let's do a C; let me write it a little farther out.1545

This is going to be the final example of an uncommon amino acid.1551

This is the alpha carbon.1559

I always like to designate that, so that I know where it is.1562

CH2, this is CH, this is CO-, there is a CO and O-.1567

So, this is alpha, this is beta, this is gamma, so this is called gamma-carboxyglutamate.1589

Let me go to blue...alpha, beta, gamma.1602

On the gamma, there is another carboxyl group, COO-, and then of course, this rest of the molecule happens to be the normal glutamate.1605

That's it, just a couple of uncommon amino acids that you should know about, nothing too fancy, nothing too difficult.1617

OK.1624

Thank you for joining us here at Educator.com and Biochemistry.1625

We’ll see you next time for discussion of the acid-base properties of amino acids.1630

Take care, bye, bye.1632

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today, we're going to continue our discussion of amino acids, and we're going to talk about the acid-base behavior of amino acids.0004

This is profoundly important.0012

If you understand this, then you'll pretty much understand all of protein behavior; so let's just jump in, and see if we can completely wrap our minds around this thing.0015

OK.0028

Amino acids have two groups that are ionizable.0030

Let me go ahead and do blue here.0035

Amino acids have two groups that are ionizable.0039

OK.0055

What we mean by that is the following: that is two groups that can release and/or accept hydrogen ion depending on what the pH is.0057

In other words, an amino acid is a diprotic weak acid.0088

It has one H that it can give up, and it has another H that it can give up under different pH conditions.0093

Maybe it has already given them up, and maybe this time it will act as a weak base, and will actually take the protons- that is all this means.0100

When we say it has two groups that are ionizable, that means it has two groups that can release or accept the proton depending on what the pH happens to be.0106

In other words, it's a diprotic weak acid, weak base.0115

OK, so let's take a look at alanine.0118

Let's look at an example.0120

Let's look at alanine.0122

OK.0128

And again, the structure of alanine is: we have that, we have that, we have our amino group, and we have CH3.0129

However, I'm going to write it, fully protonated form, COOH.0141

In fact, I'm going to actually draw out the carboxyl groups so we know exactly what we're looking at - COOOH - that's it.0147

This is our first ionizable group - that's one hydrogen that can go away - and here we have our second ionizable group, so every amino acid has at least two ionizable groups.0156

One of this Hs can go away- that's all this means.0166

OK.0171

Let's follow the titration of an acidic solution of alanine with a strong base.0172

This is going to be your standard weak acid strong base titration of a weak acid with a strong base, and see what happens as we raise the pH because that's what we're doing.0200

When we're adding base to a solution, we're raising the pH; and what we're going to be doing is, we're going to be pulling off these hydrogens one at a time as the pH rises.0225

OK.0236

Again, alanine as all amino acids - I'll just write AA - are diprotic weak acids just like carbonic acid H2CO3.0239

H2CO3 releases one hydrogen to become bicarbonate, releases another one to become carbonate.0261

Amino acids, one of the Hs is released from the carboxylic acid group, another H is released from the NH3+ group- that's all that's going on here.0266

Let me see how it is that I actually wrote that weak diprotic acids, so I'm going to write it like this, then I'm going to go back to blue.0279

I'm going to use the three-letter designation, and I'm going to write the carboxyl group, and I’m going to write the NH3+ group.0289

This includes the particular R-group, so this is what we're looking at right here.0297

That's one ionizable group; that's another ionizable group.0303

Let's go ahead and see.0307

Let me go to the next page.0310

Let me redraw this so we have it on the same page: COOH, and we have NH3+- there.0311

OK.0323

Now, the carboxyl group, this one right here, the COOH group, is the stronger acid of the two groups.0325

It's the stronger acidic group of the two, and what that means is, it loses its hydrogen ion first; so it loses its H+ first- that's it - and to become COO-, right?0342

When it loses its hydrogen ion the negative charge stays back, so it becomes COO-.0376

OK.0383

The NH3+ group loses its H+ second to become NH2, so when it loses its hydrogen ion, it's taking its plus charge with it, and it leaves this one uncharged.0385

OK.0407

Now, let's go ahead and follow the titration here, so let me draw this out; and I want to make sure that I leave room at the bottom to draw a structure.0408

This axis is going to be the pH, and this axis is going to be just milliliters of OH-.0421

This is milliliters of OH-, like I'm an adding a sodium hydroxide solution or something; it’s just a volume as I add.0432

Here is what its look like.0439

I'm going to go ahead and go, something like that.0441

Here is what's happening.0451

OK.0453

I'm going to mark off a couple of points.0454

I'm going to mark an X there.0457

I'm going to mark an X there, and I'm going to mark an X there.0458

Now, I'm going to go to red- there is that, there is that, there is that, this is pKa1.0461

Here is what happens: we're going to be starting at a low pH, an acidic solution of alanine.0472

This H is attached, and this H is attached, so what we have is this particular molecule.0479

As I add hydroxide, add hydroxide, add hydroxide - well, you remember a weak acid, all of a sudden, what's going to start happening is that base is going to start pulling off this hydrogen ion from the carboxylic acid part of the amino acid, and it’s going to leave carboxylate, well, remember what we said the pKa was? - the pKa of a given weak acid of a given group, it is the pH at which the acid form and the base form are in equal concentration.0486

So, I’m just going to write a couple of things down, but I'm going to write the reaction underneath, and it will all come together.0522

The pKa1 for alanine is 2.34.0528

Well, so that's the pKa of that group.0534

This one up here, I'll call it pKa2, that is the pKa of the NH3+ group; and it happens to be 9.69 for alanine.0546

Now, I’m going to write something here called PI = 6.01, and here's the reaction that's taking place.0561

I’m going to write: ALA, COOH, NH3+.0571

OK.0586

This is going to be ALA, COO-, NH3+, and then we have ALA, COO-, and we have NH2.0588

This is a +1 charge; this is a 0 charge, and this is a -1 charge.0608

Here's what's happening: as we proceed with the titration, we start with this form, this form right here, OK; the OH is protonated, and the NH3 is protonated, we add base, we add base, we add base, were going to pull off some of this hydrogen.0621

Well, at a pH of 2.34, there is an equal amount of this form, and this form, the form with the H pulled off.0634

This is the pK1- that's this one.0643

Now, that's this right here.0650

This the first buffering region.0653

Remember, where it's flat, that's the buffering region.0657

That is when you have the base form and the acidic form.0658

In other words, the deprotonated form and the protonated form in a concentration that allows you to actually buffer.0662

It resists changes in pH, that's why it looks like this.0671

We are actually adding a hydroxide, but the hydroxide is being eaten up by this H.0675

This H is neutralizing the OH that is added, that is why the pH isn't changing, but at a certain point, this H, all of a sudden, there is no more H for the hydroxide that we add to eat up; so it actually jumps up, the pH jumps up.0680

Now, at this point, it's all in this form: negative charge on the carboxylate, positive charge here, there is a zero total charge.0695

So here, this molecule is positively charged, this is positive 1.0706

At this point it is that, and I'll talk about what the PI means in a minute.0710

I stands for isoelectric point.0714

Isoelectric means there is a zero charge- equal electricity.0715

OK, now, but notice this hydrogen is still attached.0721

I'm going to keep adding hydroxide, now, what’s going to happen is now, the hydroxy is going to pull off this hydrogen; so now, it's going to be converted to this, or this hydrogen is gone, now, this hydrogen is gone.0725

Well, during that, we have our second buffer region.0738

Now, it's the amino group that's acting as a buffer, and again, it's buffers well between about 8.69 to 10.69, here, buffers well from 1.34 to 3.34.0742

That’s the buffering region- one unit above and below the pKa;one unit above or below the pKa.0756

The reaction that's taking place is this: I'd begin with a fully protonated form, I add hydroxide, I pull off the first hydrogen, I pull it all the way off, now, I am here, now, I start pulling off the second hydrogen from the amino group, and I get to a point once I've pulled off everything, now I'm over here, now, my molecule, my amino acid has a negative 1 charge- that's all that's going on when we titrate an amino acid.0764

One hydrogen to give up, second hydrogen to give up; two buffering regions, two ionizable groups.0794

OK.0803

Now, when an amino acid exists as follows: when it exists like this C, COO-, NH3+, H, and R, I'd switched the NH3 and the R, in this particular case, this is not a Fischer projection, I wanted to see, I wanted it to be - well, you know what, actually I don’t need to do that, why don't I just stick with what we've done.0805

We have our R-group down here, and we have our NH3+ like that.0857

When it has a zero charge, when it exists in this form, it's called the zwitterion- that's it that's the name for it.0863

When the carboxyl group has been ionized but the amino group has not been ionized, negative charge, positive charge, the total molecule has a zero charge- it’s a zwitterion.0872

14:45OK.0884

In this form, the COO- group, it can, if it has to, it can accept a proton, it can accept a hydrogen ion, so the amino acid can behave as a base.0885

The amino acid, as a whole, can behave as a base because there is a group that can accept the hydrogen ion- that carboxylate group.0920

Now, the NH3+ group can give up a hydrogen ion, so the amino acid can also behave as an acid if it has to.0933

In other words, it can be both an acid or a base depending on the pH, depending on the conditions at the time, the condition surrounding the amino acid.0967

OK.0976

It is amphoteric.0979

An amphoteric substance is something that can behave as both an acid and a base depending on the environment.0981

Amphoteric is the adjective or ampholyte is the noun, so an amino acid is an ampholyte- it is an amphoteric substance.0987

OK.1003

Now, let me go to red.1004

The pH at which an amino acid is a zwitterion is called is called the PI; it’s called the isoelectric point.1008

When the first hydrogen from the carboxyl group has been completely pulled away, but none of the hydrogens from the amino group had been pulled away, the total charge on the amino acid is 0, -1, +1, they cancel the zero- it is a zwitterion.1037

The pH at which that happens, that's called the PI- the isoelectric point.1055

In the case of amino acids that have two ionizable groups, the PI is just the arithmetic mean between the two pKas- the pKa for the carboxyl group, the pKa1, and the pKa2, which is the pKa for the amino group.1061

You just add them together, divide by two, and you'll get your PI.1078

OK.1083

Now, notice how PI for alanine is 6.01.1087

Now, PIs for most amino acids or most amino acids will be in this range.1106

Now, you understand why we wrote amino acids the way that we did with the COO-, but the NH3+.1124

This is why at pH equal to about 7, we wrote our amino acids as C, H, COO-, NH3+ and R because at normal physiological pH, somewhere in the neighborhood of about 7 to 7.4 amino acids, they exist as zwitterions.1134

So, free amino acids exist in this form under normal physiological conditions.1183

OK.1188

Now, I hope that made sense.1189

You have this amino acid, it has a carboxylic acid group, it has an amino group that's protonated under conditions of low pH.1193

Both of the groups are protonated under conditions of really, really high pH.1203

Both of them are deprotonated somewhere in the middle, which is normal physiological pH.1207

The COOH group is deprotonated, but the NH3+ group is still protonated that carries positive charge to the COO- carries the negative charge.1212

Your total amino acid is zero charge zwitterion it can act as acid or base.1222

it can go both ways depending on what needs to happen in that particular environment.1226

That's what makes amino acids so incredibly powerful.1232

OK.1236

Now, some amino acids have three ionizable groups, and I'll go ahead and list them.1237

They are tyrosine, cysteine, lysine, histidine, arginine, aspartate and glutamate.1254

These amino acids have three ionizable groups because their R-group also contains something that can release or accept a proton.1281

Now, for these amino acids, you have 3 pKas.1290

We call them pK1 for the carboxylic acid group, pK2 for the amino group, and pKR, we can call it pK3, pKR.1294

They say pKR because it happens to be the group that's attached to the R-group.1305

It can be either carboxylate or it can be an amino.1310

OK.1314

Now, let's see what we've got.1316

Now, as always, the COOH that's attached to the alpha carbon, attached to alpha C, always ionizes first; so that doesn't change.1318

The pK1 always refers to the, in other words, you have an amino acid.1347

OK.1361

That particular H is the one that always ionizes first.1363

Now, well, for NH3+ and the particular R-group, it's a toss-up.1366

Sometimes the NH3 will ionize second, and then sometimes the R-group will ionize second; and then the NH3 as the pH is rising, so sometimes this will have a lower pKa than that one, ionizes first, sometimes this R-group will have a lower pKa than this group.1378

It means it ionizes first; it loses a proton first- it's just depends.1397

OK.1404

Sometimes, one or the other will ionize first, will lose its proton first.1410

A titration curve for a triprotic amino acid on that list is going to end up having three plateaus.1424

It's going to look something like this; in general, it's going to look something like this, something like that: pKa1, pK2 or pKR, depending on which one is first, and pK3 and somewhere in here you're going to have your PI.1434

Now, you can't just add them and divide by three in this case.1452

We have to experimentally determine what the isoelectric point is for these, but that's not a big deal.1455

And again, if you look in your book, you will actually see a list- all of the amino acids.1461

It will list their three letter designation, single letter designation.1468

It will give you the molar mass.1471

It will give you pK1, pK2, pKR, and then it will give you the PI and maybe some other information too.1472

OK.1480

Let's go ahead and do an example here.1481

I think this is probably the best way to do it.1483

Let me go ahead and do it on the next page.1485

So, example, we're going to take a look at aspartate.1488

In the case of aspartate, our pK1 is less than our pKR, is less than our pK2; so in this particular case, the R-group, the carboxylic acid ionizes first, releases its hydrogen, then the R-group will release its hydrogen, then the amino group the alpha-amino group will release its hydrogen as we titrate.1495

I'm going to draw out the reactions; I'm not going to do the titration curve.1519

I’m going to write out the reaction- that's what's important.1521

We want to get the structures correct: H, NH3+, we have CH2 ,and we have COOH.1526

OK.1540

We have that one, and it's going to be H3N+.1542

And again, I’m hoping that you're actually confirming all of this because there is a whole bunch of structures going on, so I might miss an H, I might miss a C, I might miss an N.1547

I hope you are confirming this.1557

OK.1560

This is C, this is COO-, and this is CH2, and this is going to be COOH, so this is pK1.1562

OK.1572

This group right here loses first.1573

Now, our second ionization is going to be pKR, so this H is going to go next1576

What we have is NH3+, alpha carbon, COO-, H, we have CH2, and we have COO-.1584

Now, we have our final equilibrium which is going to be pK2, which is going to be the amino group, and we are going to end up with a C, a COO-, an H, a COO-; and then will going to have an NH2 neutral.1597

Notice, it went from plus to neutral, because it gave up an H; that's what's happening her- an H is being lost in each case.1619

In terms of the biochemical, an H+ is leaving - actually you know what I should do it on the upper arrow, not the lower arrow, the lower arrow is the one that is coming in - so, H+ is going away, a second H+ is going away, a second H+ is going away.1631

This is pretty typical biochemical nomenclature.1660

They actually show things coming in and going out of a reaction on the arrows, but again we'll talk a little bit more about that.1663

This is pKR, now, let's do some numbers: pK1 = 1.88, the pKR = 3.65, the pK2 = 9.60, and its isoelectric point happens to be at 2.77.1669

So, at a pH of 2.77, it actually exists in this form- 0 net charge.1694

That's all that is going on here which makes sense because you're looking at 1.88 and 3.65, because each of this contributes a negative; the only positive charge comes from this thing right here, so this PI is going to be lower than you would expect.1707

Notice the PI of alanine was 6.01- this one is a lot lower.1722

OK.1727

Let's do another example.1731

This time we'll do an example where the alpha-amino group actually ionizes before the R-group does1734

Let's do tyrosine, which is actually kind of interesting in the case of tyrosine but...so pK1 is less than pK2 is less than the pK of the R-group, so tyrosine.1739

Let's go ahead and write these equilibriums.1756

We start off with COOH, everything is protonated, we have NH3+, we have CH2, we have our phenol group or benzene, then we have OH, so everything is protonated, everything is good.1760

Now, first hydrogen to go is that top hydrogen, the alpha carboxylic acid, the carboxylic acid attached to the alpha carbon.1780

We have H3N+, C, COO-, this is H, this is CH2, this is that, and we have OH, that's our first, this is pK1.1789

Now, for pK2, this time it is the amino group that ionizes next, so it becomes H2N, neutral C, we have COO-, we have H, we have CH2, we have our benzene group, and then we have our hydroxy attached to the benzene, which is still protonated.1808

That one has not been released yet.1829

And now, of course, we reach our final equilibrium, which is going to be pK of the R-group.1832

Now, the R-group is going to release its hydrogen.1838

We have C, we have COO-, we have H, we have NH2, we have CH2, we have our benzene group, and we have O-.1842

This is the equilibrium that takes place.1858

This H goes first to turn into that, then this H+ leaves to turn into that, then this H leaves to turn into that.1861

Our numbers are: our pK1 is equal to 2.20, and, of course, all of these numbers are available in your book or on the web- wherever.1875

I would encourage you to take a look at a table showing this stuff just to get a sense of what the numbers are for all the amino acids on a single page on a list.1887

It's a great way to get a sense of general behavior because there are just going to be some numbers that are just going to stand out.1895

They are just going to be totally different than all the others, and you're going to take a look and see what amino acid that is, and chances are, that amino acid is going to play a special role when we talk about metabolism later in the course.1901

OK.1913

pK2 = 9.11, and pKR = 10.07 , and PI is equal to 5.66.1915

So, at a pH of about 5.66, the majority of the amino acid exits in a neutral state- that's it.1930

OK.1939

Now, I strongly urge you to do exactly what I've done: take a couple of amino acids at random, and then just write the equilibriums for them, see what the pKa1 is, see what’s the pKa2 is, see what’s the pKR is, and then arrange them, plot the hydrogens according to the order of the pKas, and draw these out.1942

It's an absolutely fantastic way to1, familiarize yourself with the structure of the amino acids and just being able to actively draw them out, and 2, getting a sense in keeping track of which hydrogen is being ionized and where it's being ionized- very, very important.1964

OK.1984

Now, let's talk about a rule of thumb.1986

OK.1993

If you want to know whether a given group - chemical group, not amino acid group - whether a given, I should say, ionizable group is protonated, which is the acid form or deprotonated.1998

In other words, whether it’s actually has its hydrogen ion or it's lost its hydrogen ion, which is called the base form at a given pH.2037

That's often how the problems are going to present themselves.2054

We're going to say there is this particular acid and the pH of the solution is 6.7, which one of the groups is protonated, and which one is not?2057

That's how it's going to be presented, and we will do an example in a minute.2064

Here's how you do it.2068

Here's the rule of thumb: if the pH of the solution is less than the pKa of the group - and again, we're doing this for each individual group - then the group is protonated.2070

In other words, it exits in its acid form, and, of course, the other way around if the pH happens to be bigger than the pKa; and remember, the pKa is a constant.2096

These things exist for a given species for a given ionizable group in that species.2111

The pKas don't change, pHs change.2116

Then, the group is deprotonated.2128

In other words, it exists as the base form.2133

OK.2140

Now, here's the bases for this particular rule of thumb.2142

You can either learn the rule of thumb, memorize it, or you can learn this basis, which I think, is better to know the basis and to know where it comes from, because that way, you can always reason things out.2146

Well, remember the Henderson-Hasslebalch equation?2158

OK.2160

Here's the basis for the rule; let me do this in red: the pH, we said of a solution, is equal to the pKa of the acid plus the logarithm of the concentration of the base form, the unprotonated form, over the concentration of the acid form, the protonated form.2161

Now, let me rewrite that: pH = pKa plus the log of the base concentration over the acid concentration.2193

Well, if the pH is less than the pKa, if this is less than that, which is a constant, that means that this number right here, the log of B over A, is a negative number, because I have to go a certain number subtracted something to get a lower number, then log of B over A is negative - in other words, it's less than zero - so if the log of something is negative, that means that the denominator is bigger than the numerator.2206

In other words, the logarithm of a fraction is negative, the logarithm of the number bigger than one is positive.2256

If the log of B over A is negative, that means B over A is a fraction.2263

If it's a fraction, that means A is greater than B.2268

That means that the denominator is bigger than the numerator, meaning - and don't worry we’ll be doing an example in just a minute - meaning there is more A than B.2272

There is more acid form than base form.2307

There is more protonated form than non-protonated form.2309

That’s all that means.2316

OK.2318

Let's go ahead and finish off with a nice example her, see what we can do.2319

Let's go back to blue.2326

Example: the cysteine solution was prepared and buffered to a pH equal to 8.2.2332

I would like you to describe the degree of protonation for each ionizable group.2361

In other words, I'd like you to tell me does this amino acids exists in what form.2382

What's the total charge on it?2388

Which group is ionized; which group in not ionized?2390

That's what is asking.2393

OK.2394

Well, let's go ahead and take a look at - first of all this is biochemistry, it's chemistry, it's organic chemistry - draw a structure.2396

OK.2404

So, cysteine, let's go ahead and draw it out as, you want to draw out the fully protonated form first and then make your decision, NH3+, this is an H, this is a CH2, and cysteine is a SH.2406

OK.2427

Again, begin by protonating all of them.2429

In other words, that's protonated, that's protonated, that's protonated- we have three ionizable groups.2431

At pH of 8.2, which one is protonated, which one is not?2436

Well, let's see what we've got.2441

We look up the pKs.2443

Well, the pK1 is equal to 1.96; the pKR is equal to 8.18.2446

Notice, in this case, the R-group ionizes before this group does.2456

The pK2 is equal to 10.28.2460

Well, now, we just use our rule of thumb or reason it out.2466

pH is bigger than pK1, right?2476

We said that pH is 8.2, and we said the pK1 was 1.96.2478

Because the pH is bigger than pK1, that implies that the carboxylic acid group exists as a carboxylate group.2485

It's actually been ionized; it has lost its H.2502

OK.2506

The pH which is 8.2, in this particular case, it happens to equal the pKR- that's interesting.2507

This is 8.18, the pH is 8.2, and they are exactly the same.2519

This implies that - how shall I...I'm just going to write the group - CH2, SH, and the CH2, S-, they exist in equal concentrations.2525

In this case, the pH equals the pKa of the R-group.2550

When pH equals the pKa of the R-group, that means the acid form, the protonated form and the base form, the unprotonated form, exist in equal concentrations.2554

So, in this case, they're both like that- it's a little bit of this, a little bit of that, half and half exist in equal concentrations.2563

Now, the pH happens to be less than the pK2.2575

Again, the pH is 8.2, and this is 10.28.2581

Well, this implies that the alpha amino group exists as its protonated form; the pH is less than the pKa, so it has not ripped away this hydrogen; it's still H3N+, so that's it.2587

Our final answer- we have C, COO-, H, NH3+, CH2, and SH - just want to make sure if...yes - and C, COO-, NH3+, CH2, S-, H, so, this is our final answer.2613

The amino acid actually exists as an equal concentration of this thing and this thing.2649

OK.2658

This group is completely ionized; this group, the amino group is not ionized.2660

The SH group- half of it is ionized, half of it is not.2666

That's what’s going on here.2672

OK.2674

I hope that made sense, and this is strictly based on the rule of thumb.2675

Let me write and OK.2678

And again, it's based on comparison of pH and pKa; compare pH and pKa, compare pH and pKa of each R-group- that's all that's going on here.2683

OK.2695

Now, let's say one thing about one of the amino acids, and then we will go ahead and close out this particular lesson.2698

Histidine is special.2710

When I look, I see a pK1 of 1.82; I see a pKR equal to 6.00, and I see a pK2 equal to 9.17.2717

Notice.2737

OK.2742

If you take a look at a list of all of them, this is one of the numbers that will stand out- that one.2743

It is the only amino acid whose pKR, who's pKa of the R-group is close to physiological pH, is close to physio pH found in intracellular and extracellular of fluids- the fluid inside the cell, the fluid outside the cell, physiological pH.2751

This is the only amino acid whose pKR is actually close to the physio pH.2800

OK.2805

It is, therefore, it is therefore, I should say -sorry about that , let me go ahead and erase that – so, it therefore has the potential to provide good buffering capacity under physiological conditions, under physio conditions.2808

That’s it.2868

Histidine is special because its R-group has a pKa of 6.0, which is not that far from the 7.0 or 7.2.2869

As it turns out, it has the potential to actually be a pretty good buffer in that particular range.2881

As it turns out, that's exactly what it is going to do, so keep an eye out for histidine when we start talking about enzyme reactions and when we start talking about metabolism.2889

OK. That takes care of acid base behavior for amino acids.2900

Thank you for joining us here at educator.com and biochemistry.2904

We'll see you next time, bye-bye2907

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today we are going to continue our discussion of amino acids by taking the next step.0004

We are going to put some amino acids together.0009

We are going to be talking about peptides and proteins.0011

Let's get started.0014

OK.0016

Peptides and proteins- it is just a string of different amino acids like beads on a string.0018

That's all it is- each one connected to the next.0025

Let's go; let's write that down.0029

Peptides and proteins are made up of amino acids - and again I'm just going to write AA like that - amino acids strung together- that's it.0035

Now, you’re probably wondering "Peptide, protein - what's the difference?".0053

The truth is there is no difference.0057

There are just a bunch of names that are used for proteins, amino acids that are strung together.0058

In general, if you want, you can think of a peptide as being anything less than about 10,000 atomic mass units, so molar mass of about 10,000 or less, we tend to call it a peptide.0064

A protein is 10,000 or more, and again it is not really this, you can call whatever you like.0080

We're actually going to be using several terms interchangeably.0086

So, just to sort of throw it out there, protein greater than about 10,000 atomic mass units or grams per mole, molar mass.0091

OK.0102

Now, let's go ahead and talk about the formation of a peptide bond- very, very important.0104

Bond is the bond that exists between two amino acids, and this is what holds the chain together.0114

OK....the bond between two amino acids.0124

OK.0139

Let’s go ahead and just take some generic amino acids, and see what we've got.0141

Again, when we write amino acids, I think it is the backbone that's important.0148

Yes, the R-groups are important, of course, when we're discussing them; but when we want to draw it out, it is the backbone that's connected- the peptide bond.0154

It is N, C, C, N, C, C repeating units, so if you’re going to write an amino acid, just start by writing N, C, C, and then fill in the rest.0162

We have N, C, C, I'll go ahead and put H3+ here, I'll go ahead and put the carbonyl there, and I'm going to write it in its fully protonated form just so you see that this is actually a condensation reaction.0171

The elements of water are going to be removed when we put these two amino acids together to form a dipeptide.0186

OK, so the carbonyl carbon is the second carbon.0194

The R-group goes here; the H is there, and then plus, and then, of course, we have, again, N, C, C.0197

This time I'm just going to put H there.0208

Actually, you know what, let me do it this way.0212

Let me put an H here, and let me put an H here.0215

I decided this one, I’m going to not protonate.0220

Don't worry about why, as far as where the Hs go, whether it is 3 Hs, 2 Hs, things like that, that's not what’s important right now.0221

Right now, we just want to be able to get a global sense of how these bonds form.0230

We have an H here.0237

This is the second R-group, and we have the carbonyl, and this one, I'm just going to go ahead and leave as an O-; if you want you can put an OH again.0238

Now, what happens is this.0247

I'll go ahead and do an equilibrium arrow this way and this way, so when water leaves this, the elements of water are right here.0251

Let me do this in red.0262

OK, the elements of water.0268

Basically, it is going to be, this carbon is the electrophile, and this nitrogen is going to be the nucleophile.0272

Remember the nucleophile is the one that actually has the electrons; it has the negative charge.0289

Electrophile is the one that carries the positive charge, so this thing is actually is going to attack this thing.0294

We’re not going to worry about mechanism right now, but that’s what happens.0299

What you’re going to connect is - and I’ll to do this one in blue - you’re connecting those two.0302

You’re connecting this carbon of 1 amino acid to the nitrogen of the amino group of the other amino acid.0310

OK.0320

And in this direction, it is condensation, because again, the elements of water are removed, and we're going to go ahead and write out our dipeptide; and again, we're going to go N, C, C - oops, straight lines, we definitely don't need those - N, C, C, N, C, C.0322

Now, let's go ahead and put the H3, let's leave that as plus.0348

Let's go ahead and put an R-group there; let's put the H here.0352

Here's our carbonyl, the N; let's go ahead and leave that one H that was there.0357

This is our R2 group - right? - of the other amino acid.0364

Now, we have this, and we have that.0368

The peptide bond is this thing right here; that is your peptide bond.0370

It is the carbonyl carbon attached to the nitrogen of the amino group of the amino acid to its right.0377

This is your peptide bond.0389

OK.0395

And just to let you know, in this direction, it is hydrolysis, so when we actually add water to a peptide, it actually splits this bond right here, the peptide bond; and it releases 2 free amino acids.0396

So, in this direction, two amino acids’ condensation, in this direction, it is called - got a little too much floating around here so when H2O comes in, OK, when this H2O leaves, this, well, here I will just do this - in this direction, it is called hydrolysis.0415

That's it- simple peptide formation.0440

The elements of water leave, the carbonyl carbon attaches to the nitrogen of the other amino acid, and you get your peptide bond; and it goes on like this, so N, C, C, N, C, C- it is the second carbon.0443

If this were going to attach to another amino acid, it would be this carbon that’s going to be attached.0459

If this one were going to be attached to something to its left, that would be attached.0464

That is all that’s going on here.0469

OK.0471

Under physiological conditions, under physio conditions, interestingly enough, the equilibrium of the previous reaction of peptide bond formation, the equilibrium lies to the left.0476

In other words, it lies to the formation of free amino acids, not the peptide, the left, and the reason that is so is because the hydroxy group is not a good leaving group; and you remember that from organic chemistry, just a straight hydroxy is not exactly a good leaving group.0499

It doesn't just go away to make the reaction move forward and form the peptide bond.0521

The hydroxy must be activated, and if you remember from your first year biology course or those of you who had already perhaps taken molecular biology, the amino ace will transfer RNA, ribosomes, protein synthesis, that is what is active.0537

That is activated with adenosine triphosphate and all that other stuff, so you’re welcome to look that up.0561

I'm not going to go through it here0565

It has to be activated to induce it to leave.0567

We just wanted you to know that in general, under normal physiological conditions, as is, the equilibrium tends to lie to the left, which is why you need it to be catalyzed.0577

OK. Let's do an example here.0588

Example 1: let's build the following tripeptide, so 3 amino acids, let's do Ala, Tyr and Ile, so alanine, tyrosine, isoleucine.0595

OK.0620

Let's go ahead and draw these out.0621

Again, we have N, C, C, we have N, C, C, we have N, C, C.0624

OK. I'm going to go ahead and draw them out individually so that we see- again, this is all great practice.0636

Let's see, alanine was CH3; and please, by all means, confirm that I'm actually writing the correct structures.0644

We all human; we have a bunch of carbons, nitrogens, hydrogens floating around.0652

We're going to be doing lots of structures.0657

The molecules are going to go get bigger and bigger and bigger, so, by all means, please make sure that I'm actually writing the correct structures because I get things wrong.0658

Let's see, we have that carbonyl; I'm going to go ahead and do an OH.0668

I'm going to go ahead and write an NH3+, because I'm writing them as individuals, and we said that tyrosine is the next one; so, tyrosine, CH3, I believe we have this one, the phenyl group with a hydroxy attached, and then we have the carbonyl carbon, and then, of course, we have the isoleucine, this is a plus charge, this is that, here's our carbonyl.0675

I'm going to go ahead and put an OH there, and our R-group, isoleucine - I never remember our isoleucine - this one is H, this is CH2, and I think this is CH3, yes, that is correct.0702

OK. Again, the carbonyl carbon attaches to the nitrogen.0715

The two things that we are going to connect are this and this, and we're going to connect this and this.0720

This is going to go away; this is going to go away.0730

Now, let's go ahead and draw our structure; and again, this time we go N, C, C bond, N – oops, we don't want extra straight lines because we're dealing with lines - N, C, C bond, N, C, C, OK, H3+.0734

I'm going to go ahead and put the carbonyls on first.0759

The carbonyl is on the second carbon, N, C, C.0762

The carbonyl goes on the second carbon, N, C, C, carbonyl on the second carbon, carbonyl on the second carbon.0763

This is the free end, so I'll go ahead and put that one there.0769

This is going to be alanine, so I'll do that here.0774

It is going to be tyrosine, so I'll put the OH there.0778

This one is going to be isoleucine: CH3, H, CH2, CH3.0785

I'm going to go ahead - well, you know what, that's fine - I'll go ahead and put the hydrogens.0796

In a little bit, I am probably going to start leaving off the hydrogen that's on the alpha carbon; it is there.0799

Again, from the organic chemistry, we don't always write all of the hydrogens.0809

That's it.0812

Our peptide bonds are that one - oh, you know what I should do, yes, I'm going to go ahead and put the hydrogens on the nitrogens - that's important, those are important.0813

OK.0830

Let me go back to blue.0831

That's one peptide bond right there; here is another peptide bond right here.0833

Actually, you know what, why don't we consider this whole thing a peptide bond, but that's the actual bond.0839

OK.0844

That's it.0846

This is called, if you want the name of it, basically, you just take the name of the individual acids starting from the left, you drop off the INE, and you add YL to have the "eel" sound like, remember, carbonyl, alkyl.0848

This is actually alanyl, tyrosyl, isoleucine as one word; but it is not the end of the world if you want to separate them.0866

OK.0884

This is the structure of this tripeptide, and notice, I actually use all my carbons.0887

When I do my structures, I personally like to write out every single atom; I like to write out my carbons.0893

I've never really cared for line structures myself; obviously, you want to be able to understand them, but I like to see every single carbon that I'm dealing with.0899

That is just a personal taste.0907

You have your personal taste, and by all means, don't feel compelled just because everybody else is, let's say, using line structures, that you have to use line structures, unless you have a professor that's going to take points off.0908

Understanding is more important than aesthetics at this point.0920

Later on, maybe, you can get into, maybe you’re starting to draw your line structures; but again, we want to be able to understand.0924

To this day, I actually prefer to write everything out as a straight line like this, but I'm going to go ahead and show you a couple of the other representations here, just so you know.0933

I'm going to give you the shape structure, and the shape structure is the same except it actually takes into account the angles, so it is going to look like this.0944

If you do a shape structure, it is going to be N, C, C, alternating N, C, C just like when you did alkyl change, C ,C, C, C, that little zigzag pattern, N, C, C, N, C, C, and then N, C, C.0956

Again, you just sort of fill everything in.0974

This is a plus; the carbonyl goes on the second carbon, N, C, C.0976

The carbonyl goes on the second carbonyl, N, C, C.0980

Carbonyl goes on the second carbon, something like that; and, of course, you can put your R-groups.0982

So, alanine is going to be CH3; tyrosine, it is going to be CH2.0988

Our benzene and our hydroxy and here, we have the CH, CH3, and then we have CH2, CH3, that's our isoleucine.0994

And again, let me go ahead and put the hydrogens on the nitrogen, that's important, but notice that I've left the hydrogens off the alpha carbon, so this, sort of, is another way of representing it.1010

I think it is a really nice way of doing it.1021

Again N, C, C, N, C, C, N, C, C, but notice how the carbonyls, now, they alternate, one is down one is up.1023

That's it.1030

The line structure would look like this.1031

We will go ahead and put an N, and then we’ll do that, C, C, N, C, C, then N, C, C.1041

So again, you have something that looks like this.1055

To this day, it still confuses me; it makes me crazy.1058

I really just, really like to see my carbons.1061

Carbonyl is on the second carbon, N, C, C.1064

Carbonyl is on the second carbon; that one is taken cared off.1067

Here we have CH3; here we have the CH2, the benzene, the hydroxy, and then we have the CH, we have CH3, CH2 and CH3.1070

That's it.1088

OK.1089

When you have a peptide like this- let me go ahead and put my hydrogens on, it's probably very, very important; and again, let me go ahead and do my peptide bond.1091

My peptide bond is that one right there, the carbonyl carbon and the nitrogen.1102

Where is the next one?1107

The carbonyl carbon and the nitrogen, carbonyl carbon, nitrogen, carbonyl carbon, nitrogen- that's your peptide bond, very, very important.1110

OK.1120

Traditionally, what we do- let me go ahead and put a charge on here; this is a plus charge, better not forget that, there is a minus charge.1122

OK.1129

When we write our proteins, we write them from left to right, and on the left hand side, we put the free amino group, this in red; on the right hand side, we keep the free carboxyl group.1132

This right here, this is called the N-terminal amino acid.1143

In this particular group, the N-terminal amino acid is the alanine, also called the N-terminus.1154

It is the amino terminal group, the amino terminus, so we call it the N-terminus; and this is the C-terminal amino acid or the C-terminus, C standing for carboxyl.1164

That's it.1187

We always put the amino on the left and the carboxyl on the right.1188

We read from left to right.1191

This is the alanyl, tyrosyl, isoleucine.1193

That's it.1198

OK.1200

Let's see what else we can do here.1203

All right, OK.1207

Peptide bonds are very stable once they form, having average half-lives of about 7 years.1212

So, 7 years later, you'll still have half the proteins that were made 7 years ago.1242

That's all that means- under physiological conditions, 7 years under physio conditions.1248

OK.1261

Let's go ahead and redraw our peptide here.1263

I'm going to do it as blue.1268

We've got N, C, C, N, C, C, N, C, C, 3+, carbonyl, N, C, C, carbonyl, N, C, C, carbonyl, O-.1271

We had our alanine group, and we had our tyrosine group, and we had our isoleucine group - oops, not CH2, it is CH - and then we had a CH3 out there, we had a CH2 here, and we had a CH3 there.1289

OK.1316

Now, a peptide - notice - is just like an amino acid; I mean, you've got an amino end, you've got a carboxyl end; and in this particular case, you happen to have some groups in between that also have ionizable groups.1317

That's it.1328

It is just going to behave like a long amino acid.1329

It is going to have a pKa.1332

This group is going to have a pKa; this group is going to have a pKa, and, of course, in this particular case, because this is an ionizable group, it is going to have a pKa.1334

This particular tripeptide is going to have 3 pKas.1344

The titration curve for this one is going to be exactly what you think; it is going to have 3 plateaus, 1 for each pKa.1347

That's it.1356

There is nothing strange happening here.1359

It just behaves like a really, really long amino acid.1362

Ionizable groups behave the same way they would any others.1365

Now, the pKas like for this one and this one, are not going to be the same as the pKas listed for the 3 amino acids for alanine.1369

They’re probably pretty close, but obviously, it is going to be changing a little bit because now, the environment is different.1375

OK.1382

Let's see, what shall we talk about?1385

Ionizable groups, so in this particular case, we have 3 ionizable groups.1388

OK.1392

Let's see this list: terms we'll be using interchangeably.1394

OK.1407

Let me just go ahead and write this out.1410

We talk about peptide; we talk about protein.1412

We're going to talk about polypeptide.1418

We talk about oligopeptide.1424

Again, these are just all a bunch of different terms that mean a chain of amino acids- a peptide chain.1427

As long as you specify, as long as the person that you're talking to, your audience knows what it is that you're talking about, it doesn't matter what term you use.1437

Certain teachers, they prefer you to be really, really specific, and are a little bit more pedantic about that; but for all practical purposes, again, it is understanding that matters, not little things, so we have to definitely be able to distinguish between what is important and what is not.1445

If you have a teacher that wants you to differentiate between a peptide, a protein, an oligopeptide, a polypeptide, that's fine; but other than that, don't lose any sleep over it.1461

OK.1471

Now, let's talk about some biological activity.1473

Let's see, I think I'm going to start this one on the next page, maybe.1479

Yes, here we go.1484

You know what, let me go back to blue; I really like blue very much.1492

Biological activity of a protein or a peptide, biological activity and size of a peptide or a protein have nothing to do with each other.1499

You might have a peptide that is 3 amino acids long, 6 amino acids long, or you might have one that's 417 amino acids long.1524

The size itself does not correlate to biological activity.1534

The one that is small can have incredible biological activity, and the one that's huge can have incredible biological activity; so, size doesn't mean anything.1539

It does not make it more anymore important- let's put it that way.1548

Just as an example, there is this one peptide that you know very, very well - H3, N, C, C, N, C, C.1552

This is the carbonyl here, the carbonyl here.1567

Let's go ahead and do CH2, COO-.1571

Let's go ahead and put the H on the nitrogens, and this one is going to be phenylalanine, CH2, so the tyrosine without the hydroxy, so this is called L-aspartile-L-phenylalanyl - actually, it is phenylalanine, I should say - phenylalanine methyl ester.1579

I didn't do my ester.1615

OCH3, that's an ester R-group, carbonyl, oxygen, oxygen, this oxygen connected to another carbon.1618

OK.1625

This is NutraSweet.1626

You know that NutraSweet definitely has biological activity, and it is just a dipeptide- NutraSweet, otherwise known aspartame.1628

OK.1640

Some other examples: let's see, there is a protein called cytochrome-C.1641

It has 104 amino acid residues, and it consists of just 1 chain, so one long chain of 104 amino acids.1651

OK.1665

And then, there is something like hemoglobin.1666

Yes, OK.1677

Hemoglobin- let's see, that one has 574 amino acid residues, and it actually consists of 4 different chains.1680

So, in the case of hemoglobin, you have 4 separate chains that are associated with each other.1694

OK.1709

Hemoglobin is the protein that transports oxygen in the blood, just so you know.1710

Another example would be something like a protein called hexokinase or hexokinase, depending on your pronunciation.1721

It happens to have 972 amino acid residues, but it only has 2 chains.1731

Again, just because something has more amino acid residues, doesn’t mean there are going to be more individual chains associated with each other.1740

It has almost twice as many as the hemoglobin amino acids, but it only has 2 chains instead of a 4.1750

OK.1757

And, just so you know, this one happens to be an enzyme which converts glucose - oops, and we’ll definitely be seeing this one later in the second half of the course when we talk about metabolism, when we talk about glycolysis.1758

This one converts glucose to glucose-6 phosphate, the first step in the glycolysis cycle.1821

OK.1836

Now, let's see- hemoglobin, 4 chains, hexokinase 2 chains.1839

Let's talk about this a little bit.1849

Now, proteins like hemoglobin and hexokinase - you know what, I definitely need to slow my writing down just a little bit here - which have two or more individual chains, which associate noncovalently, that's very important, noncovalently, are called multi-subunit proteins.1852

So, if you have a protein like hemoglobin or hexokinase that actually consists of more than 1 chain, well, those chains are going to fold in a certain pattern, and those chains are going to interact with each other noncovalently.1864

Those kinds of proteins, we call them multi-subunit proteins, and each individual chain is called a subunit.1878

Now, there are proteins that actually interact covalently - separate chains - insulin being an example.1903

Insulin consists of actually 2 amino acid chains, but they are actually connected covalently.1912

We don't consider those, multi-subunit, because the interaction between the chains is covalent; but when they are noncovalent, we just happen to call them multi-subunit.1919

OK.1929

Let's take a look at something.1930

This is a picture of hemoglobin; this is hemoglobin.1935

I just wanted you to see a picture of it, and see…now, don’t worry, as far as the spirals are concerned and things like that, we're going to be talking about that a little bit later- what they mean, what they represent.1944

When we talk about protein structure, specifically, we are going to get into more detail about that; but I just wanted you to see of a multi-subunit protein.1956

Notice, this red is 1 subunit; this red 1 up in here, the top right and on the bottom left, this is another subunit; and, of course, you have the 2 blues, that's a third subunit, that’s the third chain, and the fourth chain is right there.1965

So, each one of them has a series of amino acids they fold; and those 4 subunits, they come together and they form the total protein which we call hemoglobin.1981

That’s all that’s going on here.1992

OK.1995

Let's see.1996

Now, let me go back to blue.2006

Now, some proteins contain permanently associated chemical groups attached, and they're called prosthetic groups.2010

A protein could be just a long string of amino acids that has been folded into a protein, and there is nothing else that's involved with it.2045

It is a protein; it does what it does, nothing else, but there are some proteins that don’t just have the amino acid portion, but they actually have other groups that are attached to them, and these groups are called prosthetic groups.2055

Some examples would be - let's see, let's…oops…make sure these lines aren’t there - an example would be lipoproteins.2070

You know what, let me classify this a little bit better.2087

I’m going to give you the class name, and then I'm going to give you the prosthetic group.2090

So, if I talk about a particular lipoprotein, well, the prosthetic group, the thing that happens to be attached to that particular protein, is going to be a lipid, a fat, a lipid, a fat of some sort- that's it.2103

Another class is the class called glycoproteins, a huge, huge class of proteins; and the prosthetic group, a thing that happens to be attached to the protein, they're going to be carbohydrates, otherwise known as sugars.2122

There are things called hemoproteins where the thing that is attached to the prosthetic group is heme; and heme is an iron porphyrin.2144

And again, don't worry about these words; we’re going to be coming back to hemoglobin.2160

We are going to be discussing heme and iron porphyrin and things like that, so right now, I just want you to see the words and see what's going on.2164

A hemoprotein, where the prosthetic group is actually something called a heme group, and it is actually a porphyrin molecule that has an iron in the center; and this hemoglobin is actually a perfect example of a protein that has a prosthetic group, and if you look carefully, you can actually see the - let me do this one in black - you can see the heme groups right here.2170

See, they're actually inside and I know that you can see them in green.2195

There is one there; there is one in this subunit, and I think I see one in this subunit, too, right in there, if you look carefully.2199

Hemoglobin is an example of a hemoprotein.2210

It is a protein, multi-subunit, and each one of the subunits has a prosthetic group.2215

The whole protein has 4 prosthetic groups.2220

OK.2224

There is also another class, the metalloproteins, and it is exactly what you think it is.2226

The prosthetic group happens to be metal ions, for example, maybe zinc, maybe calcium, calcium 2+, maybe magnesium- whatever it happens to be.2232

Let's take an example.2250

Let's take a look at a metalloprotein to see what it might look like.2251

OK.2256

This enzyme is carbonic anhydrase; and I will write down the reaction in just a minute.2257

This is a metalloprotein.2270

This is a particular protein that has a zinc ion, actually, as its prosthetic group; and if you look really carefully, you can actually see the zinc.2272

It is right there, right in there.2283

Now, what I've done is I’ve taken this, and we’ve blown it up a little bit, so that you can actually see the interactions.2286

If we go deep inside the proteins, here is our zinc, and as you can see, its interaction is a noncovalent interaction with what looks like 3 histidine residues and this thing which looks like a hydroxy group.2291

That's what’s going on.2314

This is an example of a metalloprotein.2315

Now, carbonic anhydrase, it catalyzes the following reaction, just so you know, just to have it for information.2319

CO2, + H2O, HCO3, yes HCO3- + H+, so it catalyzes this particular equilibrium- the equilibrium between the CO2, H2O, and bicarbonate, and acid.2333

The active site of this enzyme -again, we have this as zinc ion - is coordinated to 3 histidine residues; and again, this protein has folded in on itself, but these R-groups, they are sticking out, so this particular active site, there are 3 histidine residues in different places, that have positioned themselves in such a way that they can actually trap that zinc ion, histidine resides, and what looks like a hydroxy group and NOH-.2353

That's it; that's all that's going on here.2398

OK.2400

Now, and again, as far as these little twists and turns, these arrows, these ribbons, these lines, we are going to be talking about that a little bit later.2404

Right now, what's important, I just wanted you to see some proteins, see some interactions, things like that.2414

We are going to be talking about what those mean, and they do mean specific things when we talk about protein structure- very, very important.2418

Now, let's go ahead and actually talk about protein structure just globally, real quick, and do a couple more examples of some proteins; and then later on, we'll discuss all of these in a little bit more detail.2427

OK.2445

A protein has 4 levels of structure.2446

Normally, it has 3 levels of structure, but the multi-subunits, they are the fourth level of structure.2450

The first level of structure - let me go ahead and use, that's OK, I'll go stick with black, OK - the primary structure of a protein is its amino acid sequence.2454

That's it.2465

Alanine, tyrosine, isoleucine, leusine, valine- whatever it is, that's the primary structure, the amino acid sequence.2466

Now, the amino acids, once we actually form a peptide chain, there are certain portions of that peptide chain that are going to take particular configurations.2474

Two of those configurations happen to be alpha-helix and beta-pleated sheet.2487

So, those spirals that you see, like for example over here, that means that section of the protein, the backbone, is taking on a spiral shape.2492

These flat sections like these flat sections with little arrow heads on them, that means individual amino acids have arranged themselves in something called a beta-pleated sheet; and again, we will talk in greater detail about these a little bit later.2502

This is called the secondary structure.2517

The secondary structure is where it actually folds on itself, individual portions of the amino acid chain to achieve certain basic structures that keep showing up over and over and over again; and primarily it is going to be the alpha-helix and the beta-sheet.2520

OK.2536

Now, the tertiary structure of a protein, the third level is once this is formed, and a little bit of this is formed, and whatever else is happening along the chain, that chain is going to start folding in on itself.2537

Perhaps a cysteine residue from here and a cysteine residue from there are going to bind and form a disulphide bridge.2550

Maybe there is going to be other interactions.2556

When it is actually folded, that single chain, when it has come to its final folded position, that is called the tertiary structure.2562

That is the 3-dimensional structure, and that's what we've been looking at.2567

An example of a tertiary structure protein is this one right here.2573

Notice, we have some alpha-helices; we have some beta-pleated sheets.2578

These are just regular lines; that means there is nothing is going on.2583

That is just straight amino acids, just a straight chain.2586

There is no particular uniform structure there.2590

It is just amino acid after amino acid.2593

That is this particular structure right here, just a long peptide chain.2596

So, the whole thing, this is an example of tertiary structure.2600

This happens to be the protein firefly luciferase.2604

It is an enzyme- ASE ending tells you it's an enzyme.2615

This protein is a protein that is responsible for the firefly actually being able to glow the way that it does.2617

This is an example of a tertiary structure.2625

This is a single chain.2629

It is a single chain that has folded and has taken up a particular configuration- that's the tertiary structure.2631

OK.2637

Now, when you have a multi-subunit protein, when you have, let's say, 4 chains like hemoglobin, each one takes a particular shape; and then those individual proteins actually associate noncovalently to form the entire protein.2638

When you have a multi-subunit protein, that's the fourth level of structure.2656

That is the quaternary structure.2661

So, primary, secondary, tertiary, quaternary- just wanted you to be aware of this particular setup in terms of structure; and then we are going to be talking about these in future lessons in rather great detail.2664

Let's go ahead and check out one more.2680

Again, in this particular case, this is going to be an example of quaternary structure.2684

This is hemoglobin, again, the one that we saw before.2689

You have your 1 chain right there, another chain, another chain, and another chain.2695

Each one of those represents the tertiary structure.2704

When they come together to form the entire final protein, you have your quaternary structure.2707

That's it.2711

OK.2713

Thank you for joining us here at Educator.com and Biochemistry.2714

We'll see you next time, bye-bye.2717

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

At the close of the last lesson, we talked about the levels of protein structure; we had primary, secondary, tertiary and quaternary.0004

Today, we're going to talk about the primary structure- the amino acid sequence.0012

We want to know what is the sequence of amino acids- which is next to which, which amino acid is next to which, and how are they arranged in a linear fashion.0017

That is what we're going to be working on.0028

Let's get started.0029

OK.0032

The amino acid sequence determines how the peptide is going to actually fold, how the peptide will fold, and thus, ultimately determines its structure.0034

As you can see, the primary sequence of a protein, of a peptide, is very, very important because depending on what amino acids are aware, it's going to basically guide how the protein is going to assume its 3-dimensional shape; and it is that 3-dimensional shape which is going to determine its structure and function.0080

Let me write these words a little bit better.0109

The amino acid sequence determines how the peptide will fold and thus ultimately determines its structure and function.0113

Amino acid sequence implies the function, and that's what is important in a protein, what does it do.0126

OK.0135

Well, there are many techniques for elucidating amino acid sequence.0137

We will discuss a chemical method.0160

We will discuss a chemical method still used in laboratories.0165

It is called the Edman degradation.0174

Excuse me.0186

Basically, what the Edman degradation does is it labels and removes the N-terminal amino acid for identification.0187

It labels it for identification.0210

It removes it so that it can be separated, and that way you can identify it.0211

The remaining peptide, now, has a new N-terminal amino acid; and now, what we do is we just repeat the process.0218

That's it.0243

Excuse me.0245

We're basically taking an amino acid and we're labeling the end, cutting it off, identifying it.0247

Next one, labeling the end, cutting it off, identifying it, and we just go down the list.0250

We are just chopping it up until we finally get to the last amino acid.0254

That's all the Edman degradation does, and, of course, this is an automated procedure because we have really, really good chemical control; so we can just put our sample into a machine, and it will do everything for us, and it will give us a read out at the end.0259

It is really quite wonderful.0271

OK.0275

I'm going to do a schematic representation of the Edman degradation describing each step, and then we’ll go ahead and do an example of an Edman degradation with a specific amino acid.0276

OK.0289

I wonder if I should start on a new - yes - let me go ahead and start on a new page here.0291

Yes, that's fine.0296

This is going to be the Edman degradation; let me go ahead and do this in blue.0299

OK.0306

I have to warn you there is going to be a lot of chemical names being thrown around, and there is going to be a lot of chemical structures being thrown around.0307

This is where you have to be really, really, really careful, and that includes me.0315

So, please, by all means, you definitely want to confirm that I'm actually drawing the right structures.0320

I would definitely encourage you to take a look at the Edman degradation procedure in your book to see what they have to say about the particular mechanism and how they draw it- really, really important.0330

But again, ultimately, it is just not about passive learning.0340

You don't just want to look at a diagram and say I understand it; you need to be able to reproduce it.0344

That's when you actually understand it.0350

OK, so, the Edman degradation.0352

Let's start off with just a generic peptide.0354

We have H3, N, C, C, and I'm just going to go ahead and write peptide for the other because again, we're just going to be concerned with the N-terminal, the one on the left.0360

We have the carbonyl carbon there, and we have our R-group attached to the alpha-carbon, and this is A+.0372

The first step is...where should I write this, I'll go ahead and write it here, wonder if I should do it in, this one I'm going to do in black, I think.0379

OK.0402

I'm going to be drawing this thing, N, double bond C, double bond S.0403

OK.0410

What we do is we take this peptide and we react it with something called phenyl isothiocyanate under mildly alkaline conditions.0411

OK.0418

That's the first step0419

Let me go ahead and write this as one.0421

I'm going to write the products below instead of to the right.0425

I'm going to write the steps over here; I just wanted to do it in a schematic way.0427

You know what I need a little bit more room to write this out.0438

One, phenyl isothiocyanate- that is this molecule right here.0443

OK.0454

It is abbreviated PITC, phenyl isothiocyanate, under mild basic conditions, alkaline conditions - there we go - under mild OH.0455

This is the Edman reagent, so you'll often hear it.0471

They might say PITC, or they will just say “use Edman reagent”.0475

This is our Edman reagent; let me go ahead and put that there.0480

This is called the Edman reagent; let me go back to black.0485

OK.0490

When this reaction actually takes place, what you end up with is this product.0491

Let me see.0499

It is going to be this here; let's go ahead and put the H on there.0500

It is going to be C, double bonded S, and it is going to be attached to the N, C, C.0510

This is carbonyl, and this is our peptide.0518

This is our R-group, and we have our H.0523

OK.0527

The bond is formed between this carbon and that nitrogen.0531

This is the bond that is formed- right there.0539

OK.0542

Now, again, let's keep track of our peptide.0544

Our peptide is right here- N, C, C.0548

That is what you want to look for.0550

When you're doing these yourself, again, keep track of your peptide; and you can keep track of it by looking for that N, C, C motif.0551

N to the left, C to the right, C to the right, carbonyl on the second C, R-group on the first C, counting from left to right- here is our peptide, I'm sorry, here is our amino acid.0559

This is the one that we're actually pulling off.0574

This is the isothiocyanate part here.0576

So, what I've actually ended up forming here is something called phenylthiocarbamoyl that refers to this particular arrangement of atoms.0580

Phenyl is this, thio is the sulfur, carbamoyl is this carbon attached to a nitrogen and a nitrogen here.0599

What I've done is I've taken this peptide that I have, and I've created a phenylthiocarbamoyl derivative of it, by reacting it with the phenyl isothiocyanate, the PITC.0606

This phenylthiocarbamoyl derivative, they call it PTC.0620

OK.0626

Now, we'll go to our second step.0627

Now, we'll go ahead and do another black here.0628

OK.0633

This one, we are going to react it with C, COOH; and this is going to be anhydrous.0634

What we're going to do is, we're going to react this PTC with anhydrous trifluoroacetic acid.0644

It is just a weak acid that happens to be a little bit stronger than acidic acid.0663

Actually, any acid will do; it's fine.0667

It just needs to be anhydrous.0669

Now, what happens when this reaction takes place is the following.0672

What you end up with is the following 2 molecules; this is the one that actually breaks the bond that we are trying to break.0675

OK.0683

And, I'll tell you which bond in just a minute once I draw it out.0685

Let' me see.0688

That's fine; I guess I can fit it in here.0689

Let me go back to blue.0691

We have our C, we have our NH, and we have our phenyl group, C, then we have our...here's our N, here is our C, and here's our C, that is our that, and then we have our S, and C, and we have our 1.0693

Let me make sure I have everything on here, N, trivalent, S.0728

OK.0733

Everything is good.0734

Yes, and, of course, we have that plus our new peptide, 3 peptide- the new amino-terminus.0735

The bond that we have actually broken is the following.0750

We've broken this bond; let me do this in black.0754

We have broken this bond, and again I'm going to go through the mechanism in just a little bit, but I just wanted you to see chemically what happens.0765

This thing, when we form this species or again, let me see, N, C, C, keep track of the N, C, C.0771

This is our amino acid.0781

OK.0784

This is our that; this is our that.0785

That is what this is.0787

We want to keep track of our amino acids.0788

This is called, in case you want to know, it's called an anilinothiazolinone.0790

Anilino refers to the phenyl group attached to nitrogen; thiozolinone happens to be this thing, the C, the N, the S, arranged in a ring.0803

OK.0813

Now, we take the third step - oops this is not...this is 2, not number 1, number 1 was that, sorry about that - this is step 2 of the Edman degradation.0816

Now, we're going to go to step 3 of the Edman degradation.0829

Let me move on to the next page, and let me write the molecule in blue.0832

Now, we've pulled off that other peptide.0837

We have that one N-terminus that we've actually broken off; that's the thing that we're going to react.0842

Let me redraw that one; let me draw it here.0847

C, we have NH, we have that, we have N, we have C, we have C, we have S, this is our R group, this is our carbonyl.0852

Let me see; am I missing anything here?0871

No, I don't think so; everything looks good.0874

OK.0877

Now, again, let me, N, C, C, just to keep track of our amino acid or N-terminal.0878

Now, the third step here, what we do is we're just going to react this particular molecule with aqueous acid.0888

So, step 3 is aqueous acid, and what you end up with is the following molecule.0901

Let me do this in...yes, that's fine; I'll go ahead and do it in blue.0913

We have C, we have S, we have N, we have C, we have C, and we have N, and we have phenyl, and we have an H, we have our R group, and we have that.0917

OK.0943

The reason we actually do this step is this thing is more stable than this thing, so it allows us to deal with it better.0946

This is more stable, and it is called phenylthiohydantoin; and this is PTH- that's the acronym.0952

Now, it's ready for identification.0972

There you go.0983

And now, let's go ahead and red N, C, C.0985

That is our motif; that’s what we want to keep track of.0990

All that has happened here is that this thing under acidic conditions, aqueous acidic conditions, has actually rearranged, and has formed something more stable.0993

You want to take a look what has happened form here to here.1004

The only thing that has happened is this carbon right here that is attached to nitrogen, this S, went up to where the nitrogen was, double bond; this nitrogen with the phenyl group came down to where the S was.1006

This S and this thing switched places; that's all that happened- the rearrangement.1019

OK.1026

And, of course, the last part, since now you have the H3N - oops - you have the peptide left over, the fourth step.1028

Let me do this in blue.1038

H3, N+, now, you have the rest of the peptide with a new N-terminal amino acid group.1040

So, step 4, just repeat the process.1047

That is the Edman degradation.1052

The first step is phenyl isothiocyanate, and then after that, you're going to treat it with trifluoroacetic acid; third step, you're going to treat it with aqueous acid, and you're going to form this molecule right here- this phenylthiohydantoin.1054

You've basically taken this N-amino acid, and you've labeled it with this thing.1075

You've made a derivative of this thing for that thing, and now, you can identify it; and you just repeat the process, go down the chain.1080

OK.1087

Now, let's take a look at some mechanisms.1088

It's important to talk about mechanisms, how electrons move, arrow pushing.1090

You remember from organic chemistry, electrons go this way, nucleophile, electrophile.1095

If it is something that's strange to you or perhaps you’re not too familiar with it, it intimidates you a little bit, don't worry about it.1101

I think it will just be reasonably clear what are these that's going on.1108

Don't attach any more deeper meaning than what it actually is.1111

It is just electrons moving around forming bonds.1116

You remember in general chemistry, we just sort of do this chemistry, and we wouldn’t talk about how it happened.1119

When you got to organic chemistry, that's when you started talking about "OK, this carbon is moving in here, these electrons are forming this bond, this bond is breaking"- that's all a mechanism is.1124

It is a molecular level, single step, what's happening.1134

OK.1140

Let's see if we can do - let's do this in blue - mechanism for PTC formation.1141

That is the phenylthiocarbamoyl formation.1156

This is step one.1167

OK.1175

Here we go.1176

Let's go ahead and draw out the phenyl isothiocyanate first.1177

So, I’m going to draw this vertically.1182

Actually, let me do this in black.1185

OK.1188

And, we've got N, we have C and S, so this is our PITC- phenyl isothiocyanate.1191

Now, let's go ahead and write our H2; this is N.1200

I’ll go ahead and put the electrons on the nitrogen; I’ve got N, C, C, and then I've got...I'm going to actually write out the second, N, C, C, N, C, C.1204

Carbonyl goes here; carbonyl goes here.1220

I'm just going to do it for a dipeptide.1222

This is going to be the R1 group; this is going to be the R2 group, and let's go ahead and put an H on that nitrogen.1225

OK.1232

Here is what happens.1233

These electrons, it is a nucleophile; this nitrogen is a nucleophile.1236

This carbon here that is attached to nitrogen and sulfur, it's the electrophile.1242

It is a little bit positively charged.1248

Nitrogen is an electronegative element.1249

It's going to pull electrons away from that.1252

This is negatively charged.1253

So, these electrons are going to attack here, and when these electrons come in, electrons that are there have to make room for these that come in, so they have to go away.1256

These electrons move away and they grab an H from the solution; and what you end up getting is the following.1269

Should I draw it?1280

Yes, that is fine; I'll go ahead and draw it horizontally.1284

N, H, C, double bond S, this is N, H.1292

It is going to be C, C, and then N, C, C.1302

OK.1311

Well, that's fine; I'll do this in just a minute.1312

N, C, C, this is the carbonyl, this is our R1 group, there is an H here, this is our R2 group.1315

So, the bond that we formed is this bond right here.1325

That is the bond that we formed.1329

These electrons formed this bond.1330

Now, notice, it has 2 hydrogens on it, but this nitrogen now, has 1 hydrogen on it.1333

So, I'm going to go ahead and write this minus H plus.1339

That means that it has given up that hydrogen.1342

Once this bond forms, now, nitrogen has 1, 2, 3, 4 things attached to it.1343

It is going to be positively charged.1351

It is going to release that hydrogen in the solution.1352

This is the mechanism.1357

Nitrogen is the nucleophile; this carbon of the PITC is the electrophile- standard, basic mechanism, single step.1359

OK.1369

This is our PTC, phenylthiocarbamoyl.1370

OK.1377

Now, let's go ahead and do the mechanism for the second step.1380

This is very important.1384

This one we'll do in blue again.1386

This is going to be the mechanism for ring formation and peptide bond cleavage.1389

This is the big one.1408

This is step 2.1414

This is where we add the trifluoroacetic acid.1416

OK, step 2.1419

So, we've added the trifluoroacetic acid, this is what happens.1421

Let's draw our molecule again, and we'll make sure to draw it very, very carefully.1424

And again, you need to be able to reproduce this.1429

OK.1433

It's the only way you'll have a full grasp of what it is that is going on.1434

We have N, C, C, N, C, C.1437

There is an H here, our carbonyl goes there, carbonyl goes there.1445

This is our R1 group; this is our R2 group, and let me go ahead and put the electrons on the nitrogen on that one.1450

OK.1462

Now, here is what happens.1463

Alright.1466

We are just going to be pushing arrows; electrons are going to be moving around.1467

Here is what happens.1471

Actually, you know what, I'm going to make this arrow a little bit smaller here because I want to do 3 structures on this page.1473

OK.1481

These arrows right here on the nitrogen, that is N, C, C, our N-terminal, OK, this nitrogen.1482

Again, look for the N, C, C.1489

This is N, C, N; that's not it- N, C, C.1491

This is your terminal amino acid.1493

These electrons, they go that way.1498

These electrons, they push, these electrons they attack that.1501

You know what, I'm going to do this in a different color.1509

Sorry about that.1513

Let me do this in red.1516

These electrons go down here to form a double bond.1520

They push these electrons; they attack the carbonyl right here, and these actually end up going up onto oxygen.1522

So, what you end up getting is this tetrahedral intermediate, which is very typical of carbonyl reactivity.1533

What you end up with is the following.1540

Now, I'm going to retain certain structural features.1542

I'm going to keep this C, this arrangement, while I draw a structure.1545

It's going to be...let me do this in blue.1551

I'll try to do it underneath.1554

C, C, O-, N, C, C, let me fill these up, R2.1557

Now, the S, these electrons have moved and formed a bond here, so what I have is, I have formed a bond with this sulfur.1568

Now, this sulfur is attached to this carbon.1576

It is attached to that carbon.1581

Well, that carbon is now attached to this nitrogen with a double bond, and that nitrogen is attached to this carbon.1583

That is what's happening.1595

Also attached to this carbon is the...that's it.1597

Just keep track of your carbons.1608

That is all that's happening here.1611

Again, and this is N, C, C, so the R1 group is right here.1613

Again, let's keep track of our...OK.1620

We formed this as an intermediate species.1632

Now, the next step of the reaction is the following.1635

Let me do this in red again.1642

OK.1644

This bond is the bond that we are going to break right here- right between the C and the N.1645

This is one amino acid; this is the other amino acid residue- N, C, C, N, C, C.1652

So, what happens is these electrons right here, they go back down to form the carbonyl because the carbonyl is very stable, and they kick off these bonds, and they go on to grab an H+.1659

There is an H right here, by the way.1673

And therefore, this bond is actually broken.1676

What you end up with is the following.1680

I'm going to draw this out.1690

This is going to be in blue.1692

Actually, you know what, let me draw...that's fine.1695

I'll just go ahead and do that, that's fine, but I'm going to draw this molecule over here.1702

This is going to be C…nope, do it in blue.1710

We have C, we have C, we have the carbonyl is formed again.1713

We have S, we have C, we have double...oops...we have double bonded N, single bonded C.1720

We have our R1 group, and, of course, here, we have our NH and our benzene ring, plus we have N, NH.1736

It grabbed an H, so we have NH2, C, C, O-, R2, and again, keep track, N, C, C.1750

There we go.1769

We have our amino acid; we have our derivative part, and this is the one that undergoes that rearrangement to form the final PTH; but I wanted you to see this mechanism.1771

It is the nitrogen electrons that move here to form the double bonded carbon.1788

They push the double bond on the sulfur, and attacks the carbonyl.1792

The electrons move up onto the oxygen to carry a negative charge.1795

The electrons come back down to form the carbonyl, and they kick off these electrons to have it to do whatever it does; and that actually breaks this bond right there.1798

Let me go ahead and do this in black after the fact- this bond is broken.1809

There you go.1823

That is the mechanism.1824

OK.1827

Let's go on here, see what we can do.1828

Let's go ahead and do an example.1830

OK, an example.1833

OK.1840

Write out the Edman degradation for the tripeptide Ala, Tyr, Ser- alanine, tyrosyl, serine.1843

We have this tripeptide.1873

We want you to write out structurally the Edman Degradation using arrows, not mechanism arrows.1874

We just want you to show what reagents you are using, what the products are going to be for the entire Edman degradation for this thing.1880

OK.1887

Let's just jump in, and you need to be able to do this.1888

You have to be able to reproduce this - very, very important.1889

You get practice with amino acid structures; you get practice with writing out the PITC, PTC, PTH- all of that stuff.1892

Again, you do enough of this, 3 or 4, 5 of these, you'll be perfect; but you have to do them.1900

OK.1907

Let's draw it out.1910

Let's see; let's go.1913

Shall we do it in blue or black?1915

It doesn't really matter; let's do it in blue.1917

Again, do the backbone first, N, C, C...oops...N, C, C, N, C, C.1919

And again, you can use the shape structure; you can use line structure- whatever works best for you.1931

I just love seeing everything.1935

We've got H2 or H3- it doesn't really matter.1937

We have carbonyl on the second carbon, carbonyl on the second carbon, carbonyl on the second carbon.1944

Let's go ahead and put an O- there.1948

We have alanine, which is CH3.1950

Notice, I'm not putting the H on the alpha-carbon anymore.1953

We have tyrosine.1957

I probably should have picked something a lot easier, but OK, and a little less tedious to draw out, but that's OK; it's good practice.1959

I like tyrosine and serine, which is CH2OH, if I'm not mistaken.1971

OK.1978

The first step is, you are going to use PITC, phenyl isothiocyanate, under mildly basic conditions, and you are going to form the following.1979

You are going to form N, C, S, N, H, C, C.1996

Here, this is going to be CH3, and this is going to be N, C, C, N, C, C.2014

We've got N, H, C.2022

This is going to be CH2.2026

We have OH, we have our carbonyl, and we have NH, we have CH2OH, we have that, and we have that.2031

That is our first step.2040

We have formed this thing.2041

OK.2044

This is our phenylthiohydantoin.2045

So, we've formed this PTC thing.2052

Let me just go ahead and write that in red.2053

Where do I put it?2058

That is fine; I'll just put it here.2059

We have formed PTC.2060

OK.2062

Our next step, let's go ahead and actually...that's fine; I'll just do it on the next page.2063

Let's go back to blue.2069

Now, we're going to use trifluoroacetic acid.2071

Let's just write TFA- trifluoroacetic acid.2078

When I do that, I'm going to actually form a ring, and I'm going to break a bond.2082

So, let's see which bond am I going to break.2087

Well, I have that.2091

I'm looking for N, C, C, N, C, C, N - my first peptide bond.2092

That is the bond that is going to break.2096

The ring that I'm going to form is going to be made up of sulfur, 1, 2, 3, 4, 5, 5-membered rings starting with sulfur - sulfur, carbon, nitrogen, carbon, carbon.2099

That is my 5-membered ring.2114

OK.2116

Let's go ahead and form that then.2117

Let me see; do I actually do a...yes, that's not a problem.2122

Let's go ahead and form that.2127

Let me write it out over here.2129

Let me go back to blue.2134

It is going to be C, C, carbonyl, CH3; it's going to be N, C, S, and on this C is going to be the NH, and it's going to be that thing, and what we are left with is...where am I? yes... I'm left with tyrosine and serine.2136

I'm left with H3, N+, N, C, C, N, C, C.2176

I've got a carbonyl there; I’ve got a carbonyl there.2184

This is CH2, and yes it is CH2, and this is going to be my tyrosine.2190

As you can see, keeping track of all these gets really kind of confusing.2207

So, don't feel bad if you have difficulty with this because we all do.2211

OK.2216

So, I formed this thing right here, my anilinothiazolinone; and this is what I'm going to subject to H+, under aqueous conditions.2217

And again, what we want to reverse is, this and this are going to switch places.2235

I'm going to leave everything the same; I'm just going to switch that and that.2245

I'm going to write this one in back to blue.2249

I'm going to go C, C; I'm going to go N.2252

I'm going to go C; I'm going to go N that way.2259

This one is the carbonyl; this is the alanine.2264

This C has now an S, and this actually has that phenyl group attached to it.2270

So, this is my final product; this is my PTH, my phenylthiohydantoin.2279

This is the one that I'm going to identify.2285

And again, my amino acid, N, C, C, is right here.2287

That's it.2295

Now, we go ahead and we take the next step.2297

Now, we take in this molecule, so we've gone ahead and identified one, now, we are going to subject this molecule - I hope you don't mind if I change colors here - I'm going to react now, this one for the second cycle.2299

I'm going to react it with PITC under mildly alkaline conditions, and I'm going to end up forming the following molecule.2314

Yes, that's very, very important that you write all of these out, at least a couple of times.2325

C, S, N, C, C, N, C, C, N, C, C, that carbonyl goes there, that carbonyl goes there, and here we have the tyrosine R-group, and here we have the serine R-group.2330

OK.2357

We have actually formed this bond right here with the phenyl isothiocyanate.2358

Now, this is the one that we are going to subject to trifluoroacetic acid in order to form the ring and break this bond.2365

Now, we are going to break that bond, and we are going to form a ring from this molecule.2376

Let's go ahead and form that; let's see what that looks like.2382

That is going to end up looking like this.2386

It is going to be C, C, O, we have an S, we have a C, we have double bonded N, we have that.2389

On this C, we have NH, and we have that.2402

And on this C, we have our tyrosine, CH2, and OH.2408

Now, we have plus our CH2, N, C, C.2417

We have our final serine residue which is going to be CH2OH.2425

Oops, let me do it the way that I usually do which is vertically.2430

OK.2436

That one is taken cared of, and this one is going to go on into a third cycle, which I will have you do.2437

And, let me see, this is going to be our anilinothiazolinone, which we are going to subject to aqueous acid; and when we subject it to aqueous acid, we are going to rearrange.2444

And again, what we are going to rearrange is, this thing and this thing are going to switch places.2457

Everything stays the same except those that switch places.2463

So, what I end up with is C, C, N goes there, C stays, N goes here.2466

And again, this double bond changes; it becomes a single bond that goes there.2478

That is a carbonyl; this is going to be our tyrosine group.2482

OK.2489

And this is going to happen...no, sorry about that, this and this switched, so we actually have that there.2493

This one has an H, and, of course, we have an S; but we don't want these stray lines, otherwise you are going to think that they are double bonds.2502

We don't want that.2509

So, we have that.2510

That is our final PTH, and in this particular case, our amino acid residue is right there.2513

Again, N, C, C, just follow the N, C, C, and attach the rest.2523

We can identify this, and then we just subject this to the next cycle.2530

That is it; that' all you are doing.2535

Again, it is very, very important that you do at least a couple of these by writing out the reactants, the reagents, and the product.2537

That is the only way to get a full sense of what's going on, to have full command of what's going on; and I promise you, after doing a couple of these, you'll really, really feel like you understand the material, drawing it out actively.2546

That is the only way to learn this.2561

OK.2563

Thank you for joining us here at Educator.com and Biochemistry.2564

We'll see you next time, bye-bye.2566

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today we're going to continue our discussion of sequencing a peptide or a protein, and we're going to talk about how to sequence larger proteins, and it's exactly what you might think it is.0004

Basically, you just take a larger protein, and you chop it up into fragments, and then you deal with those individual fragments, and then you just arrange the fragments.0015

Let's just go ahead and jump on in.0025

OK.0028

Before we actually get into sequencing the larger peptides and proteins, I want to talk about one other method that's used to identify the N-terminal amino acid because it is going to come up.0030

Another method for IDing the N-terminal amino acid, and again I'll just use AA for amino acid.0046

OK.0065

Frederick Sanger, who worked out the amino acid sequence of insulin, developed a reagent to identify the N-terminal amino acid.0068

OK.0103

It is called 1-flouro-2,4-dinitrobenzine.0104

I tend to separate the words; you'll see it as one word, dinitrobenzene, small b, but I tend to separate it.0115

It doesn't really matter.0121

We will just refer to it as FDNB- fluorodinitrobenzene.0123

OK.0129

Here is the procedure.0130

We will take our amino acid.0136

Let's just go ahead and go, let's see H3N+ we have C, C, we have that, and over here, actually you know what, I'll put the O a little bit better, and over here I'll just write peptide, because again, we're just going to be concerned with the N-terminal amino acids, this thing right here; and I'll just make it generic so I'll put an R1.0142

So, what we do is we react this with our FDNB, which I'll go ahead and draw the structure.0166

Let me just write FDNB, and the structure is this: 1-flouro or dinitrobenzene 1-flouro 2, 3, 4, 1, 2, 3, 4, 5, 6, numbered carbons.0172

We react it with FDNB, and what you do is you basically just create a derivative of this, so this is like a label, just like what we did before.0193

You are just labelling the amino acids so you can actually identify it.0200

What you end up with is the following.0203

Let's go N, C, C.0205

I'll write peptide, and we have a carbonyl here.0211

We have our R1 group; we have our H, and we have this right here.0216

I'll go ahead and put the NO2 over here.0229

I just flip this one around.0233

Basically, you are just replacing this fluorine with the N-terminal amino acid.0236

That is what you're doing; it just acts as a label.0240

That is it.0243

Once you go ahead and form this, once you've actually labelled this, so let me go ahead and put a little circle around our amino acid, so that's our N-terminal amino acid, and then what you do is you go ahead and react this with 6M HCl to hydrolyze, to completely break apart every single bond just to free-up every amino acid; and then when you do that, what you end up with is, of course, the labelled terminal amino acid.0244

You are going to end up with this; you're going to have the R1.0279

You have H, and then, of course, you have your label, so NO2, and I'll just put the NO2 over here plus free amino acids.0283

That is it- free amino acids.0296

When you use strong acids like that 6M HCl, you’re just going to completely hydrolyze the protein into all of its free amino acids.0298

OK.0308

And, let me write here again, 6M HCl breaks all the peptide bonds, and, of course, this thing right here, this is what we're going to be using, we just want to identify the N-terminal side.0310

That is it.0330

That is all this Sanger reagent does is it identifies the N-terminal amino acid.0331

OK.0337

You are probably wondering to yourself, well, why would we even bother with this if we can just go ahead and use the procedure that we used before, the Edman degradation, and just identify all of them and sequence them that way.0338

You will see why it's actually important in a minute when we start sequencing larger proteins.0348

OK.0354

Now, let's go ahead and start talking about sequencing larger proteins and peptides.0355

That is fine; I’ll go ahead and keep it in black.0361

Well, it might be nice if I actually wrote down the words properly.0369

How's that?0373

Sequencing larger peptides and proteins.0380

OK.0391

As we said, in the intro, these have to be broken down into smaller fragments, and these fragments are sequenced just like before with the Edman degradation.0393

Let's say you have a protein or a peptide and you chop it up into 4 fragments.0427

Well, you sequence those 4, well, you'll see the procedure in just a minute, but essentially what you do then, you have to decide which fragment is in front of which fragment; and we have some techniques to deal with ordering those.0433

That is all that's going on.0446

You are doing the same thing, except you're doing it for more fragments.0448

OK.0453

Not only do the...because the protein is a peptide, you have to break it up into smaller fragments, but you remember these disulfide bonds, the disulfide bridges that exist between the cysteine residues, they are actually covalent.0454

We have to break those because they interfere with the sequence processing.0465

We also have to break the disulfide bonds in a protein, and that is usually the first thing you do, you break the disulfide bonds then you start chopping up the protein.0469

OK.0489

Now, let's talk about the chopping up the protein part, and we'll get back to the disulfide bonds in just a minute.0490

There are enzymes called proteases which catalyze the breakage of peptide bonds next to specific amino acids.0497

So, the fragmentation process or the actual fragmentation is predictable and reproducible.0542

This is important- predictable and reproducible.0552

We use enzymes called proteases to actually take that really, really long protein or peptide, and chop it up into specific places; and depending on what enzyme we use, it actually cuts the peptide chain next to specific amino acids, maybe 1 amino acid, maybe 2 amino acids, but it does not mess with all of the other bonds, not like the 6M HCl does.0560

So, these proteases are perfect, and because they cut the peptide chain next to specific amino acids, they're reproducible and they're predictable, so we know exactly where they're going to be cut, and that's what's really nice.0587

OK.0602

In addition to enzymes called proteases, we also have some chemical methods that do the same thing.0604

Chemical methods also exist.0612

Chemical methods also exist for breaking peptide bonds at specific points, for breaking peptide bonds next to specific amino acids.0618

OK.0643

Now, as a reminder of what I just said, note again that these enzymes, chemicals, they break single bonds to create short fragments.0644

They do not hydrolyze the entire peptide.0682

They do not hydrolyze -that's a Y -the entire protein.0690

OK.0696

We are creating fragments, short peptides.0697

We are not just breaking it up into free amino acids like HCl.0699

OK.0704

Let's go ahead and list some of these.0705

Let me go ahead and do this one in red.0706

Some of the enzymes and/or chemicals - I'm going to do mostly enzymes, only 1 chemical - used for fragmentation, some enzymes and chemicals used for the fragmentation process.0711

OK.0735

We will go ahead and list the name, and we'll go ahead and list the cleavage point, in other words, where it actually breaks the peptide.0736

OK.0752

There is an enzyme called trypsin, and it breaks lysine and arginine C.0753

This enzyme trypsin, when we use that to break up a protein or a very large peptide, it will actually cut the peptide chain next to a lysine, next to an arginine.0763

I will explain what the C means in just a minute.0773

I just want to go through the list, and then I'll tell you where this tells us...well, I'll tell you in just a second.0776

Chymotrypsin, OK, I'll put enzyme so you know this is also an enzyme.0785

It breaks it up at phenylalanine, tryptophan and tyrosine next to the aromatic amino acids and C.0795

OK.0809

Cyanogen bromide, this is a chemical procedure, and it breaks it up next to a methionine, and last, pepsin.0812

It is an enzyme and it's a phenyl tryptophan tyrosine with an N.0831

Now, I'll tell you what the C and the N mean.0840

I will tell you what they are, then I'll go ahead and write them down.0843

Trypsin, when it cuts a peptide chain, it cuts it either next to a lysine or an arginine, but what it does is it cuts the peptide bond on the carbon side, the carboxyl carbon, in other words, to the right of this amino acid.0845

OK.0861

Now, notice this one right here, pepsin.0862

Pepsin phenyl tryptophan tyrosine N - this means that it actually cuts the peptide next to a phenylalanine, a tryptophan or a tyrosine, but it cuts the peptide bond to the left of these amino acids.0865

In other words, it breaks the N-terminal, the N side, the nitrogen side, right?0881

So, what you have is something like this.0888

What you have is some amino acid residue that looks like that.0889

This is going to go on on a peptide that way; this is going to go on on a peptide that way.0895

Let's say what we are dealing with is a phenylalanine.0901

Well, if I use trypsin, oh I'm sorry, chymotrypsin, and this is phenylalanine, it is going to break it up on the C side, so it is going to end up breaking that bond; and then we will have this fragment on the left and we will have this fragment, but if we use pepsin and it's phenylalanine, it is going to break that bond.0904

It is still next to a phenylalanine, but it is going to be phenylalanine the N side of phenylalanine.0926

That is all that means.0932

These Ns and Cs, it tells you which peptide bond that is actually breaking because you have 2 of them for each amino acid, the one on the right, the one on the left, the carboxyl, and the amino bond.0933

OK.0945

Let me just go ahead and write that down formally, go back to blue.0948

C, it means it cleaves the peptide bond at the C end of the amino acid, and N means it cleaves the peptide bond at the nitrogen end of the amino acid.0957

That is all that means, so either to the right, to the C end, or to the left of that particular amino acid.0997

OK.1004

Let's just do a couple of examples really quickly just to make sure we absolutely get this.1006

Let's say we have this following H3N+.1016

Let's see, we have Ala, we have Gly, we have Phe, and Met, and Leu, and Leu, and Tyr, and Ala, and of course we have that.1023

Let's say we have this little peptide right here.1045

Well, if we use chymotrypsin - I'll write it down below - if I'll use chymotrypsin, well, I notice that the chymotrypsin is the one that cleaves next to the aromatics, the phenylalanine, the tyrosine and the tryptophan, so I look here and I notice that I have a phenylalanine and I have a tyrosine and it clips it on the carbon side of the peptide bond.1048

So, chymotrypsin is going to break the bond here; and it is going to break the bond there to the right- that one and that one.1080

It is going to clip it that way.1088

So, we are going to create 3 fragments: this fragment, the middle fragment, and this fragment- if we use chymotrypsin.1091

OK.1098

Now, let me go ahead and rewrite this.1099

This time let me write it in blue again.1102

H3N+, we have Ala, we have Gly, we have Phe, we have Met, Leu, Leu, Tyr, Ala, and we have our final carboxyl group over there.1105

Now, if I use pepsin, let me go to red, well, pepsin does the same thing.1126

It still breaks it up next to the phenyl alanine and the tyrosine.1131

There is no tryptophan in here, but it breaks it up on the nitrogen side, on the left, the amino peptide bond.1136

So, we are going to break that bond, and we are going to break tyrosine to the left of that bond.1144

I have this fragment, and I have this fragment, and I have this fragment.1151

Again, 3 fragments, but I've snipped it in a different place.1154

That is all that's going on here.1159

OK.1161

Now, let's see.1164

OK, let's continue on.1166

OK.1171

Once we have a set of fragments whose sequence we have deduced by using the Edman degradation, so I break up my protein, I have my separate fragments, and then I go ahead and I sequence those fragments, once I know that, we use another hydrolyzing reagent, another enzyme, we use a different hydrolyzing reagent, in other words, an enzyme on that list or some chemical on that list, to create a different set of fragments with sequences we deduced.1175

Just like we did for the example a second ago, we used the chymotrypsin and then we used pepsin, we broke it up into separate fragments.1264

We do the first one, we break up those fragments, we sequence those 3 fragments, and then we use another enzyme or a chemical to get a different set of fragments, we sequence those, and we compare the two, and that is how we set up the order, and that's all we are doing.1276

Let me see whose sequences we deduced.1291

OK.1295

We then compare the 2 sets of fragments because now, we have to arrange the fragments in a given order to find overlap, and when we have overlap, thereby establishing the overall sequence, and I'll draw this out in just a minute.1299

This is actually really, really amazing that we can do this.1334

Let's say for example - let me do this in red - so, we have this whole peptide chain, so I’ll just write whole; let's say we use one of those chemicals on that list to hydrolyze this protein at specific points, to break it up.1338

Let's say we break it up into that fragment and then that fragment, and then maybe that fragment, and let's say that fragment.1351

OK.1362

Here we have 4 fragments, something like that, used in one of the enzymes or one of the chemicals on that list to break it up.1364

Now, we take this whole peptide protein; we do it again, but this time with a different hydrolyzing reagent, so maybe we end up with this.1376

We end up with that fragment, that fragment, and let's say maybe that fragment.1384

Now, we have 3 fragments.1393

Well, now, the thing is, I have a bunch of fragments, and I know the sequences of each of these fragments.1397

I know those sequences, but I don't know what order they go in.1403

I have to arrange the order.1406

What I do is I compare this with this, and I set up overlaps.1409

That's it.1416

Notice here, I have some overlap.1418

There is overlap there; I have here.1425

There is a bunch of overlap here and here.1427

OK.1430

Of course, here there is overlap between those two, this one and this one.1431

This overlaps these two; this one overlaps this, this, and this.1436

This one overlaps this and this.1440

By comparing and seeing what the overlap is because I know the sequences, I know where every amino acid is, I can establish which fragment goes where.1442

Does this one go first?1450

Does this one go first?1452

Now, that way I've drawn it, I've cut it in certain places; but once I’ve cut the fragments, they are just randomly ordered, the fragments are.1453

I have to see which fragment goes first, which fragment goes second.1462

That is all we are doing.1465

OK.1467

Now, using Sanger’s FTNB, like we talked about at the beginning of this lesson, we know the N-terminal amino acid.1469

That information, that info gives us our first fragment, gives us our first fragment.1499

In other words, let say I come up with these four fragments here, when I break it up with my first cleavage enzyme and let say I run a Sanger procedure on these fragments and I end up discovering that the N-terminal happens to be some amino acids, well, let’s say it ends up over here, well, that automatically tells me that this fragment is my first fragment.1511

That is the one that goes on the far left, so that's why this is important.1536

I hope that makes sense.1542

And again, if it doesn't, it's ok.1544

We are actually going to be going through a process, going through in detail what it is that we're actually doing here for a specific example.1546

OK.1555

Now, let's go back and talk about breaking the disulfide bonds, how we do that chemically, and then we'll go ahead and run through an example of a protein sequencing bifragmentation.1557

All right.1568

Breaking disulfide bonds.1571

OK.1581

I'm going to list 2 procedures for breaking disulfide bonds.1583

OK, and recall what a disulfide bond is.1599

It is a covalent bond between these sulfurs on cysteine residues along a protein chain.1606

They don't need to be next to each other.1633

They can be anywhere, but if there is a cysteine and a cysteine, they'll tend to come together, which helps in the folding process but actually a covalent bond.1634

OK.1645

The first procedure is using a chemical called performic acid, and the chemical structure of performic acid is OOHH.1646

It is a peroxy acid using performic acid.1665

OK.1667

Let's go ahead and do this.1668

Let me draw out…let me do this in blue.1670

I've got N, C, C, and peptide goes this way, peptide goes this way, and, of course, this is CH2, this is S, and this is S.1677

Let me make this S a little bit clearer.1692

So, we have this disulfide bond between two cysteine residues.1695

This is C, this is C, this is O, and this peptide bond goes on, this is N, this peptide bond goes on.1702

This is the bond that we want to break right here.1709

Ok.1713

This is the disulfide bond; that's the one that we want to break.1714

OK.1716

We do that with performic acid, so let me go ahead and draw that.1717

Let me go ahead do H, C, O, O, O, H- this is oxidation that's taking place here.1724

I'm oxidizing this, OK, in order to break it; and when I do break that bond, this bond right here, here is what I end up with.1734

I end up with N, C, C, this is O- - oops, that's not O-, this is - I haven’t broken the peptide bond here.1741

This goes on to a peptide; this goes on to a peptide.1755

I've got CH2, and then I have S, double bond O, double bond O, O-, that is one of them, and then I have O-.1758

I hope I have enough room here S, double bond O, double bond O, CH2, C, C, O, that goes on that way, and this goes on this way.1772

What I've done, I have broken this bond, and I’ve created, I’ve basically added a bunch of oxygens to the sulfur.1784

I have just broken the bond in order to free things up, so that things don't stay attached.1792

Again, because there is this covalent attachment between 2 sulfurs on the cysteine residues, I have to break that; I have to free it up, so that the chain is completely free to be sequenced like a string.1798

I can’t have it folded.1809

I can’t have 2 disulfurs attached to each other.1811

That won't give me anything.1815

This is one way to break a disulfide bond.1817

This particular process is oxidation.1819

What I end up producing here is 2 cysteic acid.1823

This dysfunctional group here, this is a cysteic acid.1831

OK.1836

I guess I can write that: 2 cysteic acid residues.1839

That is it.1850

I have broken it, and now, because they are like this, the bonds are not going to reform.1851

These are fully oxidized.1855

They are not going to actually go back the other way at this point.1858

This is one method using performic acid to break the disulfide bond.1860

OK.1864

Now, the second procedure for breaking the disulfide bond is using a reagent called dithiothreitol followed by iodoacetate.1865

Dithiothreitol looks like this: C, CH...actually you know what, that's fine, I'll go ahead and do CH2, I'll go C, I'll put my H there, I'll put my H there, and I'll go CH2.1887

This is OH; this is OH.1910

This is SH, and this is SH.1912

So, this is the dithiothreitol, and my iodoacetate.1916

Well, acetate looks like that.1920

I'll go ahead and write the acid, iodoacetic acid.1924

It has...I'll put an H there; I'll put an H there.1929

One of the Hs is actually just replaced by an iodine.1935

That is all it is.1938

This is just acetic acid and you have an iodine.1939

OK.1942

Here, chemically it is going to look like this.1943

Let's go ahead and do this one in blue again.1946

We have got N, C, C.1950

We have CH2, and we have S, and we have S.1956

We have CH2, we have C, we have N, we have C.1964

Peptide bond goes that way; peptide bond goes that way, peptide bond, peptide bond.1972

That is the protein chain.1976

This is a carbonyl here.1979

This bond, again, is the...oops...do this in red.1982

Again, the disulfide bond that we are trying to break is that bond.1987

OK.1991

The first thing that we do is we react it with the dithiothreitol, and this process is actually a reduction.1992

OK.2009

When you reduce this, it is going to end up looking like this.2010

I wonder if I have any room here.2015

Yes, that's fine; I should be able to do this.2018

So, we have got N, C, C.2022

It is going to be that way, CH2, SH.2026

What you do is you break it up into cysteine residues again.2031

OK.2037

That is the first part, CH2, C, C.2039

That goes that way, and, so, the peptide, peptide.2044

So, you have broken this bond.2048

That bond is now broken.2050

And now, what you have to do, is you actually have to react this with the iodoacetate, and the reason you do that is because if I did not do that then these 2 will just react again and form the disulfide bond again.2054

So, once I've actually reduced it to the thiol, the 2 individual cystein residues that are not covalently attached, I have to do something to these, so they don't go backwards; because this reaction actually wants to be over here.2076

It would prefer to be this way, not this way.2088

So, I have to do something to it.2091

When I do that, I end up with the following.2093

I'm going to do one of them over here, and I'm going to do the other over here, see what they look like.2095

You are going to end up with N, C, C; and you are going to end up with C, H, S, CH2, COO-.2101

Basically, I have just added an acetic acid.2120

I have attached an acetic acid.2123

I have acetylated it.2125

I have attached an acetic acid to this, so that this S and this S can't react anymore.2127

That is all that I've done.2131

I have created basically 2 molecules of that.2133

The peptide goes on this way; the peptide goes on that way, and, of course, this one over here, you’ve got N, C, C.2135

The peptide goes on this way; the peptide goes on this way.2146

You have CH, you have an S.2150

You have CH2, and you have COO-.2154

That is it.2158

You have taken your protein that is stuck together with a disulfide bond.2159

You have broken that disulfide bond.2164

Now, the protein can unfold, and now, you have a nice straight chain that you can go ahead and sequence.2166

That is what we have done.2172

So, 2 ways of breaking a disulfide bond, you have the performic acid, or you have the dithiothreitol; and these are the reactions that take place.2174

And again, you probably don't have to worry too much about reproducing these.2184

You just have to actually know what happens is that this disulfide bond breaks, you end up forming the thiols; and then here, you go ahead and you acetylate them.2189

You add these groups to it.2200

It is OK if you can't reproduce these, but I did want you to see it.2202

It is very, very important that you actually see these chemical structures over and over and over again.2206

That is how we develop the familiarity with these things because again, there is a lot going on here.2213

OK.2218

Let's see what we can do.2220

Now, let's go ahead and actually follow a process of taking a protein, breaking the disulfide bond, hydrolyzing it, splitting it up into fragments, sequencing those fragments, and seeing if we can arrange it in order.2222

We are going to run through this process, so let's just jump on in.2239

Let's see; let me go to...OK.2245

Here we have this random protein, and I'm going to go ahead and draw a couple of disulfide bonds in here.2249

I don't know if you can actually see, or I wonder if we can actually do it on here.2256

No, that's OK; it doesn't really matter.2266

Let me go ahead and this one in blue.2269

We have this random protein that we start with.2272

The first thing that we are going to do is we are going to break the disulfide bond, and we are going to hydrolyze this protein completely, so we can count the amino acids.2274

So, this is the process we are going to use.2282

We want to sequence this protein.2285

That is what we are doing, trying to find the sequence of this really long protein.2287

Alright.2290

Here is what we are going to do.2292

We are going to...oops...should I do it on this page?2295

Yes, that's fine.2302

The first thing we want to do is we want to break any disulfide bonds.2303

We want to free-up the chains, so it is just one long straight chain.2309

Break any disulfide bonds- that is the first step.2313

OK.2316

The next thing we are going to do is we are going to hydrolyze it.2318

We are going to hydrolyze it completely.2322

OK.2324

We are going to use probably some really, really heavy acid like 6 mono-hydrochloric acid.2325

We want to completely break up this thing, so hydrolyze.2329

We want to separate all the amino acids, and we want to be able to count them.2338

And, we can actually do this with other techniques.2346

We don't need to necessarily go through an Edman degradation for that.2347

We have other techniques for separating individual amino acids, proteins.2350

This is just one thing that we can do, one step that we can do.2356

We are going to count which amino acids are there.2359

This is an extra process that we can run.2361

When we go ahead and do this, break the disulfide bonds, we hydrolyze the protein, in other words, break it up into completely free amino acids, separate the amino acids using whatever technique that we have at our disposal, and then, count to see what amino acids are there, this is what we end up with.2367

Again, this is just an example, protein; and this is the data we end up with.2384

We end up with 5 As; we end up with 2 Hs, 1 R.2390

We have 2 Cs, we have 3 Is; these are amino acids, the A, H, R, C, I, right?2399

The arginine, cysteine, isoleucine, things like that.2405

I'm just using these single-letter designations instead of the 3-letter designations.2409

S, we have 2 of them, D, we have 4 of them.2415

Yes.2423

K, we have 2 of them, L, we have 2, M, we have 2, P, we have 3, R, we have 1.2427

Wait, A, H, R, C, I, S, yes, there we go.2441

And then we have S, we have 2 of them, T, let me do it over here.2446

T, we have 1 of them, V, we have of them, and Y, we have 2 of them.2456

We have taken this protein; we've broken the disulfide bonds.2465

We have completely hydrolyzed it, broken every single peptide bond.2468

We have separated the free amino acids, and we have counted them, and these are the counts we get.2471

So, these are the amino acids, their single-letter designations, and this is how many we have.2475

There is some information that we can extract from this.2481

Let's see what sort of information we can extract from this.2484

OK.2486

Now, this gives us a total of 38 amino acids in our protein.2489

This protein that we now know contains 38 amino acids.2499

We basically take a look at this and see what it is that we have, because now, what we want to do the next step, we want to break this up.2505

We want to take this protein, and we want to chop it up.2516

Well, notice I've got 2 lysines.2517

I have 1 arginine, 2 methionines.2526

OK.2531

If we use trypsin...let me do this in red.2533

Using the enzyme trypsin, trypsin breaks peptides next to lysine and arginine.2540

So, using trypsin we have 2 lysines and 1 arginine, that means we're going to make 1, 2, 3 cuts, right?2553

1, 2, 3 cuts, it’s going to give us 4 fragments.2564

So, using trypsin, we can recover 4 fragments.2567

That is pretty good.2575

OK.2578

And now, if we use cyanogen bromide, cyanogen bromide is that chemical way of clipping a peptide chain next to a methionine residue.2579

So, if I use cyanogen bromide, well, I have 2 methionines, so I have 2 points where I'm going to clip this protein; if I clip it in two places, I'm going to end up with a first, a middle and a third.2592

This one will give us 3 fragments.2609

This should be pretty good.2613

I'm going to go ahead and do a trypsin cleavage, and then I'm going to do a cyanogen bromide cleavage, and then I'm going to compare those fragments, and see if there is an overlap or see what kind of information they give me, and see if I can arrange the final sequence, what is the sequence of all these free amino acids.2614

And again, I got that just by sort of taking a look at this list.2633

There are other proteins I could have used and other reagents, but I notice the arginine, I notice the lysine, I notice the methionine, so trypsin and cyanogen bromide, they just happen to be the ones that I have picked in this particular case.2637

OK.2649

Now, let's go ahead and run through the fragmentation process.2651

All right.2656

Now, the first thing I'm going to do, however, is I want to find out what the N-terminal amino acid of this protein is.2658

So, this protein ends somewhere.2665

There is an amino side, and there is a carboxyl side.2667

I want to know what the N-terminal is.2670

All right.2673

I take my intact protein, and I react it with the Sanger reagent, the FDNB- fluorodinitrobenzene.2674

OK.2690

Hydrolysis separation and then, of course, the N-terminal has been labelled with the FDNB, so N-terminal identification.2696

This is the identi…oops let me write this out…identification.2713

My results are, when I do everything, dinitrophenol glutamate is detected.2724

When I run this process, I label it with FDNB; I hydrolyze it.2742

I separate it, and now, I Identify the N-terminal.2747

It happens to be, the molecule that I detect is 2,4-dinitrophenol glutamate, so glutamate is my N-terminal amino acid.2750

So, E, which is glutamate, is the N-terminus.2760

Now, next step of the process that I take when I actually do my fragmentation, one of my fragments on the left is going to have an E; I know that is my first fragment automatically.2775

I don’t have to worry about that; no matter whether it's short or long, I’ve identified the left hand side of the protein to which nothing else is attached.2785

That is why this process is good.2794

OK.2795

Identified glutamate as the N-terminus, now, I can go ahead and start with the breaking up process.2797

OK.2803

I take my intact protein again.2804

First thing I do is, of course, let me see; let me write it over here, yes, so my first step, I’m going to break the disulfide bonds- that's the first step.2808

OK, the second step.2831

Here's what I'm going to do, now, I'm going to run my fragmentation process.2833

I'm going to use trypsin first.2837

Trypsin cleaves the protein on the carbon side, the carboxyl side of lysine and arginine, OK, an R.2840

I separate the fragments; I sequence each individual fragment, sequence the 4 fragments, and my results are as follows.2857

OK.2876

My T1 fragment - T1 for trypsin - my T1 fragment consists of the following sequence: D, C, V, S, H, D.2877

My second fragment is as follows: L, Y, I, A, C, G, P, M, T, K.2892

I'm doing now the results of this process, added the trypsin; I've cut I up.2906

I've separated the fragments; I've sequined the 4 fragments, the Edman, and these are the results: T3, I have E, A, G, G, H, Y, F, E, D, P, I, D, P, R, that's my third fragment.2912

And, of course, I have a fourth fragment, right?2940

Four fragments because we cut it in 3 places.2944

I have got G, A, A, M, I, L, A, K.2948

Notice, there is my K, that's my lysine.2957

There is my R, that's my arginine; and there is my K, that's my other lysine.2960

So, these 3 fragments are where I clipped, right, because it’s going to clip it next to a lysine or an arginine- the K or an R.2968

These are going to show up at one end of the fragments.2976

Notice, this particular fragment doesn't have a K or an R.2980

Chances are, this is our final fragment, the one at the end.2983

OK.2988

Now, let's sort of take a look and see if we can extract some information from what it is that we've done here and we can; let’s see.2989

Here's our E, remember we said that E glutamate we know that that's our N-terminal amino acid on the protein.3002

So, T3 is our first fragment.3011

Let me write that over here.3014

T3 this is the far left fragment.3016

I know that the T3 is our first fragment because the N-terminal is E glutamate.3025

T1, I can also make a comment about T1.3043

T1 is the far right fragment.3048

It is our last fragment in the order.3051

I've got 4 fragments.3053

I need to place them in some sort of an order.3054

I know that T3 is the first; I know that T1 is the last.3057

I know that T1 is the last fragment because it does not end with a lysine or an arginine.3059

In other words, when we cut this, we're cutting it to the…we have our lysine or our arginine residue, we're cutting it to the right of it, right, because we're cutting, we're snipping the carboxyl side, we're breaking this bond and this bond.3082

That means anything over here is not going to have anything attached to it.3102

There is going to be no lysine and no arginine anywhere because this one doesn’t have a lysine or an arginine that tells me that it's the final fragment.3106

These cannot be the final fragments.3113

I hope that makes sense.3116

If not, just stop and think about what it is that I've just said.3117

OK.3121

Now, we have our first fragmentation procedure.3122

Now, I'm going to redo this procedure, this whole thing, except instead of trypsin, I’m going to use cyanogen bromide to create a new set of fragments that I can compare to the first.3124

OK.3135

Let's go again.3136

Let's do this in blue.3141

Again, our first step is to break the disulfide bonds, and, of course, our second step, let's go ahead and use cyanogen bromide.3142

Now, once we go ahead and we clip it with cyanogen bromide, we're going to need to separate the fragments by whatever separation procedure we happen to have at our disposal in that particular laboratory, separate the fragments, and then we're going to sequence the fragments using our Edman degradation, or again, whatever other procedure you happen to have in your laboratory.3161

Maybe you have some mass spectrometry machines, those work also, sequence the fragments, but in this case, let's just say using Edman, and now, these are the results of this particular analysis.3190

Our C1 fragment consists of the following E, A, G, G, H, Y, F, E, D, P, I, D, P, R, G, A, A, M- that's our first fragment.3203

OK.3226

Notice, here's our E.3227

We notice that immediately, so we can almost automatically say this is our first fragment, C1, C2. 53:58 And remember, cyanogen bromide, it breaks things; it clips right next to a methionine, it clips the carboxyl end of the methionine.3230

We had 2 methionines, so in our long peptide, this protein, when I stretch it out after I’ve broken the disulfide bonds, there are going to be 2 methionines.3250

I know that already from the initial amino acid count, because there are 2 in there, I'm going to clip it in two places; I’m going to break it into 1, 2, 3 fragments, so I normally get 3 fragments.3261

This one is T, K, D, C, V, S, H, D, and our C3 fragment is going to be I, L, A, K, L, Y, I, A, C, G, P, M.3271

These are the results of my analysis.3290

Now, let's see what we've got: something to the right of C1, something to the right of C3.3293

Well, I have my methionine here and here.3307

Well, because I'm cutting it to the right of methionine, right, that's what cyanogen bromide does, I know that there is going to be something to the right of C1.3313

Well, because of these, I also know there is going to be something to the right of C3.3321

So, my results: something to the right of both C1 and C3, but I notice that C1 has the E, which is the N-terminus, therefore, I know that C1 is the far left fragment.3327

I know that.3373

OK.3374

We're almost there.3375

Now, in this case, I notice that I've got 3 fragments here.3377

Actually, what's happened is, in this case, I don’t necessarily need to compare overlaps, but I’m going to go ahead and compare the overlaps just so you see it.3384

Let's go ahead and do that; and then we’ll discuss how it is that this cyanogen bromide procedure actually go ahead and gives me my final answer.3391

I didn't actually need to go through the trypsin procedure, but again, live and learn.3399

You don't really know this in the lab because you are just given a protein and they say sequence it.3402

OK.3408

Now, let's go ahead and write out this sequence, and hopefully I can do it in one line here.3409

So, lets' go E, A, G, G, H, Y, F, E, D, P, I, D, P, R, G, A, A, M, I, L, A, K, L, Y, I, A, C, G, P, M, T, K, D, C, V, S, H, N, D.3414

Yes, so what we have is the following from E all the way to the first R.3455

Let me do this one in black.3460

From E to R, this was our T3 fragment.3464

From here to M, that was our C1 fragment.3472

From G to K, this was our T4 fragment.3477

And from I to G, P, M, T, this here to here, this was our C3 fragment.3484

T, K, from here to here, this was our T2 fragment, and then, of course, our T1 fragment, and then from here to here is, of course, our C2 fragment; and notice, we have a C, and a C, so this is our disulfide bond.3499

Disulfide bond is there between those 2 Cs, and notice the overlap between the T3, T4 and the C1, the overlap between the T4, T2, the C3, and the overlap between the T2, T1 and the C2.3527

Here are our points of overlap.3544

Now, in this particular case, I don’t actually need to compare these fragments, the ones from trypsin to the ones that I got from the cyanogen bromide, and the reason I don't need to do that, the cyanogen bromide gave me 3 fragments.3546

Well, those 3 fragments told me that what's going to be on the right one, what's going to be on the right of the other, and the fact that I knew that glutamate was my N-terminal amino acid, I knew that that was going to be my far left fragment, so that automatically put me, I was already able to arrange this overall sequence based on just the cyanogen bromide.3561

OK.3585

Let me go ahead and write this out, and we will go ahead and close out this lesson.3586

In this particular case, since we already knew from the first hydrolysis, remember when we did our initial count, we knew from the first hydrolysis that we had 2 methionines, and also knew that glutamate was the N-terminus, was the N-terminal amino acid.3591

OK.3657

Since we already knew from first hydrolysis that we had 2 methionine, and also knew that glutamate was the N-terminal amino acid, we knew we would have 2 cleavages, which implies 3 fragments.3658

Well, would you have 3 fragments, 3 fragments are easy to order.3687

If you know the far left one, you know what the other 2 are because you know something has to be to the right of the methionine, and something has to be to the right of the other methionine.3690

Well, that's it.3699

We can actually use the cyanogen bromide just to go ahead come up with the final sequence.3701

However, we're not always that lucky where we have 3 fragments.3707

If we don’t, 4 fragments, 5 fragments, or if something else is getting in the way, then we have to use the other information from the other fragmentation procedure to actually set up an overlap.3712

So, what we do is we take these fragments and we see where overlaps occur.3722

When we find those overlaps, we just rearrange those fragments; it’s just like putting a puzzle together and we come up with our final sequence.3726

So that's it.3735

I know that this appears a little bit complicated.3737

Hopefully, you'll see a couple of these problems in your book and you'll run through them.3739

It does take a while to go through, but it is worthwhile to go through it.3744

I promise you.3748

Thank you again for joining us here at Educator.com.3749

We'll see you next time, bye-bye.3752

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

In our last lesson, we closed off by talking about breaking a protein down to elucidate its amino acid sequence.0004

Now, we want to go on reverse.0012

What if there is a particular peptide or protein that I want to build.0014

Today, that is what we are going to be talking about- peptide synthesis.0019

And, the process that we are going to be talking about is something called the Merrifield process, after Bruce Merrifield who created this back in the late 60's.0022

OK.0032

Let's just jump right on in.0033

OK.0036

Now, let's use blue.0038

We can synthesize small peptides with a sequence of our choosing.0049

Now, if there is a particular sequence that I need, I need the peptide, I can actually do it- the sequence of our choosing.0069

And by small, we generally mean up to about 100 amino acids.0081

We actually do that pretty well.0091

And, we also have methods for attaching these fragments to create larger proteins.0093

Before, we took a large protein, we fragmented it, we sequenced the fragments.0122

Now, we are just going in reverse.0126

We are creating sequences up to about 100 amino acids long, and then, we can take those, let's say, 5 fragments, stick those 5 fragments together, and create a really large protein.0128

This is really, really fantastic.0140

OK.0142

So, what we are going to do, we will describe the synthesis of a peptide on a solid support; and I'll tell you what that means in just a second.0143

And, this is called the Merrifield peptide synthesis.0170

OK.0183

The Merrifield peptide synthesis, so here is the general scheme.0184

OK.0194

Now, this thing that we call a solid support, all these fancy terms, I tell you, you are going to want to cross all kinds of fancy terms in the biology, physics, chemistry, and science in general, and well, I guess they are necessary.0195

I don't know; a part of me always wonders whether they actually are necessary.0212

Why don't they just call things for what they are, but I guess, you know how it is; you have to give things certain names.0216

OK.0221

Solid support, it refers to insoluble polymer beads.0222

Just think of really, really, really tiny, small pieces of plastic- that's all it is.0237

Solid support refers to insoluble polymer beads that have reactive chemical groups attached to them, and they look like this.0241

I guess I can do this in blue; it's not a problem.0270

So, I have this bead, and then in this particular case, the group that I have attached to it.0272

That's fine, I'll do it down here, CH2Cl.0285

This is our polymer bead.0294

This is the bead, and it is insoluble, so it doesn't dissolve.0297

You can actually see it like sand at the bottom of a glass of water, and this our reactive chemical group right here.0299

In this particular case, it happens to be an alkyl halide.0309

Alkyl halide, chlorine, alkyl halide, this chloride, this chlorine is going to be a good leaving group, chloride.0312

So, you can imagine, the chemistry is going to take place right here.0318

Some nucleophile is going to come in, and it's going to displace the chloride.0320

This is the reactive chemical group.0325

Basically, the chemistry takes place here; and basically, this solid support, because we can see it, always see it and keep track of it, we are just going to build the molecule in a long chain all along there, and when we are done, we are just going to pull the beads out, and they are going to have these hairs of peptides.0329

That is all that's happening here.0346

A solid support refers to the insoluble polymer beads that have reactive chemical groups attached to them, really, really tiny pieces of plastic that have these chemical groups attached to them, and the reaction can take place.0347

That is it.0359

And, because they are insoluble, we can keep track of them; we can filter them.0360

OK.0365

So, what we do is we start with the bead, and just add amino acids one at a time; and just add amino acids extending from the bead.0369

That is it.0408

Basically, let's just take our...I only have some bead like that.0409

Well, we add an amino acid, that's one, and then we another amino acid, and then we add another amino acid.0417

We add another amino acid; we add another amino acid.0423

We go as long as we want, and we know what sequence we are going in because it's the sequence of our choosing.0425

When we are done, we split this, we break that bond.0430

There we go.0434

We have the protein that we wanted, and because we are actually just adding one at a time, we can always keep track of this because it is insoluble.0435

We see it; we know exactly where it is, and it is very, very easy to deal with.0440

It's a fantastic, fantastic procedure, and in fact, it is fully automated.0445

You just put the reagents in, and you let the machine do your work for you.0449

You come back a day or two later, depending on how long you want your protein to be.0452

It's done; it's ready to go.0456

OK.0458

Now, OK, in the body, proteins are synthesized.0460

Proteins are built from nitrogen end towards the carbon end.0478

In other words, on a piece of paper, they are built from left to right, from nitrogen to carbon, nitrogen toward carbon.0488

OK.0496

In the Merrifield synthesis, it happens in reverse.0497

In the Merrifield process, synthesis is...I wrote the N and the C in the same place, but now, I reversed the arrow.0506

It goes from the carbon end to the nitrogen end.0521

So, when we build this protein, we are building it from the carbon; we are going to the left, right?0524

Because we are adding this first, and then we are adding something to the left of it, to the left of it, to the left of it, to the left of it, it is in reverse; but again, the final product is what's important, but you should know that in the body, it is actually built from N forward.0532

You are adding amino acids; here you are adding amino acids backward.0546

That is it.0549

You are starting from the carboxyl end, and you're going back that way.0550

OK.0553

Let's go ahead and illustrate the procedure with an example.0555

We are actually going to build a tripeptide.0559

So, let's illustrate the procedure with the synthesis of the tripeptide Ala, Gly, Ser- alanine, glycine, serine.0563

We want to make this tripeptide; how are we going to do it?0593

OK.0596

Well, let's take a look at this, just some things we want to observe here.0597

Alanine is the N-terminal amino acid.0603

That is the N-terminus.0608

Serine is the C-terminus, the carboxyl terminus.0611

So, we are actually going to be starting with serine, and then we are going to add glycine, and then we are going to add alanine; because again, in Merrifield synthesis, we move from the carboxyl end towards the nitrogen end- the opposite of what happens in the body.0615

OK.0628

What I'm going to do is I'm going to give a general schematic of all the reactions that take place, and then I'm going to go back, and I'm going to number these reactions, and then I'm going to go back and go through the details of each of these reactions including the mechanism.0631

Now, I have to warn you, there is going to be a lot of chemical structures flying around all over the page; but it is really, really important to be able to do this, to understand what is happening at the atomic level.0649

It is very, very important0661

It makes all the difference between a marginal understanding and a complete command of your material.0663

This is why I'm doing it by hand, and again, this is why I'm not showing you an actual illustration, and just going through the illustration, because you have the illustration in your book already, and they are wonderful illustrations in the book; and I absolutely encourage you to look at it, but that is passive.0668

You need to be able to reproduce this.0683

You need to be able to write what is happening.0685

Draw the structure, write the reagent, give the product- that is where full command comes in.0688

OK.0695

Hopefully, I can actually fit this all in in one page; if not, it's not a problem.0697

I'm going to give the general schematic, and again, I'm going to go back and go through it in detail.0702

Alright.0707

Here is where we start.0708

I'm going to start on the left here.0711

I'm going to start, remember we said serine?0712

Let me just...where can I put this?0716

I'll put her on top.0718

Again, our peptide we are trying to build is the alanine, glycine, serine.0719

We want Ala, Gly, Ser.0726

We are going to start with serine, add glycine, then add alanine.0728

OK.0731

Here is the process.0732

I'm going to take serine, and I'm going to react it.0734

Actually, that's fine; I can go a little bit further to the right here.0744

I take serine, and I'm going to react it with something called Fmoc chloride; and again, this is a general schematic.0748

I'm going to go back and go through all the details.0757

When I do that, I'm going to end up with: Fmoc Ser.0759

And then, I'm going to react this with the bead, and I'm going to end up getting Fmoc serine attached to the bead.0767

OK.0785

Then, I'm going to react this with trifluoroacetic acid or mild base - either one is fine - and I'm going to end up with serine, and a bead.0786

OK.0808

Now, over here, I'm going to do the reactions in red.0810

This is going to be reaction 1; this is going to be reaction 2.0818

This is going to be reaction 3.0822

Now, let me go back to blue.0824

OK.0826

Over here, now, I'm going to deal with the glycine.0827

I'm going to make it ready.0830

This is going to be coming in this way.0832

I've got glycine.0835

I'm going to react glycine with my Fmoc chloride in order to produce Fmoc-glycine, and then I'm going to react that with something called DCC; and I'm going to end up producing Fmoc-Gly-DCC.0838

Now, I'm going to take this, and this is going to be my reagent for this reaction.0862

It's going to come in here, and what is going to end up leaving is DCC.0869

This is going to react with this, and what I'm going to end up with is the following.0876

3...let me label some reactions here.0883

This is reaction 4; I'm going to call this one 4a, and I'm going to call this one reaction no. 5; and let me go back to blue.0885

So, what I end up is Fmoc-Gly-Ser, and bead.0895

OK.0906

And again, I subject this to my trifluoroacetic acid or base, mild base, and what I end up with is Gly - I really hope I can do this in one page - Ser and bead; and let's go ahead and call this one reaction no. 6.0908

I should be able to go, yes, that's not a problem...oops.0940

OK.0948

Now, let me go back over here and prepare my alanine.0949

I'm going to take alanine, and I'm going to react it with this thing called Fmoc chloride in order to get Fmoc-Ala, and I'm going to react it with this thing called DCC, in order to get Fmoc-Ala-DCC.0952

I'm going to take that.0977

That becomes the reagent for this one.0980

DCC leaves the procedure.0985

This is going to react with this, and what I'm going to end up with is the following.0989

I end up with...I have to write it over here: Fmoc-Ala-Gly-Ser with a bead, and then I'm going to subject this to trifluoroacetic acid or mild base; and I'm going to end up with, let me see, so I've got Ala, Gly, Serine, bead, and then of course I subject this one to hydrofluoric acid, and I'm left with Ala, Gly and Serine- what it is that I wanted.0994

This is my general schematic.1051

Let me go through this again, and then I'll go ahead and go through the details.1052

Let me label my reactions.1056

That is reaction 6; this is reaction 7.1058

This is reaction 7a; this is reaction 8.1063

This is reaction 9, and this is reaction 10.1068

I start with my serine; I react it with something called Fmoc chloride to create this Fmoc protected serine.1073

I react that with a bead; I attach it to the bead, so now, I have the bead, a serine residue and Fmoc.1080

React it with trifluoroacetic acid, I take off the Fmoc because I need to attach the glycine directly to serine.1086

I prepare my glycine reagent; I take glycine.1094

I protect it with Fmoc, and then I add DCC to the right side of it.1097

I take that; I react it with my bead and serine.1101

Now, I have my bead, my serine, my glycine, and I have my Fmoc, DCC has gone away.1104

I subject that to trifluoroacetic acid.1111

I break off the Fmoc, so now, I have the glycine that is free to react.1114

I prepare my alanine residue - alanine, Fmoc chloride, this is Cl - to produce Fmoc-Ala.1117

I take the Fmoc-Ala; I react it with DCC to create Fmoc-Ala-DCC.1127

I use this reagent to react with the bead, serine, glycine.1131

Now, I have bead, serine, glycine, alanine, Fmoc.1134

I do TFA to get rid of the Fmoc, and then I use hydrofluoric acid to break this thing off the bead.1137

I'm left with my tripeptide.1145

This is the procedure.1146

Notice, it goes in cycles.1147

This is why anything that goes in cycles like this, it's perfect for automated procedures.1149

OK.1153

Now, we are going to go through each one of these reactions in detail and mechanisms.1154

Alright, so, here we go.1160

Here is where the structures start to fly around all over the page.1164

Let's hope to God that we can keep this straight.1167

It's actually not that bad.1169

If you can follow this, believe me, you can follow anything.1170

OK.1175

Our first reaction, let me see, what should I do this in?1176

You know what, I think I'll do these reactions in red.1179

Let me go ahead and do.1181

So, we said we started with our serine.1183

OK.1187

We have H2N, and we had C, C.1188

We had that, and we had O-.1197

Now, our serine is going to be CH2OH.1201

I hope that I got my R-group right; I hope that is correct.1204

If not, I hope you'll correct me.1207

Now, our first reaction, reaction no. 1, we reacted this with something called Fmoc chloride.1210

This was reaction no. 1, and I'll go ahead and write the reaction number in blue.1219

Let me go back to the red, and what we ended up with was Fmoc, N, H, C, C, O-, CH2, and OH.1227

OK.1244

Let me say what happened here.1246

We are reacting it with this reagent called Fmoc chloride in order to protect the amino group, and here is the reason why we need to protect the amino group.1248

Well, if I take serine, let me do this in black, if I have H2N, or if I had just any random amino acid, notice, I have electrons here.1258

This nitrogen is nucleophilic.1271

Well, I have this O- here.1273

This carboxyl group is also nucleophilic.1274

The next step I'm going to take is I'm going to react this with a bead.1278

Well, the bead is an alkyl halide.1281

Well, since this is a nucleophile and this is a nucleophile, I don't want this to attach to it by accident.1284

I want strictly the carboxyl group to react with the bead end, to react with the bead.1290

I don't want this to react, so I have to protect it.1296

I have to cover it up, keep it from being nucleophilic, so that the only nucleophile on this molecule is that.1298

This is why I'm actually reacting this Fmoc chloride.1305

I'm attaching a protective group, so nothing reacts with the nitrogen, right, because this is nucleophilic, in other words, carrying a negative charge, and this is nucleophilic.1308

I need just that to react with my bead.1325

I don't want this to just randomly react with it, so I have to protect it.1329

That is what's going on.1332

I hope that made sense.1338

OK.1340

Now, let's go ahead and draw the structure for Fmoc chloride just so you see what it looks like, but I'm just going to refer to it as Fmoc chloride, and it looks like this.1341

We have that, and then we have another one, and then, of course, we have this, then we have CH2COCl.1356

OK.1369

This is an acid chloride; it's a carbonyl chloride.1370

OK.1375

This chloride is going to be your leaving group.1376

Sorry, let me go ahead and put my aromatic, because this is definitely aromatic.1377

OK.1384

This right here, that's the Fmoc part, and, of course, this is the chloride.1385

Just so you know, this stands for 9 fluorenylmethoxycarbonyl, all these names, methoxycarbonyl chloride.1395

Wait, am I...I think I have my structure wrong here.1418

Oh well, it doesn't matter; we are only concerned with the Fmoc chloride.1430

This is what we are concerned with right here.1433

I think I'm missing an oxygen here somewhere, but that's OK.1435

Again, Fmoc chloride, it is the chloride part that we are concerned with.1440

This is where the reaction is going to take place.1444

We are just going to call this Fmoc; it's just a protecting group, and here is how it happens.1445

The mechanism is, these electrons, they attack that carbon, and they kick off a chloride, so now, you have this Fmoc group attached to the nitrogen.1450

That is how it ends up attached to the nitrogen, and this is our final product.1470

OK.1474

Here we go.1479

Great.1482

Now, we have an N-protected serine.1485

OK.1499

Yes, that's fine; let me go ahead and do this in red.1503

Let's take our Fmoc, and let's take our serine, N, C, C.1506

So, that's our serine.1517

Now, let's go ahead and do reaction no. 2.1523

Should I do it vertically, or should I do it...that's fine, I guess I can do it this way.1527

Actually, let me go ahead and do it this way.1534

Now, I'm going to react this with the bead, and I'm going to actually draw out the bead here.1538

Boom, boom, boom, boom, boom here.1544

That is that; we have CH2.1549

We have Cl, and here is what happens.1554

This is going to be reaction no. 2.1556

Let me erase something here and do this over.1569

Alright.1583

Let me put my H2s down here, and then let me go ahead and do my blue.1584

The mechanism is this nucleophilic oxygen attacks that carbon, and it kicks off that.1591

What ends up leaving here is our chloride.1600

This reagent attacks the bead, and now, it's going to be attached to the bead.1604

Now, what we have is the following.1611

Let me go back to red.1614

I have Fmoc; I have N, C, C.1616

This is carbonyl; this is O, and this is, I'm just going to write bead.1630

OK.1636

And this is our serine, so this is CH2.1637

This is OH, and let me go ahead and put my H there.1642

N-protected serine, I react it with the bead; now, I have bead, serine, and I have Fmoc.1647

There we go.1653

Now, I'm going to go ahead and react this with...this is going to be reaction no. 3.1655

Go back to red.1666

This is going to be our trifluoroacetic acid or our mild base, and this is to take off the Fmoc group, to deprotect the nitrogen.1668

So, what I end up with is the following.1679

I end up with, let me just write it as H2N, C, C.1683

There is that; there is that...oops.1691

There is no minus there.1696

I'm actually attached to the bead.1698

Let me see, yes, attached to the bead.1705

Then, of course, I have my CH2, and I have my OH.1708

There you go.1714

Now, I have the bead, and I have my serine.1715

So, this is my serine residue, and that is the third reaction.1718

OK.1726

Now, let's go ahead and deal with our fourth reaction and our 4a reaction- the preparation of glycine.1728

Let me go back to red.1736

Let me go ahead and draw out the...so, we have H2N.1737

This is C, C; this is O-, and, of course, glycine has just a hydrogen there.1745

This is glycine.1753

Now, what I'm doing is I'm reacting this again with my Fmoc chloride to create my N-protected glycine.1760

This is going to produce Fmoc, N, C, C.1769

I have that; this is glycine.1779

There is an H there.1783

OK.1785

This is my N-protected glycine residue.1786

Now, I'm going to subject that to my DCC, dicyclohexylcarbodiimide, and I'll discuss this in just a minute.1790

Let me go ahead and draw the structure.1799

When I react it with my DCC, I end up with the following.1800

I end up with Fmoc on the left; I end up with my amino acid on the right.1804

O, D, C, C, and I end up with the DCC on the right attached to the amino acid.1815

This is glycine.1821

I'll go ahead and put an H here.1823

I have Fmoc; I have my glycine, and I have my DCC.1825

I've prepared my reagent to react with what's on the bead.1829

Let me go ahead and write the fact that this is glycine, and now, let me start talking about what is going on here.1833

Let me draw the structure of DCC, and then I'll show you the mechanism going from this step to this step.1843

The mechanism from here to here is, again, this nitrogen, it attacks the Fmoc, and it kicks off the chloride, so chloride actually goes away.1849

This is reaction no. 4.1865

This is reaction 4a.1868

Now, what happens is this.1871

Now, let me go ahead and draw dicyclohexylcarbodiimide.1872

That's fine; I guess I can draw it here.1878

This is N, double bonded with a C, double bonded with an N, and another cyclohexyl group, something like that.1885

OK.1894

I'm going to go ahead and put just a floating H...not yet...OK.1895

So, this is DCC.1902

This is called dicyclohexylcarbodiimide.1907

We will just call it DCC.1918

Once I've created my N-protected glycine, now, this is no longer nucleophilic that's why I protected it.1919

This is going to be nucleophilic.1927

Let me do my mechanisms in blue actually.1933

This is going to attack that carbon, and, of course, we are pushing electrons, so it's going to push this out towards grabbing some H, and what you end up with, now, I'm going to draw this structure with this thing attached.1942

Let me do it in red just so you see it.1958

I'm actually just going to be writing Fmoc amino acid and DCC, but I wanted you to see the picture because again, this is biochemistry, and it's all about the chemistry, the organic chemistry, the structures.1961

N, C, C, this goes there; this is O.1973

This is going to be C; this is going to be NH-cyclohexyl.1980

This is going to be N, and this is going to be a cyclohexyl.1989

So, this DCC; that is this thing right here.1992

That is that.2000

That's it.2001

Now, we have this reagent.2002

Now, we are ready to go through and do reaction no. 5.2003

OK.2008

We have 4, 4a; everything is good.2013

Yes, everything is good.2017

OK.2019

Now, let's see what we've got.2020

We have our serine attached to the bead.2022

Let's go ahead and do that.2025

Let's go to red because that is what we are doing our reaction in.2027

We have H2N, C, C; that is a carbonyl.2030

That's O; this is CH2.2040

I'm just going to write bead.2044

O, and this is the bead; this is CH2.2048

This is OH.2057

OK.2059

So, this reaction that takes place is going to be the following.2060

Should I go straight down?2064

Yes, that's fine; I can go straight down.2065

Now, I'm going to react it with the reagent that I just made.2069

I'm going to react it with the Fmoc, N, C, C, O, DCC, our glycine, right?2076

So, here is how this one works.2098

Now, mechanisms we do in blue.2101

These electrons, that is nucleophilic, that is going to come around.2106

It's going to attack this carbonyl because it is electrophilic.2115

These electrons are going to pop up here.2120

It is going to form a tetrahedral intermediate.2122

Then the electrons are going to pop back down here, and they are going to kick of this DCCO group.2124

Our final product, what you end up with is the following2130

You end up with this thing on the left of this thing.2133

So, our final product is going to be Fmoc, N, C, C, and then, N, C, C, carbonyl, carbonyl, O.2138

This is attached to the bead.2159

This is there; this is our glycine.2163

This is there; this is our serine, CH2OH.2168

This is what I've made.2174

Now, I have my serine, and I have my glycine to the left of it.2175

That is what I've done.2185

OK.2187

Now, let's stop and take a look at where we are just to make sure we've got everything.2189

OK.2192

Let's go ahead and label some reactions.2193

This is reaction no. 5.2196

And, of course, the thing that ends up leaving from this reaction is DCC, because that is our leaving group.2199

Now, what we do, we are going to subject this to trifluoroacetic acid or mild base in order to deprotect, to take off this Fmoc group.2206

So, what I'm left with now, is my glycine residue, my serine residue, attached to my bead.2217

I've got H2N, C, C, N, C, C.2225

I have a carbonyl on the second carbon, carbonyl on the second carbon.2233

This is attached to our bead covalently.2236

I'm going to go ahead and put that there.2240

I'm going to go ahead and put an H there.2243

Now, I'll go ahead and do my R-groups.2245

This is glycine, and this is CH2OH.2247

That is my serine.2251

I have my bead, my serine, my glycine.2252

Now, I'm ready to start the next process.2256

This reaction was reaction no. 6 in our general scheme.2258

OK, we are almost there.2265

We are more than halfway through.2266

Now, we have to prepare the alanine in order to have it react with this thing, and attach it to the bead; and that is going to be our last amino acid.2268

So, let's go ahead and do that.2276

Let's take alanine.2278

Let's go ahead and do it in black actually.2280

We have got H2N, C, C.2285

We take our alanine which is CH3.2292

We react it with Fmoc chloride, and again, let's go ahead and do the mechanism in blue.2299

This nitrogen is nucleophilic.2305

It will attach that; it will kick that off, chloride leaves.2310

This is reaction 7, and what we end up forming - let me go back to black - is our Fmoc-protected alanine residue.2316

N, C, C, O-, I'll go ahead and attach an H, and this is alanine, so this is CH3.2326

We go ahead and activate the carboxyl by reacting it with our DCC.2336

Now, I go this way, and I react it with DCC.2341

This is reaction 7a, and when I do this, what I end up with is the following reagents: Fmoc, N, C, C.2348

This is H; this is carbonyl.2364

This is O; this is our DCC, and this is alanine.2367

So, this is CH3, we have this alanine.2372

Fmoc-alanine-DCC- this is ready for the next step.2376

Now, let's go to reaction no. 8.2382

OK, reaction no. 8.2387

Now, we have our bead, our bead has a serine and then a glycine.2391

We are going to react that with the reagent we just produced- the alanine reagent.2395

Let me go to red.2399

I've got H2N, C, C, N, C, C, N, C, C.2403

There is a carbonyl here; there is a carbonyl here.2415

There is an O, and there is a bead.2418

Our first residue is serine, that is there.2422

Our second residue is glycine, and I have these electrons on the nitrogen.2427

What I'm going to do now, I'm going to react this with my - let me write this in black - the reagent that I just produced, which is...no, I said we are going to do this in black.2432

Let's hope this cooperates.2450

Yes, there we go.2452

I have Fmoc; I have N.2455

I have C; I have C, O, DCC.2459

This is H; this is my alanine reagent, right?2466

This is my alanine reagent.2472

I'm going to stop writing this down.2474

When it reacts with that, DCC actually leaves.2477

This reaction is reaction no. 8, and here is the mechanism.2484

Let me see, where are we?2491

Yes, yes, here we go.2496

I have blue ink.2503

I'm telling you, this gets kind of crazy trying to keep track of everything, but this is actually the nice part, taking the time to keep track of everything to be able to know where you are, to search through all of these chemical jungle that we are looking through to actually make sure you know exactly which electrons are going where.2504

This is where you have a complete understanding of the material.2521

These electrons come here; they attack that.2524

These electrons go there; they fall back down.2530

They kick off the DCC leaving group, and what you end up with is the following.2532

Let me write this in red.2538

You end up with Fmoc, N, C, C, N, C, C, N, C, C, carbonyl, carbonyl.2540

Those are my points of reference, my carbonyls.2556

I have my O; I have my bead.2560

My first residue on the bead is my serine, CH2OH.2563

My second is my glycine, which is just H.2570

My third is my alanine, which is CH3.2572

There is a hydrogen attached to the nitrogen, a hydrogen attached to the nitrogen, a hydrogen attached to the nitrogen.2576

There you go.2582

I've got my serine.2583

Sorry, I'm not going to actually do this.2587

I've got my bead, serine residue, glycine residue, alanine residue, and now, it is protected.2590

Now, I need to deprotect it and break it off from the bead.2600

That's it- reactions 9 and 10.2603

Now, I'm going to do these as single arrows.2606

Let me go ahead.2611

This first step is going to be the trifluoroacetic acid.2614

This is reaction 9 or mild base- to remove the Fmoc.2619

Once I remove the Fmoc, now, my no. 10 reaction, I'm going to react it with hydrofluoric acid to remove the bead, or actually, because that is the solid support, I'm going to remove the peptide from the bead; and what I end up with is exactly what I wanted, and this one I'm going to do in blue.2631

I end up with alanine, glycine - I never, ever, ever get that right, OK - and serine.2659

Serine is my C-terminus.2672

Alanine is my N-terminus.2678

That is it.2683

I just run through the Merrifield process with mechanism, with structure.2685

I've kept it as Fmoc; I've kept it as bead in order to concentrate just on the structure of this, but running through the same thing.2689

You protect the nitrogen group.2699

You add an amino acid to the bead.2702

You prepare the next group.2704

You protect the nitrogen group.2706

You activate the carboxyl group with the DCC.2708

You react these two.2711

Now, you have something else on the bead.2714

You deprotect the nitrogen group, prepare it.2716

You prepare your third amino acid, react it, protect the nitrogen group, activate the carboxyl group, react it.2718

Now, you've added one more chain, deprotect the nitrogen, prepare it, go to your fourth amino acid- over and over and over again.2725

That is the Merrifield synthesis; it is an automated procedure.2734

Now, what I would like to do is I'd like to actually go ahead and show you how this takes place physically, as far as the reaction vessel that it actually takes place in.2738

So, let me go ahead and just go forward to my picture here.2748

OK.2757

This is a chromatography column.2758

Now, you can do this on a chromatography column, or you can also do it by hand by just having it...well, you know what, let's just deal with the chromatography column.2761

This stationary phase right here, these are our beads.2771

Basically, what you are looking at is you are looking at, just imagine a bunch of really, really, really, really tiny beads- that's it.2778

You've got hundreds and hundreds and hundreds of thousands of beads- that's it.2786

It's like sand basically.2791

It's just like really, really, finely ground sand is what it is.2792

So, you take this column, this chromatography column, which is just a hollow tube that comes in different sizes.2796

It could be really, really thin; it could be really, really thick.2803

Generally, they tend to be pretty small for the Merrifield synthesis, and you just fill it up with a bunch of beads.2806

Well, you remember the process that you went through- the schematic.2814

Let me go back to the schematic.2817

Now, because you are actually constructing this molecule, this peptide, on a solid support, all of your solutions, you just pour them in.2819

The reaction takes places as it filters through and passes through the beads, and then it exits when you want it to exit; and, of course, you can control the flow down here.2829

That's it.2838

So, for example, you create your serine and you react it with Fmoc chloride, so you create your N-protected serine residue, well, now, you're going to react it with the bead but the bead is already in the column, so what you do is you take that solution and you just pour through.2839

That is going to be reaction no. 2, and it is going to go ahead and react, and the serine is going to stick to the bead, and everything else is just going to wash through.2857

That is what makes this great.2867

You don't have to purify anything.2868

All the excess reagents, everything that you don't need, just washes through, you can throw it away.2870

Well, now you go ahead and you pour in your trifluoroacetic acid or your mild base to deprotect again.2876

The reaction will take place as it passes through, passes over all of the beads; and any excess will run off.2882

Now, you go ahead and you create your glycine reagent.2890

You pour it in here, it will react as it passes over the bead, any excess will run off.2894

You put your trifluoroacetic acid, it will run through, run over the beads, and that is what you are doing- stationary phase, it doesn't move.2899

You are literally just washing all of the beads with all of the reagents that you need.2907

The beads just stay there; the reaction takes place on the bead.2913

When you are done, you take the column; you empty out the beads.2917

Actually, no, you don't even have to do that.2920

In the final step, the hydrofluoric acid, you just pour it in, the hydrofluoric acid runs over the bead.2922

It breaks the bond between the bead and the first peptide, and then, of course, you just wash it into a flask; and now, you have your peptide in a flask down here for further processing.2931

That is the Merrifield synthesis.2945

Thank you very much for joining us here at Educator.com.2948

We'll see you next time, bye-bye.2950

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

We just finished discussing amino acids and peptides, and what I thought we would do is just do some more example problems with amino acids and peptides just to make sure we have a complete understanding, give us a little bit of review of the things that we have done before, and just get familiar with it.0004

Let's jump on in.0020

The first example, example 1.0025

OK. Given the following data, answer the questions that follow.0033

Let me do this in blue actually.0059

We have 100mL of a 0.20M serine solution at a pH of 1.60, is titrated with a 2.0M sodium hydroxide solution.0062

We don't care about the sodium; it is the hydroxide that we are concerned about.0098

OK. The pK1 = 2.21.0100

We have a pK2, which is the pKa of the amino, that is equal to 9.15; and then I'll go ahead and give you the molar mass.0108

I'm just going to write m/m for molar mass; m/m and this happens to be 105g/mol for serine.0120

Now, the first question that we want to ask is "What is the pI of serine?", and draw the correct structure.0131

In other words, draw the correct structure at that particular pH, which is the pI.0155

Well, you have got 2 ionizable groups on serine.0162

You have the carboxyl group, and you have the amino group.0165

So, the pI, that means...let me just draw the general...so you've got N, C, C, that; and you have that.0168

Starting off at a pH of 1.6, these are both going to be protonated.0180

Well, so this serine...sorry, I'll go ahead and just do R there; we are not concerned with this serine yet.0184

This is carrying a +1 charge.0192

A pI is the isoelectric point.0194

It is the point where the net charge on the molecule is going to be 0.0196

So, the pI is going to be where the carboxyl group is full.0201

In this particular case, since you have 2 ionizable groups, it is where it is completely ionized.0206

There is no H left on there at all because then the carboxyl group is going to carry a -1.0210

The NH3+ is going to end up carrying a +1 for a net charge of 0.0217

That is the isoelectric point.0224

Well, with 2 ionizable groups, the isoelectric point takes place at the arithmetic mean of the 2 pKs.0225

So, you just add the 2.21, 9.15 and divide by 2.0232

We have pI...oops, not a capital P; this is a small P.0238

The pI, the isoelectric point is equal to pK1 plus - I'm always, always, always doing capitals - pK2 divided by 2.0245

2.21 + 9.15 / 2 gives us a pI of 5.68.0260

There we go.0270

When the pH is 5.68, that means this serine molecule has a net charge of 0.0271

The carboxyl is ionized; the amino is not ionized.0278

OK.0281

Now, let's go ahead and draw the structure of serine.0283

We have got the N; we have C, and we have C.0285

We have our ionized carboxyl group; we have this one which is not ionized.0292

Let me make my carbonyl carbon a little bit better here.0297

And then we go ahead and we have the H, and then, of course, we have CH2 and OH.0300

This is our structure at 5.68.0306

That is it. OK.0310

Let's see, part B.0316

Excuse me.0320

There we go.0324

Our pen didn't work there for a second.0327

Alright.0328

How many milliliters, specifically, of the sodium hydroxide solution, of the NaOH solution, have been added at this point, in other words, at the pI?0329

OK.0355

So, we started off with a pH of 1.6, and now, we are at a pH of 5.68- that is the isoelectric point.0357

We want to know how many milliliters of this 2M sodium hydroxide solution we've actually added to bring it to that point.0363

OK. Let's think about this.0371

First of all, we want to know how many moles of serine there is in there because it is going to depend on how many moles of ionizable groups that we have to titrate because that is what the hydroxide is doing.0374

The hydroxide is going in there, and it is pulling off the hydrogen from the carboxyl group; and when that is done, that is going to be the isoelectric point, and then it is going to go and start pulling off the hydrogens from the amino group until it's done.0386

That is the second plateau on the titration curve, which we will draw in just a minute.0402

Let's find out how many moles of serine we have.0405

Well, we have 100mL of a 0.2M solution.0409

Let me do this one in red.0415

We have 0.100L x 0.20mol/L.0418

This will give us 0.02mol of serine.0426

Well, serine has 2 ionizable groups, but 0.02mol of serine contains 2 moles of ionizable groups, right?0432

You have the carboxyl, and you have the amino.0454

With every amino acid, there is a minimum of 2 ionizable groups per mole of serine, which means that we have 0.04mol of serine cancels, 0.04mol of ionizable groups.0456

Well, the isoelectric point - in this particular case, you have 2 ionizable groups - is when one of them is completely ionized.0478

So, if you have 0.04mol of ionizable groups, when half of them have been ionized, that is your isoelectric point.0484

That means - let me write that out - one of these groups is fully ionized.0496

That is what the pI means- isoelectric point.0513

You have 0.04mol of total ionizable groups.0515

Half of them have been ionized, which mean 0.02mol.0520

Well, since we have 0.02mol of ionizable group that have been ionized, that means, and it's a 1 to 1 ratio, 1 hydroxide per 1 ionizable group, right, is pulling off 1 hydrogen, that means there are 0.02mol of the hydroxide that I have to use.0533

Let me actually, specifically write that.0557

0.02mol of OH- times, and it is 2mol/L, 2mol of hydroxide per liter.0560

What we end up with is 0.01L or 10mL of sodium hydroxide solution.0576

There we go; that's our answer.0588

I hope that made sense.0590

At the pI, we know that 1 group is fully ionized.0592

Well, I need to know how many moles there are, but each mole of serine actually brings 2 ionizable groups, so I have a total of 4 moles that can actually be ionized.0597

Half of them have been ionized at the pI, at the 5.68, so of the 0.04, 0.02mol have been ionized.0608

Well, the 0.02mol that have been ionized comes from, they have been ionized by 0.02mol of hydroxide because it is a 1 to 1 ratio, OH + H.0616

That is what forms the water.0626

This is a titration, the acid base titration.0627

That is where this comes from.0630

I take the 0.02mol of hydroxide; I multiply by the reciprocal of its molarity, and I get its volume, so, 10mL of NaOH are added.0631

That tells me that in order to fully ionize the serine completely, I would just add another 10mL, so I would need 20mL of sodium hydroxide to completely ionize it.0639

OK. Let's see, part C.0650

I hope you don't mind that I'm jumping around from color to color.0656

OK.0658

Actually, you know what, I'm going to go ahead and do this one in...I'm going to go back to black- part C.0663

Excuse me.0673

At what pH is the average charge on serine -½.0674

OK, we want to know the pH where the average charge on the whole molecule is -½.0690

OK. We have 2 ionizable groups.0696

Let me go ahead and just draw this out really quickly.0699

We have N, C, C, O-.0702

I'm just going to put an R1 here.0707

Actually, let me go ahead and put the H+.0712

OK. When both of these are protonated, the molecule is carrying a charge of +1.0716

At the pK1, right, the pK1, that is when half of the carboxyl groups have been ionized.0723

The other half is still protonated, so half protonated, half deprotonated.0730

At that point, this side is carrying a charge of -½, right?0733

Because if it were fully ionized, it would carry a charge of -1, but at the pK1 - remember we said that's when half of the groups are ionized - the protonated and deprotonated form are in equal concentration.0740

So, instead of -1 total, just a solid -1, you get a -½.0754

Well, -½ + 1 gives you a +½, and then when this is fully protonated, that is the pI, then you are going to have -1 and +1, that is going to be 0.0758

Now, at the pK2, that means half of these are going to be ionized.0773

This side of the molecule is going to carry a charge of +½, and this side is already fully ionized because we’ve passed the pK1 mark, that is a -1 charge.0781

So, -1 + ½, that gives us the negative half.0791

The pH at which the total serine molecules carrying a charge of -½ is the pK2- the 9.15.0795

Now, let me go ahead and write that out.0804

At a pH = 1.60, that implies that the charge on this molecule is +1 because nothing has been deprotonated.0808

OK. At the pK1, that implies the -½ + 1 = +½.0817

That is the charge at the pK1.0828

Well, at the pI...I didn't even write it.0832

Actually, you know what, it doesn’t matter; I'll just go ahead and do that here.0839

That is going to be -½ + 1 = +½ and then at the pI, we have, of course, -1 + 1, we have a charge of 0, and now, the pK2.0843

We have the -1 from the deprotonated carboxyl group, and then we have +½ from the half-deprotonated amino group, and that gives us a +½.0857

At a pH that is equal to the pK2; that is equal to 9.15.0869

At that pH, the serine carries a +½ charge.0875

That is it; I'm sorry, -½ charge.0880

-½- that is the one we are looking for.0882

OK, part D.0885

Draw the titration curve for this situation.0892

OK.0907

Well, we know how to do this, not a problem.0908

So, we are going to do this; we'll do something like that.0910

OK. Let's mark off some, let's do 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, I'll just go ahead and put 11, and then, so let's see, this is 1.0916

We said we started at about a 1.6, 2.12.0930

So, this 2.12, that is the pK1, and we said that the pI is 5.68.0937

I'll do 1, 2, 3, 4, 5, 5.68, so the pI is somewhere around there.0947

This is the pI.0953

You know what, let me write what these are here.0955

This is pK1; this is 2.12, 5.68, and then 9.15, 6, 7, 8, 9, 9.15.0959

So, this puts our pK2 right there, and this is 9.15.0972

Here we go.0978

We are going to start right about here.0980

We are going to plateau out.0982

We are going to plateau out, and we are going to go up, and then we are going to plateau again, and then we are going to go up.0987

So, it looks something like that.0993

That is our pK1 mark; this is our pI, the isoelectric point.0997

This is our second dissociation, the pK2, and that is it.1002

It is at this point that we've added the 10mL of sodium hydroxide.1007

We have fully ionized the carboxyl group, and then, of course, somewhere over here, we have added the 20mL.1014

This is going to be mL of OH-, and that is it.1022

The titration curve, that's what it looks like, 2 ionizable groups; you have 2 plateaus.1024

If you have 3 ionizable groups, you'll have 3 plateaus.1029

That is it, nice and simple.1032

This point, this point and this point, those are the important points- pK1, the pK2 and the pI, the isoelectric point.1034

You should be able to go back and forth.1043

If you are given a titration curve with numbers on it, you should be able to say this happens here, this happens here, this happens here; or if you are given a numerical data, you should be able to produce the titration curve.1044

OK.1056

Let's see, E.1058

The 10mL of NaOH added to the solution at the pI, so the 10mL to bring it to the pI is how many equivalents?1063

OK.1088

In general chemistry, we mentioned this thing called an equivalent.1090

We haven't really talked about it very much.1096

In biochemistry, they tend to use it a little bit more.1098

It is not really that big of a deal.1101

You are not going to see it all that often, but equivalents just means how many.1103

So, 1mol of serine, we said, has 2 ionizable groups, right?1110

It has 2mol of ionizable groups.1115

It has 2 equivalents of ionizable groups.1119

Equivalent just means how much OH do I have to add for each H that is going to be deprotonated.1123

In this particular case, in order to get to the pI, I have ionized 1 of the ionizable groups, so I've added 1 equivalent.1130

If I ionized the whole thing, I've added 2 equivalents.1140

That is it.1144

It's just, instead of talking about specific volumes like 10mL, 5.6mL, we just speak of equivalents.1147

We are speaking more globally.1153

A particular amino acid might have 3 ionizable groups.1156

Therefore, in order to fully ionize that amino acid, I have to add 3 equivalents of hydroxide: 1 for the carboxyl, 1 for the alpha-amino, and 1 for the R-group.1160

That is all that's going on.1174

So, in this particular case, 1 equivalent has been added to bring it to the pI.1175

A second equivalent would be added to bring it to full ionization- not the pK2.1181

Remember, the pK1 and the pK2 represent half deprotonation.1187

These are the buffer regions.1194

It is fully ionized here, and then the second one is fully ionized here.1197

That is the second equivalent.1201

This is really, really easy; this is just 1 equivalent, and they will often refer to it that way, qualitatively instead of quantitatively.1203

Quantitative, 10mL- they are giving you a number.1214

1 equivalent is just, they are speaking about how many ionizable groups.1216

That is all they are doing.1221

OK, F.1223

Now, would a serine buffer solution, so let's say I went ahead and had a serine solution, and I created a buffer solution out of it.1228

With a serine buffer solution, the appropriate for an experiment requiring for an experiment that needs to be maintained at a pH equal to 8.7.1242

If I am running an experiment and I need to maintain the pH at about 8.7, would it be appropriate for me to use a serine buffer solution?1280

In other words, I create a serine solution; I add enough acid or base to bring it up to this particularly.1288

Is this a good buffer solution?1294

Is serine a good buffer solution?1297

Well, the answer is yes, and the reason is, well, take a look at the pKa.1298

Where does this 8.7 fall?1303

Well, I need it to be in the buffer region, so it either needs to be here, 2.2 + or - 1, so 1.12 to 3.12- that is the range of a good buffer, or in this case 8.7.1305

Here, the pK2 is 9.15, which means our particular buffer region - let me go ahead and - this right here is going to be from 8.15 all the way to 10.15.1320

So, as long as the pH falls, if I need the pH to be in that range, this serine is actually a good buffer solution to use.1334

That is it.1342

You are just looking at the pKa.1343

That is all you are doing.1345

Note the pI; this is not the buffer region.1347

The buffer region is the horizontal region.1349

The buffer region is actually the part that resists changes in pH the more you add.1351

Notice, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, you keep adding a hydroxide; the pH doesn't go up all that much.1356

That is why.1366

So, in this particular case, experiment 8.7, yes, it falls within the range of the second pK for serine, so serine would be a perfectly good buffer solution to use in this particular experiment.1367

OK.1380

Let's go on to example no. 2.1385

OK.1394

Quantitative analysis reveals that a certain protein contains 0.60% tyrosine by mass.1396

I am going to use the...oh, that's fine; I'll just go ahead and write by mass.1427

Remember m/m, sometimes as w/w, they often say a tyrosine by weight instead of by mass- it is the same thing.1434

OK.1444

Further analysis estimates the molar mass to be about 135,800g/mol, and the molar mass of tyrosine is equal to 181g/mol.1445

This is the information that is given to you.1482

OK.1485

Quantitative analysis reveals that a certain protein, it contains, by mass, 0.60% tyrosine.1486

In other words, all of the tyrosine in there accounts for 0.6% of the total mass of protein.1493

Well, they do a little further analysis, and they are able to estimate the molar mass that is somewhere in the range of about 135,800, give or take.1498

The molar mass is 181g/mol of tyrosine.1507

So, the first question we want to ask you is...and this is the process that you will use.1511

Assuming the protein contains only 1 tyrosine residue, calculate the minimum molar mass of the protein.1520

What we are going to be doing by assuming that we actually have just 1 tyrosine residue, we have the percentage, if we assume just 1 tyrosine residue, we can place a lower limit on the molar mass; and then from there, given this other bit of information, the 135,800, then we can go ahead and find out a little bit more about it.1557

Let's go ahead and do the first part.1580

Well, let's see.1582

Let me do this one in red.1586

You remember, when we speak about a residue, it is the amino acid minus the elements of water.1591

It is minus the elements of water and the elements of water are H2O.1598

What you get is, what you end up with is 181g/mol minus the 18g/mol which is water, so a residue of tyrosine actually is 163g/mol.1607

OK.1626

Because, when we form a peptide bond, when we are forming the protein, it is a condensation reaction.1627

We are removing the elements of water, right, reverse of hydrolysis.1633

A tyrosine residue in a protein is actually missing an OH and an H from the original structure.1637

When we give the mass of the amino acid, it is a181g/mol, but when that amino acid is tied up in a peptide, in a protein, it is actually missing an oxygen and 2 hydrogens.1644

It is missing 18g/mol.1655

So, in this case, 163g/mol is the mass of the residue.1658

Well, we take the 163g/mol, divided by the total protein mass, times 100, and they already told us that this is equal to 0.60, because it’s 0.60%m/m tyrosine, part over the whole, tyrosine over the whole; and when I solve for the mass, I end up with 27,166g/mol.1663

This is the minimum per mole.1692

This is the min. molar mass, and it is based on just using 1 tyrosine residue.1696

Well, interestingly enough, I already have this, an estimation of the total molar mass, the 135,800.1702

Well, if I know that I have a minimum mass of 27,166, now, my second question to you is the following.1709

How many tyrosine residues are there in this protein?1718

Well, I have the minimum for 1 tyrosine residue- that is the relationship.1733

I have the total, so I'll just take the total, divided by the minimum, and that will give me, hopefully, somewhere near a whole number.1740

That is how many residues I have.1748

I hope that makes sense.1750

I’m going to do that on the next page, running out of room here.1751

Actually, you know what, I should be able to do it here; that’s fine.1755

I've got 135,800; that is the total estimated mass of the protein.1759

Based on this 0.6% for 1 residue, I've got 27,166g/mol, and when I divide that, I get about 4.99; so you are looking at 5 tyrosine residues.1768

There you go.1783

You use the basis of one to find the minimum, and then you work up; you divide it by the total.1785

This is not altogether different than the empirical formula, molecular formula calculation, that you do in general chemistry.1790

OK.1798

Let's go ahead and do another example here.1801

Let's go back to blue; I like blue a lot.1804

OK.1810

This is example no. 3.1811

Now, a peptide called methionine enkephalin was subjected to the following procedures with the following results.1815

OK.1853

We are going to ask you to take these results, take a look at them, and deduce the structure of the methionine enkephalin.1854

OK.1876

The first procedure that it was subjected to was 6M hydrochloric acid hydrolysis.1877

Oops...hydrolysis, there we go.1884

We basically just completely broke every single base, released every single free amino acid.1886

Let's see.1895

6M hydrochloric acid hydrolysis and we ended up with the following results: methionine to glycine to phenylalanine to tyrosine, the molar ratios were 1:2 to 1:1.1896

OK.1915

We were not able to count how many of each, but we were able to get the ratios of the amounts of the amino acids in this particular analysis, so 1:2 to 1:1, Met to Gly to Phe to Tyr.1916

OK.1928

The other procedure we subjected it to was FDNB and hydrolysis.1930

I should write FDNB then hydrolysis, then we broke it up - fluorodinitrobenzene, that is the Sanger reagent - and we ended up with the following.1942

We ended up with DNB-Tyr, this molecule's derivative of tyrosine was detected, and also, there were no free tyrosines detected.1959

OK.1985

Well, now, our third procedure, we had fragmentation.1987

We did fragmentation by pepsin, and pepsin breaks up the aromatics on the amino side.1995

OK.2013

And what we ended up was with the following.2016

We ended up with a dipeptide containing Phe and Met, and we ended up with some Tripeptide.2018

We did not sequence this; we did not know the sequence.2037

We just know that this dipeptide contains Phe and met; we don't know which one actually comes first.2039

And a tripeptide containing tyrosine and glycine in a ratio of 1:2.2044

OK.2059

We need to deduce the structure, in other words, what's the amino acid sequence.2060

OK.2066

Well, let's see what we've got.2067

Let me go ahead and go to the next page here.2070

Let me see.2074

Our glycine is the one that's 2, so I'm going to write the glycine first.2075

Glycine to methionine to phenylalanine to tyrosine- that is going to be the 2:1 to 1:1.2086

OK, so I've got this.2095

Well, they said that the FDNB gave me a DNB-Tyr.2097

OK.2108

This tells me that the tyrosine is the N-terminal residue.2109

OK.2122

So, we know that it is tyrosine.2123

Well, let me see.2124

Here is something else they said.2125

They said that there was no free tyrosine after we actually hydrolyzed it, once we reacted it with the FDNB.2128

Remember, the FDNB attaches to the N-terminal, and then once you break it up, everything comes apart, we detect this.2134

We have labeled it.2142

We detect that.2143

The others, we can also check to see what's in there, but there was no free tyrosine.2145

OK.2151

And, since there was no free tyrosine, that means there was only 1 tyrosine residue, and it happened to be the one on the end.2153

So, remember, we labeled it, the FDNB can only react with what's on the end.2180

Once that reaction is done, that's when we break up the protein, and now, you have a bunch of free acid.2188

Well, if you can have a bunch of tyrosines that are also free in addition to the one on the left, but there is no FDNB in there, so it is not going to react with those.2193

That is the whole idea.2201

It reacts with the last one first, and then if you have any other free acids, then that tells you how many that you have, but in this case, there was only 1 DNB-Tyr that was detected, but there were no free tyrosines, which means there is only 1 tyrosine, so this 2:1 to 1:1 is not just a ratio, it is exactly how much we have.2202

We have 2 glycines; we have 1 methionine, 1 phenylalanine, and we have 1 tyrosine.2221

This is going to be a pentapeptide.2228

In other words, we have 5 amino acids that make this up- really, really nice.2232

We know what the far left one is; it's going to be the tyrosine.2235

OK.2239

Now, let's go ahead and take a look at our relationship.2240

Now, pepsin cleaves, like we said, pepsin cleaves the phenylalanine, the tyrosine and tryptophan, but we don't have to worry about tryptophan on the left.2245

When it cleaves it on the left, that means, in other words, if you have, OK, if this is either phenylalanine or tyrosine, it is going to break it right there.2260

So, one of your fragments is going to have a phenylalanine or a tyrosine on the left.2277

One of the dipeptides, they said, contains phenylalanine and methionine.2282

Well, we know that that dipeptide has to have a phenylalanine on the left.2286

So, we know that we are looking at Phe and Met- that is our dipeptide.2291

Well, we also know that tyrosine...there is also a tyrosine and a glycine on a 1:2 ratio.2297

Well, I have already accounted for the phenylalanine and the methionine.2307

I know that tyrosine is on the left, so that I know that I'm looking at this- Gly and Gly.2310

I know there is nothing to the left of the tyrosine because that is the N-terminal amino, therefore, all of this information points to the following: Tyr, Gly, Gly, Phe, Met- tyrosine, glycine, glycine, phenylalanine and methionine.2318

This is the structure or the sequence of methionine enkephalin.2343

There you go; I hope that made sense.2350

You are just, sort of, putting pieces of the puzzle together.2353

There is no one way to do this; you just have to use your intuition, use the things that you know one piece at a time, put it together.2358

This one was reasonably simple because we are dealing with a pentapeptide, not going to be so simple all like that.2367

Other times, you are probably going to have to use a couple of cleaving procedures to see where you have overlaps; and, in fact, that is what we are going to do next.2373

OK.2382

Let's go ahead and take a look at another example of a sequencing of a peptide, but this one a little bit more complicated, a little bit longer.2384

Let's see what we've got.2394

OK.2397

This one I wrote out because there is a lot more analysis going on.2398

Let's take a look.2402

Glucagon is a peptide hormone that is secreted by the pancreas in response to low levels of glucose in the blood.2404

It induces the liver to convert glycogen to glucose - glycogen is a carbohydrate, glucose is a carbohydrate, glycogen is made of - and release it into the bloodstream.2411

Glucagon was subjected to several analytical procedures with the following results; use these results to deduce the - I'm sorry - amino acid sequence of glucagon.2421

So, glucagon just does the opposite of what insulin does.2431

If the blood sugar gets too high, insulin is released.2435

If the blood sugar gets too low, glucagon is released.2439

It's a way of maintaining the blood sugar level at some stable level, hopefully.2441

OK.2448

In this particular case, we did a 6M hydrochloric acid hydrolysis of the whole thing, and an amino acid separation, so we were actually able to count the number of amino acids.2449

Here are the results of the hydrolysis and the counting.2460

Histidine, serine, Gln, Gly, Try, Asn, Phe, Asp, Tyr - these are all the numbers that we have.2467

OK.2473

Now, let's see what other analyses.2475

So, we did an FDNB, and we ended up with the DNB-His.2480

OK, that's good.2485

We have that; we know that histidine, and we noticed that we have the one histidine.2486

In this particular case, that 1 histidine happens to be the N-terminal, so that's some good information.2493

We did a couple of fragmentations on this using 2 different enzymes.2498

We used the Asp-N protease, and what it does is that fragments that cleaves the Asp, the cleaves of the protein to the left of the Asp, and that is what the N means, the amino side.2503

So, when we fragment it, the fragment is going to start with an Asp.2515

Let's see here.2524

Fragmentation and Edman sequencing gave the following fragments.2526

OK, so, in this particular case, we not only fragmented but we also sequenced it; and we came up with these fragments.2529

Here, we are just using the single letter designations for the proteins, and don't worry about them if you still haven't memorized them.2535

I mean, at some point, you are going to have to; but no big deal for right now.2540

This is the first fragment; that is the second fragment, the third and the fourth.2545

So, this A1, A refers to the aspen protease, then we go ahead and take an intact protein; and we subjected it to a second fragmentation procedure with trypsin.2549

Now, trypsin, tends to break lysine and arginine.2560

It cuts them at the lysine and arginine residues, and it cuts the carboxyl sides, so to the right of it.2567

In any particular fragment, you are going to end up having an arginine or a lysine at the end.2573

OK, so let's see what we've got here.2580

And thus, fragments gave us this, and this, and this, and this.2582

Excuse me.2588

Well, OK, so, let's see if we can put this together; and let's see if we can find some overlap between these fragments and these fragments.2590

Let's see what we've got.2600

Let's see what we can do.2603

Alright.2605

Well, we know that this is what we know.2606

We know that the DNB-histidine means that histidine is the N-terminal residue.2609

Yes, so that is nice.2619

It means that His is the N-terminus of this particular protein, which is really, really great.2621

And, we have a couple of the fragments, so let's see.2630

A1 and if I take a look at T3, I notice that T3 is the one that actually has the H on the left, so we know that that fragment goes first.2634

That is fantastic.2645

We have already taken cared of about ¼ of this.2646

Let me go ahead and list this as...so, I've got H, S, E, G, T, F, T, S, D, Y, S.2650

This is the T3 fragment.2668

The T3 fragment goes there.2671

Now, let me go to blue.2673

OK.2676

I take a look at some of the fragments on the A side, and I notice that this thing, this T3 overlaps A4.2677

I'm going to go ahead and put the overlap A4 right underneath.2690

So, it is going to be D, Y, S, and then K, Y, L.2693

OK.2700

Now, let me switch colors again.2701

Now, I go back to my T, and see if there is an overlap there, turns out there is.2704

This actually overlaps the T4 fragment.2710

And again, you can switch back and forth and see that this is absolutely correct.2715

In this particular case, I write the overlap this way.2719

I just tend to do it on a stair step fashion, and then just read it off at the end.2722

You can do it anyway you like, however you want to put the pieces of the puzzle together.2725

D, S, R, OK, now, let me see.2731

I take a look at my A fragments, and yes, there is an overlap here.2735

Let me go back to red.2738

This overlaps A3, so I go ahead and write A3 underneath.2742

That is going to be D, S, R, R, A, E.2751

Let me go back to blue.2758

OK.2760

This one overlaps T1.2761

I go ahead and write T1 underneath.2767

I've got A, E, D, F, V, E, W, L, M, N, and T; and this one, it overlaps A2, and that will be my final sequence here, A2.2770

And, of course, that is the...let me do this in black.2797

This is going to be the D, F, V, E, W, L, M, N, and T; and there you go.2802

Now, I just sort of read it off, and just make sure to skip the overlap part.2811

There and there, and there, and here, and I just read it off.2817

I'm going to do this final one in...I guess I'll do it in red, how's that?2827

So, our final sequence is H, S, E, G, T, F, T, S, T, F, T, S, D, Y, S.2831

I have that overlap, so it is going to be K, Y, L, K, Y, L.2842

I have the Y, L, so it is going to be D, S, R, D, S, R, R, A, E; and then, the rest, D, F, V, E, W, L, M, N, and T.2847

There you go.2872

This is the final amino acid sequence of glucagon.2874

Again, hydrolysis- to count2878

Sanger reagent- to find out where the N-terminal is.2881

Let me go ahead and make sure that there is this little mark here.2885

Fragmentation with one enzyme or chemical procedure fragmentation with another enzyme, check for some overlaps.2890

You are just sort of putting it together.2897

Again, I like to put it together in a stair step fashion like this, and then just read it off.2899

There you go, pretty straightforward.2903

There is nothing difficult going on here.2907

It is just a question of putting the pieces together.2909

That's it.2914

OK.2915

Let's see what we've got here.2918

OK.2921

Let's close this off with one final example.2923

Excuse me.2926

Let's go ahead and do this in blue, and hopefully we can fit this in one page, so example 5.2927

I want to create as much room as possible, so I'm going to say...oh, that's fine.2941

Give a schematic for the Merrifield synthesis of the following tripeptide: Gly, Phe, Leu, so glycine, phenylalanine, and leucine; and use only the 3-letter designations.2952

You don't actually have to write out the structure, not a problem.2986

OK, so let's go ahead and do it.2990

Remember, the Merrifield synthesis, we actually synthesize from right to left, from the carboxyl end toward the amino end.2992

So, we are actually going to be starting with- oops, let's do this in red.3000

We are actually going to be starting with leucine, then phenylalanine, then glycine.3005

It is the opposite of how nature does it.3009

Nature goes amino to carboxyl; Merrifield goes carboxy to amino, because we attaching the first one to an insoluble bead.3010

OK.3019

Let's start off over here.3020

Hopefully I can do it, so let's start off with our leucine, and I'm going to react it with our Fmoc-chloride, and that is going to give me Fmoc-leucine; and then, I'm going to react this with a bead, those polymer beads.3024

What I end up with is Fmoc-leucine, and then the bead; and then I'm going to subject this to trifluoroacetic acid or mild base to deprotect that leucine, get rid of the Fmoc.3050

So, I end up with leucine and the bead.3067

OK.3072

Now, over here, I'm going to go ahead and do this in black.3073

Now, I'm going to go, and I'm going to work with my phenylalanine.3078

I'm going to take phenylalanine, and I'm going to react it with Fmoc-chloride.3082

I'm going to protect its amino group, so I get Fmoc-phenylalanine; then I'm going to react it with the DCC, dicyclohexylcarbodiimide, and I get Fmoc.3087

I get the phenylalanine, and I get my DCC, and this is what I'm actually going to take and react, goes in here; DCC comes out, and what I'm left with is Fmoc.3101

I'm left with Phe; I'm left with Leu.3122

And now, it is still attached to the bead.3125

OK.3128

I subject this to trifluoroacetic acid to release this Fmoc.3129

So, I end up with phenylalanine, leucine; and I end up with a bead.3134

Now, I'll go ahead and prepare my second amino acid; that is going to be glycine.3140

I'm going to react glycine with Fmoc-chloride.3146

I'm going to get Fmoc-Gly.3152

Notice that the Fmoc is on the N-side.3154

OK.3156

And then, I'm going to react this with the DCC to activate the carboxyl, so it'll actually react.3157

I end up with Fmoc-Gly, DCC.3164

I take this, and I react it with that; and DCC comes out, and what I'm left with here is Fmoc-Gly, Phe, Leu, and a bead.3171

OK.3193

You know what, I can keep this on 1 page; it's not a problem.3194

Well, nah, that's fine.3197

Let me go ahead and do the next page.3200

We have got Fmoc; we've got Gly.3203

We have got Phe; we've got Leu, and we've got a bead, and we want to go ahead and subject that to trifluoroacetic acid, and we end up with glycine, phenylalanine, leucine, still attached to a bead, and then we wash this with some hydrofluoric acid.3210

We end up releasing that; we end up breaking this bond, the leucine and bead bond, and we end up with our final glycine, phenylalanine and leucine.3236

There you go.3254

That is a Merrifield synthesis of that particular tripeptide.3255

OK.3261

Well, that takes care of the examples for the amino acids and peptides.3262

Thank you so much for joining us here at Educator.com and Biochemistry.3267

We'll see you next time, bye-bye.3270

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today, we are going to start our discussion of carbohydrates, of sugars, otherwise known as saccharides.0004

Today we are going to talk about monosaccharides- absolutely fantastic, fascinating area of biochemistry.0011

I personally, I can't decide which is more exciting.0018

I love proteins, and then when we start doing sugars, carbohydrates, I love carbohydrate chemistry, and then when we talk about lipids, when we talk about enzymes, each area is more fascinating than the next; and it is amazing when all of this starts to come together.0021

Anyway, let's just jump right on in and see what we can do.0038

I have to warn you, there are going to be lots of structures being written out.0042

And again, I don't use pictures; I like to draw everything out.0047

My recommendation, again, I can't stress it enough especially for carbohydrates, because you have lots of carbons and lots of hydroxys.0051

It's one thing to be able to look at an illustration, and by all means, you definitely want to use your book.0060

Your book has fantastic illustrations of most of these things that I'm going to be drawing, but being able to say "Yes, I can see what is going on." is very different from being able to actually produce what is going on with your hand.0065

You want to draw these things out as many as possible, and you'll discover that just by the time you draw your fifth or sixth one, you have a really, really good command of the structures.0079

So, by all means, pictures are great, illustrations are great, but you have to be able to do it with pencil and paper.0090

You have to be able to do it with your hand.0095

OK, enough said; let's jump on in, and hopefully I don't make mistakes, because again, there is lots of carbons and oxygens and hydrogens that are going be floating around.0098

OK, monosaccharides.0109

Well, let's talk about carbohydrates first in general.0110

Carbohydrates are nothing more than aldehydes and ketones with several hydroxy groups attached to the non-carbonyl carbons.0114

That is it, or carbohydrates yield these things; they yield these aldehydes and ketones upon hydrolysis.0163

Do you remember when the lesson, when we did the example problems for peptides and proteins, we talked about glucagon; and we talked about how it induces the liver to actually break up glycogen to release free glucose into the blood?0197

Well, glycogen is a carbohydrate when you hydrolyze it.0212

When the liver hydrolyzes it, it actually releases free glucose, which is the monosaccharide.0215

So, carbohydrate is just that.0222

It's either the monosaccharide itself or the aldehyde or ketone, or it produces those things when you have actually hydrolyzed it.0225

That is all a carbohydrate is.0234

It's either an aldehyde or a ketone that has a bunch of OH groups attached to the other carbons; and you'll see the structures in just a second.0235

OK, most carbohydrates have the empirical formula CH2O.0245

That's where the name comes from- carbohydrate.0266

Hydrate for the H2O, carbo for the C.0269

That's the empirical formula.0273

Now, some carbs - I'll just call them carbs - contain nitrogen, sulfur, or phosphorus.0276

OK.0287

Now, there are 3 major classes, if you want to call them that; I mean, probably not necessary, but we tend to break them up like this.0290

There are 3 major classes of carbohydrates.0299

We have the monosaccharides, which we are going to talk about today, and saccharide just means sugar, the oligosaccharides, and the polysaccharides- long words.0307

Saccharide just means sugar; mono means 1 sugar unit.0335

Oligo means a few sugar units, and poly means a whole lot of sugar units attached to each other.0338

OK, monosacchs are the simple sugars, and consist of single aldehydes and ketones with those additional hydroxy groups attached to the carbons that are non-carbonyl- OH groups.0405 OK, let's go ahead and draw some structures out; let's do some examples.0345

Examples of monosaccharides- I'm going to do a lot of these.0410

OK, there is going to be a lots of structures.0413

I want you to see them over and over again until they are just completely natural for you.0416

OK, let's go ahead and do blue, because I like blue.0422

We have examples of monosacchs, and these are going to be the, I'm going to do the aldehydes first, and they are called aldoses.0430

Aldose, D-O-S-E - that just means carbohydrate, sugar; that is the ending.0442

Glucose, mannose, galactose- they all end in O-S-E.0447

Aldose means all the sugars that are aldehydes that is a broad class.0452

Examples of monosacchs, the first one I'm going to do is one that you've already seen before.0457

It is 3 carbons; I will do the aldehyde group up here, and I think I'll put the H on...that's fine, I'll go ahead and put the H on this side: CH2OH, and I will do the OH on this side, and H.0463

This is 3 carbons; this is the diglyceraldehide.0482

You have seen this one before.0485

Remember when we were talking about protein configuration, L-glyceraldehyde and D-glyceraldehyde?0492

Remember the L-glyceraldehyde had the hydroxide on the left?0498

The D-glyceraldehyde has the hydroxide on the right.0500

That is it.0505

The glyceraldehyde, it is a 3-carbon sugar, because it has 3 carbons.0507

The carbonyl carbon is the first.0513

This is the second; this is the third.0515

Notice, hydroxy attached to the second carbon, hydroxy attached to the third carbon.0517

Also notice that his one has 4 different groups attached to it.0524

So, this carbon is actually a chiral carbon center, but we know that already because of the glyceraldehyde.0527

We have a D; we have an L.0533

Those are the enantiomers of glyceraldehyde, the D and the L.0535

OK, that is a 3-carbon sugar.0540

Well, let's go ahead and do this one.0543

Let's do C, O, H, oops let me draw the backbone first, always a good idea.0547

H2OH, do the aldehyde first; do the last one.0555

Put the H2OH on there, and now, just go ahead and attach the hydroxys.0558

An hydroxy over here, and a hydroxy over here, we will put an H.0563

We will put an H, and this is D-erythrose.0567

And again, these are just examples; you don't have to know all of these.0572

There is only a couple that you are going to have to know.0575

Well, you are certainly going to have to know the glucose, and maybe galactose and mannose, but don't worry about that.0577

I just want you to get comfortable with what's going on here and how these are drawn.0583

This is a 4-carbon sugar.0587

1, 2, 3, 4, hydroxy, hydroxy, hydroxy, attached to the non-carbonyl.0591

This is the aldehyde group, the carbon double-bonded to the oxygen, hydrogen.0598

Carbon, double-bonded oxygen, hydrogen.0603

That is it; nothing actually changes.0605

The chain just gets longer; that is all that's going on here.0607

OK, let's go ahead and do a...this is 1, 2, 3, 4, let's do a 5-carbon sugar.0610

This is going to be 1, 2, 3, 4 and 5.0620

I go ahead and put my aldehyde group up there.0625

I go ahead and put that down here, and then, I go OH, OH, and OH.0628

This is the, 1, 2, 3, 4, 5; yes, this is D-ribose.0639

This is a 5-carbon sugar.0648

OK, now, let's go ahead and do a 6-carbon sugar- probably our most important one.0654

Well, not probably- our most important one.0659

OK.0662

Actually, you know what, let me go ahead and put the Hs here.0665

And again, if you ever see a carbon that is missing a bond, the last bond is going to be an H.0669

Sometimes I forget the Hs, sometimes I don't.0676

Anyway, 1, 2, 3, 4, 5, and 6, let me put the aldehyde group up there.0680

Let me put my H2OH up here.0692

Now, glucose happens to be OH here, OH here, OH here, and OH here.0694

This is D-glucose.0703

This is a 6-carbon sugar.0709

OK, now, let's make some observations here.0713

All have aldehyde groups.0717

They are called aldoses.0731

These are all aldoses because they have the aldehyde, aldehyde, aldehyde, aldehyde.0736

Notice, the only thing that happened is the chain got longer.0744

That is that.0749

The carbonyl carbon, the carbonyl-C, is no. 1.0751

So, 1, 2, 3; this is 1, 2, 3, 4, 1, 2, 3, 4, 5 carbon.0758

This is the 1 carbon, the 2, the 3, the 4, the 5, and the 6 carbon.0767

OK, here is the important part.0774

Again, as with amino acids, the DL system applies.0778

In other words, the reference carbon, the reference C, is the chiral carbon furthest from the carbonyl carbon.0797

OK.0828

If its configuration, I should say if its configuration matches D-glyceraldehyde, then the monosacch is a D-monosaccharide.0832

It is a D-sugar; it is a D-aldose.0858

There we go.0866

So, the reference carbon in this particular case, we are going to be looking at the chiral carbon that is furthest from the carbonyl carbon, and we are going to check to see whether the hydroxide is on the right for D or on the left for L.0868

That is why if you notice, in this particular case, this right here, let's look at the D-glyceraldehyde; this is our reference molecule, so let's not worry about that.0883

This is the chiral carbon; this is our reference.0891

This configuration, if it matches this configuration, the reference carbon for the others, that is what's going to designate it as D or L.0894

In this particular case, here is our carbonyl carbon; this is chiral.0901

This is chiral; this is not chiral.0905

It has 2 of the same thing attached to it- 2 Hs attached.0907

This is not a chiral carbon.0911

So, this is our reference carbon.0913

Well, reference carbon, the hydroxide is on the right hand side.0914

It matches the hydroxy on the right hand side of D-glyceraldehyde; that is what makes it a D-erythrose.0918

If I took this OH and this H and switched positions, put the OH on the left and the H on the right, then it would be L-erythrose because it would match L-glyceraldehyde.0924

In the case of D-ribose chiral carbon, third is a chiral carbon, fourth is a chiral carbon, fifth is not, so this is our reference carbon, sure enough, hydroxys on the right.0935

Notice, on all of these, the hydroxy on the last chiral carbon, the one that is furthest away, is all on the right.0947

Glucose, second is chiral, third is chiral, fourth is chiral, the fifth is chiral, the sixth is not chiral, so this is the reference carbon.0953

The hydroxy is on the right; it makes it D-glucose.0963

If the hydroxy were on the left as written in the Fischer projection, these are still Fischer projections, it would be L-glucose.0965

Notice, these others, I can put these anywhere, and I'll get the other isomers, but we will talk about that in a minute.0973

These are just some examples.0980

What we want to realize here is again, look for the chiral carbon that is furthest from the carbonyl, that is the reference carbon.0982

The configuration at that carbon is the one you compare to glyceraldehyde to see whether it is D or L.0990

We are going to be dealing with D-sugars exclusively, physiologically- D-sugars.0999

Just like for proteins, physiologically, there were L-amino acids, but for sugars, the body uses the D-monosaccharides.1003

OK.1013

Let's go ahead and look at some other examples of monosaccharides, but this time let's do some ketoses, the ones that have ketones- draw out their structures.1016

Let's go ahead and do this in, I think I'm going to do this one in black.1026

We have examples of monosacchs, and this time, we are going to do the ketoses.1033

And again, ketose, they have a ketone group in them- all of them.1043

This is a general class.1046

Well, let's look at the 3 carbon.1049

We have 1, we have 2, we have 3, OK.1052

Because this is a ketone, the first carbon is not going to be the carbonyl carbon; it is the next one.1055

This goes here; this is going to be CH2OH, and this is going to be CH2OH.1063

This is called dihydroxyacetone.1071

The only difference between this and the glyceraldehyde is that now, the carbonyl is not on the first carbon, and the second carbon does not have the hydroxy; they have switched places.1080

The hydroxy now goes on the first carbon, and the carbonyl drops to the second carbon.1091

That is going to be the pattern of ketoses.1095

This is still labeled, though, first carbon, second carbon, third carbon.1098

Now, the carbonyl is always going to be on the second carbon of the ketose.1104

OK.1109

Let's go ahead and go back to black; let's do a 4 carbon here.1112

We have 1, 2, 3, 4; we said the carbonyl is on the second carbon, so let's put that there first.1116

Let's put RH2OH here, and there is an H2OH here.1125

Let's go ahead and stick a hydroxy on the right over there.1132

This is going to be, and let me go ahead and make sure I have my Hs.1136

These are the ones that I always tend to forget, and I also have a really, really bad habit, sometimes, of attaching an H to the carbonyl carbon.1141

Sorry, I hope you are catching those.1150

You have to be really, really careful with these structures; there is a lot floating around.1154

It is not hard; it is just detailed.1158

This is D-erythrulose.1161

And again, the carbonyl is on the second carbon, the hydroxide has gone up to the first carbon.1168

Now, this is the hydroxide; the only chiral carbon in this one is this one right here in this particular case.1174

Again, we have first, second, third, fourth; the second carbon is the carbonyl for ketoses.1180

Let's go to black; let's do the 5 carbon.1190

1, 2, 3, 4, 5, we'll put the carbonyl there.1193

We will put the H2OH here; we will put the H2OH here.1201

We want to set up our frame.1206

Now, let's go ahead and put, yes, let's leave the hydroxys on the right.1209

We have an H, and we have an H.1215

This is D-ribulose.1217

Now, know that the reference carbon is still the same.1223

The reference carbon is the chiral carbon that is furthest from the carbonyl.1225

In this particular case - I'll do this in blue - here, the reference carbon is this.1229

Hydroxy is on the right; that is what makes it D.1235

Here, this is a carbonyl; this is chiral.1238

This is also chiral; this is not chiral.1240

This is the furthest carbonyl; hydroxy is on the right.1244

That is what makes it D.1247

So, that part is still the same; you are still looking for the chiral carbon that is furthest from the carbonyl, comparing that, checking the configuration against glyceraldehyde.1248

And again, we are going to be dealing exclusively with D-sugars of the hydroxyl, and that carbon is always going to be on the right.1258

The other hydroxys, left or right, so the other ones might change.1265

OK, now, let's go ahead and do a 6 carbon.1270

Let's go back to black.1275

We have 1, 2, 3, 4, 5, 6; put the carbonyl there.1276

Put our H2OH here, H2OH on the last one.1285

Now, let's go ahead, and as it turns out, in this particular case, I'm going to go OH, OH.1290

And again, these are just examples; OH, H, H.1300

Now, you are probably wondering why it is that I put this particular OH here, and I left these.1310

Well, you remember, when I did the D-glucose, well, actually, you know what, don't worry about that.1315

Remember, we said the chiral carbons, so first of all, let's list the reference carbon.1323

That is chiral; that is chiral.1330

That is chiral; that is not chiral, so this is our reference carbon; hydroxy is on the right.1333

This is D-fructose, and again, this is the first, the second, the third, the fourth, the fifth, and the sixth.1338

Now, this one we leave alone; these 2 hydroxys, I can actually put them anywhere I want.1349

I can be left, right; this one can be left, right.1355

So, there are 4 possibilities: both left; both right; this left, this right; this right, this left.1359

And again, those sugars do exist.1366

I'm just drawing out examples for the sugars that we tend to run across the most often.1368

That is why I did D-glucose, and here, D-fructose, which is fruit sugar.1373

There is no reason why I put this hydroxy on the left and this hyrdoxy on the right other than the fact that this is the one we are going to run across most often.1381

The other sugars, the other isomers, do exist; and we will draw them out.1391

Don't worry about that, but again, these are just examples.1394

OK, now, let me go ahead and I'm going to draw out the glucose for comparison.1398

This is C, C, C, C, C, C, glucose, the aldehyde.1403

The carbonyl is on the first; let me go ahead and put this here, and we have OH, OH, OH, and OH.1411

This is D-glucose1423

You notice, the only thing that has changed is, the carbonyl has come from this carbon- red.1426

1, 2, 3, 4, 5, 6 - excuse me - the carbonyl has gone from the 1 carbon to the 2 carbon, and the hydroxy has come from this carbon up to this carbon; but notice, everything below that happens to be the same.1435

This structure is all the same; that is the only thing that makes D-fructose and D-glucose different.1461

They have different chemistry; they behave differently, but that is it.1468

That is all that's happened here.1471

On the aldoses, the no. 1 carbon is the carbonyl.1473

On the ketose, the no. 2 carbon is the carbonyl, but the numbering is still the same- this way.1476

OK, just wanted you to see what it is that actually happened here.1481

OK.1486

Let me actually write that down.1490

For ketoses, the carbonyl carbon, the carbonyl C is no. 2; and the reference carbon is still the chiral carbon, the chiral C furthest from the carbonyl.1496

That is the reference C for that one; that is the reference C for the glucose.1527

OK, I hope this is starting to make sense.1533

Again, we are just concerned with some structures here.1536

Let me go back to blue actually; there we go.1547

Let's take a closer look at D-glucose.1555

OK, let's draw the structure again: C, C, C.1564

You can never draw it enough times, trust me on this one.1567

C, C, C, 1, 2, 3, 4, 5, 6, carbonyl goes there; let me put that there.1569

Let me put H2OH here; it is right, left, right, right.1576

That is the pattern for D-glucose.1585

Once you actually draw in the carbonyl on the first carbon, then you put the hydroxy on this last carbon, the H2 which is not chiral, your pattern for the chiral carbons is right, left, right, right, so D-glucose.1586

So, D-glucose1599

And again, there are hydrogens here, but I did not put the hydrogens.1602

That is fine; I'll leave them off, but just know that there are...well, that's OK.1607

That is fine; I'll just put them in.1610

It is probably a good idea.1611

I should not leave them off.1613

OK, alright.1615

Now, notice, it has 1, 2, 3, 4 chiral centers.1617

It has 4 chiral centers, chiral carbons.1626

OK, because it has 4 chiral carbons, it has a total of 24 stereo isomers.1638

Remember what we said.1652

OK.1654

Well, let me say, in general, n chiral centers means 2n isomers- stereo isomers.1659

In other words, I have 4 chiral carbons.1684

That means I have 24, which is - 2 times 2 is 4, 8 and a 16 - I have 16 possible ways that these hydroxide, this 1, 2, 3, these 4 hydroxides can arrange themselves on these carbons.1688

This does not change; this does not change, but here, the hydroxy can be left to right, left to right, left to right, left to right, all right, all left, couple left, couple right, 1 left, 3 right, 1 right, 3 left.1704

All of those combinations contribute to the 16 isomers.1720

D-glucose happens to be one of those 16- that's it.1725

In this particular case, it is the one where you have, on the no. 2 carbon, 2, 3, 4, 5, 6, where the arrangement is the 2 carbon is on the right.1728

3 carbon is on the left; 4 carbon is on the right, and 5 carbon is on the right- that's it.1741

This is one of the 16 stereo isomers for this particular hexose.1745

OK, hexose, 6 carbons, just add that OS.1751

We also speak of pentose, tetrose, triose when we are talking about the number of carbons.1755

OK.1761

Of these 16, 8 are D-hexoses.1763

In other words, they are D in the sense that this final thing, the hydroxy, is on the right.1773

It matches D-glyceraldehyde, and 8 are L-hexoses.1779

In other words, this hydroxy on the reference carbon is on the left.1791

And again, hexose just refers to the number of carbons- that's all.1796

Hexose is just another general.1812

So, the breakdown would be something like this.1815

An aldose, that is the general, aldehyde.1820

Of the aldoses, you have the trios; you have tetrose.1824

You have a pentose; you have a hexose, 3 carbon, 4 carbon, 5 carbon, 6 carbon.1830

Of the hexoses, now, you have your D-glucose, D-mannose, you have D-allose, etc.1837

You have 8 of them, and then, you also have the L-glucose, the L-mannose, the L-allose, etc. - the other 8.1852

I hope that makes sense; what is important is number of chiral carbons.1860

Two to that number gives you how many stereo isomers there are.1865

Now, don't worry, we are actually going to draw all of these out.1869

OK, let's look at all 8 of the D-hexose stereo isomers.1873

OK.1881

Let's do this in black.1884

Let's get this again; there we go.1888

Let's look at...alright...let me see here.1892

Let's look at all the D-hexose stereo isomers.1902

If you have a 6 carbon sugar, of those 6 carbons, 4 of them are chiral centers.1919

That means we have a total of 16 stereo isomers.1924

Of that 16, half are D, half are L.1928

We are not going to be concerned about the L because physiologically D-sugars are what is important.1932

So, we are going to look at the 8 D-hexoses.1938

Alright, let's go.1940

1, 2, 3, 4, 5, 6, OK, this is an aldose.1944

I'm going to go ahead and put my H over there.1954

Now, OH, OH, OH, OH, H2OH, I'm going to leave the hydrogens off.1958

Notice, all the hydroxys are on the right.1969

The particular stereo isomer where all the hydroxys are on the right, this is called D-allose- that's it.1972

Repeat, 1, 2, 3, 4, 5, 6, put my aldehyde group up there; put that there.1984

Now, this one, I'm going to put on the left; but the others, I'm will leave on the right.1998

So, the first chiral carbon, this one, which is the carbon no. 2.2005

All I have done is I have switched the configuration on that one.2012

That gives me D-altrose.2015

OK, now, let's go 1, 2, 3, 4, 5, 6.2020

This time I think I'm just going to put my H on the right; I hope you don't mind.2028

Here, I put them on the left, on the right; it doesn't really matter because it is not chiral.2030

It does not really matter where it goes.2034

I'm going to frame it with my H2OH right there.2037

Now, I'm going to go back to the original2040

I'm going to leave the first chiral hydroxy to the left, and now, I'm going to move the second hydroxy to the left.2043

I'm sorry; I'm going to leave the first hydroxy on the right.2052

I'm going to take the second hydroxy, and move that one on the left, leaving everything else there.2055

This is our D-glucose.2060

That is the one we want: 6 carbons, 1, 2, 3, 4 chirals.2065

The pattern is right, left, right, right, right, left, right, right, as you go down.2073

That is D-glucose.2079

So, D-glucose happens to be the isomer of the 8 D-hexoses where the third carbon happens to have the hydroxy on the left.2081

The other carbons, the second, the fourth, and the fifth, the chiral carbons have the hydroxys on the right.2093

OK.2102

Now, C, C, C, C, C, C.2104

Aldehyde is here; H2OH is here.2111

Now, if I leave the first chiral and the second chiral on the right, this time, if I take the third chiral, I end up with D-glucose.2117

And again, these are all very, very different.2130

I mean, they behave the same way, but they are not the same thing- completely different molecule.2131

The configuration here is different than here.2136

OK, let's do C, C, C, C, C, C.2140

We have our aldehyde group; we have that.2156

Now, I'm going to, so here I have the first one, the second chiral, now, the third chiral.2159

Now, I'm going to go ahead and do the first 2 to the left.2169

Let me put the Hs a little bit closer, OH, but I'm going to leave this one on the right, and this one on the right.2176

This one has to stay on the right because that is the D; that is what designates the D.2185

If I were to do the L-hexoses, I would just move this hydroxide to the left because that is the reference carbon- the fifth carbon.2190

This is D-mannose.2197

Now, let's do this one, C, C, C, C, C, C, H2OH; we have our aldehyde.2203

Now, I'll do that one, and that one.2214

I'll leave that one on the right, and that one.2222

This is called D-idose.2225

OK, we are almost there.2229

Let's go, 1, 2, 3, 4, 5, 6, CH2OH; we draw our aldehyde.2236

Now, we will leave the first chiral on the right.2248

This time we will take the second and the third, and then, we will leave this one on the right.2251

This one is going to be D-galactose- also a very important sugar that tends to come up a lot.2256

And, of course, our last one, C, C, C, C, C, C, H2OH.2265

As you can see, by the time you've actually gone through these 8, you should have a pretty good command of drawing these structures; that is that one.2274

Now, we are going to have all 3 on the left.2283

That one, no, this is not going to work.2290

I need the hydroxys a little bit closer.2296

OH, OH, OH, and, of course, this one stays on the right because that is what designates it.2299

That is a reference carbon that designates it.2306

D...this is talose.2308

There you go.2314

Those are the 8 stereo isomers of hexose- that's it.2315

One of those happens to be glucose.2326

The one that has the pattern, right, left, right, right, for the hydroxys that are attached to the non-carbonyl carbon.2329

That is going to be the important sugar.2338

OK, now, some things to notice; put this in red.2341

Very, very important to notice this.2349

The no. 5 C, the no. 5 carbon, does not change configuration.2354

That one does not change configuration.2365

It is the reference carbon, and it makes these hexoses D-hexoses.2376

If I drew the hydroxy on the left, then, it would be an L-hexose.2395

It might have the same thing, except now, this one is on the left; I’d have another 8.2399

Now, let's go ahead and define something called an epimer or epimer; some people say epimer.2407

Again pronunciation, completely irrelevant.2413

Despite what some people might tell you, it is not relevant.2417

2 monosacchs that differ, if you have 2 monosaccharides that differ in the configuration around 1 carbon, those are called epimers.2421

2 monosacchs that differ in the configuration around one carbon, those are called epimers.2450

Let me give you an example of that.2456

OK, I'm going to draw glucose in the center.2460

So, let's go 1, 2, 3, 4, 5, 6, CH2OH2463

I have my aldehyde; I have my right, left, right, right patterns.2472

So, this is my D-glucose, my primary central monosaccharide.2480

Now, over here, let me go ahead and draw C, C, C, C, C, C, 1, 2, 3, 4, 5, 6, yes.2487

This is CH2OH; this is my aldehyde, and what did I use?2497

I did, oh, I did mannose.2502

OK, this is going to be left, left, right, right.2505

Notice, the only thing that I've changed is that one.2515

This one is left, left, right, right, right, right; everything is the same.2521

The only difference is, at the no. 2 carbon, these differ in configuration.2525

Here, hydroxy is on the right; here hydroxy on the left.2535

This is D-mannose, and it is called a C2-epimer of glucose, or you could say that glucose is a C2-epimer of D-mannose, or you can just say that they are C2 epimers- relative.2538

So, the only carbon that's different is the 2 carbon.2560

Now let’s do another one.2564

Let’s go 1, 2, 3, 4, 5, 6, we have that; we have that.2567

Now, what did I do?2576

I left that one over here; I think I left that one over there.2578

Oh, I changed that one and that one.2583

Now, let me go to black.2588

Now, the only difference is, this one and this, those are the same; this one and this, those are the same.2591

Ah, I changed that one on the 1, 2, 3, on the 4th carbon.2598

On glucose the hydroxy is on the right; on... is this galactose?2605

Yes, that's galactose; yes.2609

On the galactose the hydroxy is on the...this is a no. 4 carbon, so this is a C4-epimer of glucose- that's it.2613

When we talk about epimers, we just mean that, on one of those carbons, usually they will specify which carbon the configuration is reversed, is different- that's it.2627

Now, notice, this does not change the DL.2638

The DL is based on this carbon, the reference carbon.2642

Notice, hydroxy is on the right, D; hydroxy is on the right, D.2647

Hydroxy is on the right, D.2650

It is the one, the carbon that we matched against the configuration of glyceraldehyde to decide whether it is D or L.2655

OK, you are never going to find a 1, 2, 3, 4, 5, you are never going to have a C5-epimer- you're not.2660

I suppose you can talk about it, but we will never talk about a C5-epimer.2670

OK.2674

Now, let's go ahead and get to the, well, get to further elucidation of a monosaccharide structure here.2678

Let’s go ahead and go back to blue.2689

Now, oops, actually you know what, that's fine; I'll go ahead and leave it as black.2693

Now, in aqueous solution, so these monosaccharides, they are very, very soluble.2702

All these hydroxy groups, lot of hydrogen bonding, they are almost infinitely soluble.2709

I mean, you can dissolve a whole bunch, you know that already; you can dissolve a whole bunch of sugar in water, in a very little amount of water.2713

In aqueous solution, in other words, our bodies, aqueous solution, the monosacchs with greater than or equal to 4 carbons, they tend to exist in their ring formations.2720

In other words, if you were to take like D-glucose as a straight chain sugar like this, and if you were to drop this in water, one of the hydroxys and this carbonyl would actually react with each other in an intramolecular reaction; and it will for a ring.2755

In aqueous solution, most of these monosaccharides, they exist predominantly in their ring formation; and we are going to actually talk about, we are going to draw out how the ring forms in just a minute.2775

So, in aqueous solution, monosacchs with greater than or equal to 4 carbons tend to exist in their ring formations.2786

What that means is that 1 of the OH groups on the monosacch has reacted with the carbonyl carbon, has reacted with the carbonyl group, to form a hemiacetal or a hemiketal.2793

OK.2843

Now, some of you may be coming to this biochemistry course having taken only 1 term of organic chemistry; and my guess is that that particular term definitely discussed alcohols, but you may not yet have seen carbonyl chemistry.2846

The chemistry surrounding the carbon oxygen double bond, probably the most important chemistry of organic chemistry, and certainly of biochemistry, the most important functional group.2862

When an alcohol, the hydroxy group reacts with the carbonyl, it form something called a hemiacetal or hemiketal,2874

We will do the chemistry in just a minute.2884

It is not the name that I want you to know.2890

I mean, yes, it is nice to know that it is a hemiacetal or a hemiketal.2892

In other words, when one of the aldehyde, one of the aldoses reacts, you are going to get a hemiacetal.2896

When one of the ketoses reacts, you get a hemiketal.2901

It is the chemistry that I want you to understand.2905

That is what's important.2909

Let's just go ahead and make sure that that is well-understood.2910

The name itself, you might see it occasionally here and there, but it is the chemistry that's important.2915

OK, now, here is the important part.2920

OK.2926

In the process of reacting, the carbonyl carbon is converted to a chiral carbon.2928

So, the carbonyl carbon is not chiral.2953

The double bond, there is no chirality there; but when it reacts, the double bond breaks and now becomes a single bond.2955

Now, you have 4 different objects attached to that carbonyl carbon.2961

In the process of reacting, forming the hemiacetal or hemiketal, the carbonyl carbon is converted to a chiral carbon, because the COO becomes a COH.2969

It becomes an alcohol.2994

Now, it has 2 configurations available to it.3002

Now, that it is chiral, it also has 2 configurations, 2 enantiomers at that carbon, 2 configurations available.3016

We call them alpha and beta.3030

OK.3035

Now, I'll actually go ahead and leave it that way.3037

So, once it reacts, that carbonyl carbon is converted to a chiral carbon.3042

That chiral center has 2 configurations.3047

One of them, we call alpha; one of them, we call beta.3050

Now, let's go ahead and follow this very, very carefully.3052

Let's follow the formation of the 2, actually you know, I'll make sure to write everything out; I mean, I know we know what is going to happen, but OK.3056

Let's follow the ring formation of the alpha and beta configurations of D-glucose.3086

OK, let's go ahead and draw out D-glucose.3106

This is going to be...do this in blue.3110

I have got 1, 2, 3, 4, 5, 6, 2, 3, 4, 5, 6, yes, I have CH2OH.3112

I have my aldehyde group, and, of course, I have right, left, right, right.3123

OK, here is what I'm going to do, and here is how I think about it.3132

When we do the final structures, you can actually arrange it; and you can think about it anyway you want to, but here is how I think about it.3137

I take this vertical arrangement of the glucose, and what I do is rotate it 90 degrees to the right.3144

In other words, I take this molecule, and I just rotate it 90 degrees to the right; then what I do is, I've got the aldehyde on the right.3150

I have got this on the left, then I take this side of the molecule, and I go around to my left from the back, and I attack the carbonyl from the back on the right-hand side; and now, I'll show you what that looks like.3158

Let me go ahead and draw this as...actually, you know what, I'm probably going to need a little room here.3175

I'm going to go COH, and I'm going to go ahead and put the electrons there.3184

C, C, C, C, there is my carbonyl, and I'm going to go ahead and put my H down here.3190

Here, I have got OH; this is right, left.3206

That is there; this is going to be CH2OH.3218

OK, see what I've done.3225

I've rotated this to the right; I've put this carbonyl over on the right.3227

Now, I have taken this group, and I've brought it around to the back.3231

So, this part is the front; from here back, this is actually going back behind the page.3235

I have the aldehyde part, and I take this hydroxy, and I loop it around the back.3242

My carbonyl is here from your perspective.3250

The carbonyl is here, I take this OH group as on the left, and I loop it around behind, so that the hydroxy group is actually coming from behind this way because I want this oxygen on the back and on the right from your perspective.3254

OK, and here is what happens.3268

Well, these electrons, this is nucleophilic hydroxide, right?3270

These electrons, this is an electrophilic.3275

This is going to attack there, and it is going to cause those electrons to move; and it is going to go ahead and grab an H+ from solution, and turn this into a hydroxy.3279

I'm actually going to show that.3289

You know what, I'm going to do this in a different color; sorry.3293

Let me go ahead and put the electrons here.3298

Let me go ahead and do the mechanism in black.3300

These attack the carbonyl, and it goes ahead and it grabs this.3304

Now, here is what happens.3310

You are going to get 2 different structures here.3314

Now, the carbonyl carbon, this is flat.3317

If I have the C and that, this is flat; the carbonyl is flat.3322

This is the carbon, this is the double-bonded oxygen.3331

This hydroxy, you remember, the carbonyl can be attacked from 2 sides, OK.3335

It isn’t attacked that way; it's attacked from the top and from the bottom.3339

If it is attacked from the top, it is going to push the oxygen down.3343

If it is attacked from the bottom, it is going to push the oxygen up, because now, this double bond is turning into a single bond; and it is going to assume a tetrahedral arrangement.3347

It is true that we took it, and we are attacking it from this side; but what is happening is, we are actually attacking it from the top, or we are attacking it from the bottom.3362

That is what's going on.3371

So, you get 2 possible things going on.3373

This one, we will say, this is an attack from above, and this is attack from below.3377

There is attack this way, or there is attack this way.3388

OK.3391

Now, let's go ahead and draw the structures that we end up getting.3394

You end up with the following.3398

I'm going to draw these in black actually, and I'm going to write out all of my carbons because I love drawing out everything.3401

C, C, C, C, O, OH, H, this goes down.3409

This one is up; this one is down, and this is CH2OH.3421

Did I forget anything here?3428

Nope.3430

This is called the alpha-D-glucose; this is D-glucose, OK.3433

The D-glucose part is the configuration of the hydroxys.3436

The alpha part means that it is attacked from above.3442

Now, this hydroxy here is down.3445

If I'm looking at the ring this way.3450

Imagine this is flat; I'm looking at it like that.3454

If the hydroxy is below the ring, that is the alpha-D-glucose.3459

That means it was attacked from above, so it pushed that oxygen down.3462

This right here, that is the carbonyl carbon.3466

OK, this was originally the carbonyl carbon; that one right there.3471

OK, now, that is alpha-D-glucose.3474

If I have attacked from below, so that the hydroxide ends up above the ring, it is going to look like this: C, O, C, C, C, C.3480

I'll make it a little bit more uniform here.3491

C, C, C, this time, when the tetrahedral arrangement is such that this hyrdoxy is above the ring.3494

And again, notice, everything stays the same.3502

That one is down; that one is up.3504

That one is down, and this one doesn't matter.3508

This is beta-D-glucose.3512

The hydroxy is above the ring.3517

OK, hydroxy is up here.3521

Here, the hydroxy is below- that is alpha-D-glucose.3524

OK.3528

Now, here we go.3532

The carbonyl carbon, the carbonyl C, which is this one, which is now a hemiacetal, and all that means is that this was a carbonyl carbon.3537

Now, it has a hydroxy group attached to it, and it also has an oxygen connected to a carbon group.3557

OK, there is an ether function, COC, and there is an alcohol function.3566

Both oxygens are attached to this carbon.3576

That is the carbonyl carbon.3580

We draw it like this, specifically.3582

We put the carbonyl carbon on the right; we put the oxygen on the back right, and we arrange it like this, but I'll talk more about that in just a second.3583

So, the carbonyl carbon, which is now a hemiacetal - and again, hemiacetal means hydroxy group, ether group, hydroxy group, ether group attached to that - is called the anomeric carbon, and the 2 isomers namely alpha and beta.3591

It is called the anomeric carbon.3602

And, the 2 isomers namely alpha and beta, OK.3617

OK. This is the only place that the configuration is different; everything else is the same.3625

OK, down up down, down up down, nothing, nothing, up here, down here.3631

It is the only place, OK.3637

The isomers, and the 2 isomers of the anomeric C are called, well, you guessed it- anomers.3640

Alpha-D-glucose and beta-D-glucose are anomers of each other because the configuration is different only at the anomeric carbon.3657

The anomeric carbon was originally the carbonyl carbon, the aldehyde.3669

There was an intramolecular reaction.3673

So, 1, 2, 3, 4, 5, there was an intramolecular reaction of this hydroxy group attached to the no. 5 carbon that reacted with the carbonyl to form this hemiacetal.3679

Hemiacetal is a hydroxy group attached to that carbon, an ether group attached to that carbon.3691

That is what's going on here.3697

OK.3698

Now, let's go ahead and follow the same thing for fructose.3702

Yes, this one I'll do in blue.3707

I want you to see it again, that's why I'm going to go through it.3709

Let's follow the ring formation for fructose.3712

And again, fructose is a ketose.3726

Let's go ahead and draw it out.3729

Again, we have 6 carbons; we have 1, 2, 3, 4, 5, 6, but this time, we have the carbonyl here.3733

We have the H2OH here; we have the H2OH here.3746

This is there; this is there, and this is there.3752

Again, it is going to be the hydroxy on the no. 5 carbon, 1, 2, 3, 4, 5; let me number them- 1, 2, 3, 4, 5.3758

It is going to attack the carbonyl, but notice, now, we have 1, 2, 3, 4, 5 members in the ring, not 6 members in the ring because now, the carbonyl is not on this carbon, it is over here.3774

Let me go ahead and turn this around so you can see it.3789

Again, rotate it to the right.3790

Rotate the molecules to the right; take this side, and bring it around the back.3794

That is what we want to do; we always want the anomeric carbon to be over here.3800

OK, when I do that, I'm going to end up with the following.3807

Let's go, should I do this in blue?3812

Yes, let's do this in blue.3815

I have got C, C, C, C, and I've got OH, 1, 2, 3, 4; oh, yes, of course, sorry about that.3817

I have got this one over here, CH2OH, and this is my carbonyl; there we go.3838

I got lost for a second there.3843

OK, this one is up; this one is down, and this one is there.3845

Now, again, we have attacked from above, attacked from below.3855

Yes, that is fine.3861

Let me go ahead and draw the mechanism.3863

Let me put an H+ out there; this attacks the carbonyl.3867

This goes ahead and grabs that, and again, we have 2 possibilities.3871

This is attack from above.3879

OK, this is attack from below, and you end up with the following.3889

This time, for the 5-membered rings, we put the oxygen on the top; and we go C, C, C, C, C.3896

And now, if we do attack from above, that is going to push this oxygen down.3909

So, you end up with the hydroxy down here.3916

You end up with CH2OH there.3920

You end up with CH2OH here, and here, we have the OH up; and this is the OH down, and this is alpha-D-fructose, alpha, because the hydroxy is below the ring.3924

Attack from above...wait...that's from above.3944

From below, we are going to push the oxygen up.3948

We are going to end up with the following.3951

Let's go back to black here; put our oxygen there, carbon, carbon.3953

Let me number my carbons, by the way.3960

This is 1, 2, 3, 4, 5, so the hydroxy on the no. 5 carbon again.3963

OK.3972

This time from below, we are going to push the hydroxy up.3975

Let me go ahead and draw my ring first; let me close that one out.3979

This one is going to be CH2OH; this is going to be CH2OH.3982

This is going to be OH here; this is going to be OH there.3990

This is beta-D-fructose.3994

Again, arrange it horizontally, come around.4001

This is 1, 2, 3, 4, 5; the hydroxy attacks the no. 2 carbon.4006

In this particular case, this is your anomeric carbon.4015

Again, it is on your right-hand side.4018

Oxygen is on the top here, top here.4020

It gives you the beta-D-fructose, beta, because the hydroxy is above the ring.4024

When you look at this, you are looking at it like this, but really what you are looking at is - I've drawn it this way oxygen, carbon, carbon, carbon - you are looking at it that way.4029

And, we will actually do a prospective drawing in just a moment called a Haworth projection, but you are looking at it that way.4039

That is what's happening.4044

You have the hydroxy either up here or the hydroxy down here.4046

Hydroxy down here is alpha; hydroxy up here is beta.4048

OK, now, let's finish this up here.4053

So, we have got 6-membered rings.4059

6-membered rings are called...and again, you know, well, let me write down the name- pyranose.4067

One of the most frustrating things about biochemistry for me, personally, has always been the vocabulary.4080

You have got aldose; you've got hexose.4086

You have got pyranose, so 6-membered rings are called pyranose.4089

See, you have got all of these names for the same thing.4092

And again, in any conversation that you'll have with a professor or a student or something like that, anyone of those people is going to use anyone of those terms.4095

So, it is a little annoying to have to have all these terms floating around.4106

It gives the impression that you are talking about a whole bunch of different things- you are not.4110

You are talking about 1 molecule.4114

It is just, all these names that are attached to it depending on what we want to emphasize, and a lot of this is just historical garbage in the sense that this stuff has just, sort of, stayed, and we have used it, and we have used it; and now, we have this build-up of all these stuff from the history of biochemistry that we now have to synthesize, that we now have to bear on our shoulders.4118

All I can say about that is "I'm sorry"'; it is just as annoying to me as it is to anybody else.4137

I never use the word pyranose, but there it is.4142

OK, 5-membered rings, and I'll tell you in a minute why they are called pyranose and furanose.4147

5-membered rings are called furanose, and I can never remember which is which.4157

Is pyranose 6; is furanose 6?4165

Anyway, OK, now, let's talk about something called a Haworth projection.4168

Let's go ahead and do this in black.4178

Haworth projection- this is a way of looking at these sugars in 3-dimensional way.4183

We will do a Haworth projection of the pyranoses, and these are 6 carbons, yes.4192

I'm going to draw the projections, the I will draw the bases of the name pyranose.4201

OK, here is what you've got.4206

This time, I'm not going to draw out all of the individual Cs.4208

I'm going to do it in a line structure.4212

This is going to be O, boom, boom, boom, boom.4215

I want a little bit better than that.4225

OK, let's go ahead and do the alpha, OH, OH, OH, CH2OH.4241

OK, this is alpha-D-glucose.4254

Yes, it is fine; I'll just go ahead and write it- alpha-D-glucose.4261

Notice this particular projection, how we have done it.4266

Remember we said the sugar, so now, it's a ring.4268

You have go this 1, 2, 3, 4, 5, 6; the oxygen is on that side.4270

You are looking at it that way; that is what you are doing.4274

That is what this projection is.4277

The single lines, the normal lines, those are in the back; these bold lines, it comes out as a wedge, and then it stays bold like that.4280

Those are coming out towards you, and what it does is it gives you a way of seeing what is above the ring and what is below the ring.4287

Now, you remember that anytime you have a 6-membered ring, you don't have a flat ring.4294

What you have is a chair and boat confirmation.4299

Won't talk about that right now, I will in the next lesson, a little bit; but this is a really, really great projection because it shows you what is above and what is below the ring.4302

In this particular case, you have the hydroxy below the ring, so you have the alpha-isomers.4310

This is alpha-D-glucose, and notice how this is arranged.4318

The oxygen is on the top right, and the anomeric carbon is on the right.4321

This is why we said, take your molecule, rotate it to the right, make sure the anomeric carbon is right there.4327

The carbonyl, bring this side around the back, and your oxygen will actually end up staying back there.4333

That is the way you want to think about it.4338

Rotate to the right; bring it from the back, and attack above or below to create the ring.4339

OK, this is alpha-D-glucose.4346

Now, let me go ahead and do beta-D-glucose.4347

Again, we have our oxygen, we that, that, that, that.4352

I'm telling you, I don't think they will ever improve drawing these things year after year. 4367 OK, that is that, there, there, there, there.4361

Again, we've got, comes out as a wedge, bold, something like this.4374

Now, we have our beta with a hydroxy above there, and this stays.4382

This is down; this is up.4386

This is down, and this is like that.4390

This is the beta-D-glucose.4393

OK, now, some things, blue.4399

I love jumping around with these colors; it's really, really great.4407

OK, oxygen is at the back right always.4411

OK, the anomeric carbon is on the right.4425

Anomeric carbon, oxygen, anomeric on the right, oxygen, back right.4440

If you want, you can put the electrons on the oxygen, it doesn't really matter.4444

Now, here is why we call it a pyranose.4447

Yes, that is fine; I'll go ahead and do it in black.4457

Well, basically what you have is this.4463

This molecule is called pyran, and if I were to draw it in perspective, it would look like this.4470

OK, this is pyran.4479

It is based on this thing with the hydroxys attached, so they call it a pyranose- that is why.4481

That is where the name comes from.4487

I never cared for it very much, in fact I rather dislike it, but there it is.4490

OK, now, let's do our Haworth projections for the furanoses 5, yes.4496

OK, let's go back to blue, and this time we put the oxygen on the back, but the anomeric carbon is still on the right.4520

Again, we have...you know what, let me start again.4530

OK, we have got O, boom, boom, boom, boom, boom.4538

So, we have got, this comes out into a wedge, and this is a bold line here; and this comes out to a wedge, or you can just make them all bold.4545

It doesn't really matter all that much.4554

We have got an OH here; we have got CH2OH here.4557

In this particular case, that is up; that is down.4563

This is CH2OH; this is the hydroxys down below the ring, right?4570

This is this way; oxygen is back here.4574

1, 2, 3, 4, 5, we are looking at it like this.4577

The hydroxy is down below.4582

This is alpha-D-fructose.4585

I think I have got everything there, I hope, yes.4590

And, now, let's go ahead and do beta, boom, boom, boom, boom, boom.4594

And then let's go ahead and bold this out, bold this out, bold this out.4603

And now, we have the hydroxy on top, the CH2OH below.4609

This CH2OH stays.4614

This hydroxy is up; this hyrdoxy is down.4618

Is that correct?4622

Yes, that is correct.4625

So, this is beta-D-fructose.4627

A lot of structures we're drawing.4632

And, they are called furanoses because of this molecule.4634

This particular molecule is called furan.4641

They consider it a derivative of some sort, just a whole bunch of hydroxys attached to it.4649

So, that is it; that is our introduction to monosaccharides.4654

Thank you so much for joining us here at Educator.com4660

We'll see you next time for a further discussion of carbohydrates.4662

Take care, bye-bye.4665

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today we are going to continue our discussion of monosaccharides.0004

I'll finish that off a little bit, and then we are going to start talking about disaccharides, 2 sugar units attached together.0006

OK, let's go ahead and get started.0013

Before I begin, I wanted to make sure there was one thing, point of clarification from the previous lesson.0018

It concerns the formation of the actual ring form from the linear form of the sugar.0026

I just wanted to make sure that everything was OK there.0033

So, let's go ahead and discuss that before we actually go on.0036

Point of clarification forming a cyclic sugar from a linear sugar.0043

OK, let's go ahead and do a linear sugar.0067

Let me go through the entire process.0069

Remember when I said rotate it to the right, and then grab it and bring it around.0071

I just wanted to make sure that we have everything OK.0075

There is lots of hydroxy groups that can actually react with the carbonyl carbon, with either the ketone or the aldehyde; and I just wanted to make sure which hydroxide is going to be reacting.0077

You know, most of the time, we are going to be dealing with the hexoses, so glucose.0089

I just wanted you to see exactly how we arrange things, how we rotate it, how we pull.0094

I just want to make sure that that's clear.0099

Once that that is clear, again, once the basics are clear, everything after that is absolutely perfect.0102

Let's go ahead and draw out this: 1, 2, 3, 4, 5 and 6.0108

Let's go ahead and draw out our glucose here.0116

We have right, left, right, right- that is the pattern, so CH2OH.0120

Now, what we are going to do here is we are going to go ahead and rotate this 90° to the right from the top.0130

OK, when we do that, we end up with the following.0140

I'm going to draw out the 6 linearly this way, horizontally: 1, 2, 3, 4, 5, 6.0143

Now, the aldehyde group is over here; and now, I have this OH down at the bottom, this OH up at the top, this OH at the bottom, this hydroxy at the bottom, and over here at this end, I have my H2OH.0150

OK, so far so good.0164

Now that I have it arranged like this, I grab the left end.0166

I grab this end and I pull it around to the back to have it come around in a circle and set itself up for attack.0171

Let me go ahead and do that.0179

From your perspective, what you are looking at is now, the carbonyl side is here; the CH2OH is here.0181

I'm going to pull it around to the back and just arrange it.0186

OK, I'm going to draw that now.0190

You know what, I'll stay with black; why not?0195

Let's go that, that, that, that.0200

Nope, we have, of course, this is the carbon; there is this.0204

I'm going to go ahead and put the H there.0207

Now, this hydroxy, this is this carbon.0209

This is down; this hydroxy is up.0212

This hydroxy is down.0216

Now, I've just pulled it straight back, so what I've got here is the following.0219

Let me see.0224

This is down, up, down, down.0226

On the no. 5 carbon...wait...1, 2, 3, 4, 5...yes.0228

And now, I have 1 more down; this hydroxy is down.0235

And, of course, this is the CH2OH.0240

Now, this is the no. 1 carbon; this is 2.0244

This is 3, 4, 5 and 6; 1, 2, 3, 4, 5, 6.0246

It is the hydroxy on the no. 5 that is actually reacting with the carbonyl.0255

Here, when I pull it around, when I'm doing this systematically to form the ring, when I pull it around, I still have to make an adjustment.0260

At this carbon right here, let me do this in red, I have to actually rotate 90° up.0270

Now, the CH2 group which is here and the hydroxy which is here, I need to do this.0276

So now, it is the hydroxy on the no.5 carbon and it is the CH2OH group, that is actually pointing up.0282

That is where we get our glucose structure.0289

I'm going to rotate this 90° that way, at the no. 5 carbon.0291

Let me redraw this ring structure here.0298

Let me go ahead and draw it down here.0300

OK, I'll write "Rotate the no. 5 carbon, so the hydroxy group is now horizontal, and the CH2OH group, which is this thing right here, this thing, the no. 6 carbon is pointing up, is vertical".0305

OK, I just want to make sure that this is absolutely clear.0342

It is not the hydroxy on the no. 6 carbon that attacks; it is the hydroxy on the no. 5 carbon that attacks.0346

Once we rotate it, now, we get this structure.0353

We get this, this, this, this.0356

Let me go ahead and write the C.0361

It's always a good idea to write the Cs.0363

Actually, you know what, I'm going to do this in black still.0367

Let me go back to black.0373

1, 2, 3, 4- this is our aldehyde.0375

Now, our OH is here and now, our CH2OH is up.0381

See here, it is that way.0386

That is just from turning it, pulling it back.0388

Now, we still have to rotate this one in such a way that the hydroxy is now ready for attack.0390

It is the no. 5 carbon, it is the hydroxy on the no. 5 that attacks the no. 1 carbon to form the 6-membered ring, so now, it is ready.0396

It can attack on the top; it can attack from the bottom, and, of course, what you end up getting is our glucose ring.0407

That and I'm going to do a little wavy line here because again, I'm not specifying the stereo of chemistry here.0418

It can be either alpha or beta.0422

If the hydroxy ends up below, in other words, if it attacks from above, it ends up being an alpha-glucose.0424

If it attacks from below and pushes the oxygen up, so that the hydroxy is above the ring, then it is going to be the beta-glucose.0430

So, this wavy line means I'm not specifying the stereo chemistry, but this CH2 group is up there; and of course everything else stays the same.0437

This is down; this is up, and this is down.0445

Now, we have our glucose.0449

I'm not going to be calling it D-glucose anymore, where just the assumption is, the sugars that we deal with are going to be the D-isomer.0451

If they happen to be an L-isomer, we will specifically say L, so I'm just going to write glucose.0460

There we go; that is what you are doing: rotate, pull around, make a little rotation.0466

So, it's the no. 5, the hydroxy on the no. 5 carbon that gives you this arrangement.0471

This is the conventional arrangement- oxygen on the back right.0478

And, let me go ahead and draw the actual projection here- the 3-dimensional projection.0481

We have that like that, and there we go.0487

This is our nice, basic glucose structure.0491

OK, I hope that helped.0495

Now, let's go ahead and move on to a discussion of conformation.0500

Let's recall the alpha and beta anomers of glucose.0505

And again, you can never get enough practice in actually drawing out these structures.0508

I hope you are not getting sick of actually, be drawing them out, drawing the out- it is really, really important.0513

Repetition is what keeps these things solid in your mind- repetition with your hand, in fact.0517

OK, let's recall - excuse me - the alpha and beta anomers.0523

And remember, we call them anomers because the carbonyl carbon is the anomeric carbon- once it has been converted to a hydroxide, once you form the ring, the anomers of glucose.0535

OK, we have 1, 2, 3.0546

OK, let's go ahead and draw the alpha.0550

This is that; this is that.0555

This is that, and CH2OH.0559

Let's go ahead and give a little bit of projection here.0563

OK, this is our alpha-glucose.0567

Hydroxy is down below the ring, and of course, we have our beta-glucose, where the hydroxy is above the ring, but everything else is the same, down and CH2OH.0572

Let's go ahead and put a little perspective on it.0587

There we go, good solid pictures of alpha and beta-glucose.0591

OK, now, these projections give you stereo chemistry, but they don't give you conformation.0599

These tell you exactly where the hydroxys are.0606

Here, it is below the ring; here, it is below the ring, above, below.0610

CH2 is above.0614

They give you stereo chemistry, but you remember from organic chemistry that a 6-membered ring is not a flat molecule.0616

It is not benzene.0621

OK, It achieves, these are the 1 Ns.0624

These Ns are fluked like this, so what you end up getting is 2 chair conformations.0627

When we work with them, we'd like to keep these forms simply because they are easy to see.0631

They are easy to see relationships, but how it really looks is a little bit different.0638

I'm going to go ahead and draw out - just so you see it - the chair conformations of these glucose molecules.0641

Let's see.0652

I'm going to go ahead and just do the alpha-glucose.0654

I said we are not going to be writing the D, so for alpha-glucose.0661

I'm having a hard time spelling today.0668

OK, let's try this again.0669

For alpha-glucose, OK, let's see if we can do this here.0673

Let me go ahead and just do...let me do it over this way.0676

This is that; that is that.0681

That is that; that is O.0684

Let me go a little bit further down here.0687

It is going to be there; this is going to be there.0689

This is going to be there.0693

And now, I'll draw in my axial position.0696

Axial is vertically down, 1, 2, 3; and, of course, boom, boom.0699

And now, I'll do my equatorial positions.0705

There is 1; there is another.0707

There is another; there is another, and this is always an interesting one.0711

It goes that way, and I have the oxygen.0718

OK, let me go ahead and do, here OH.0723

That means OH is down; OH is down.0727

This OH is up, so it is up here, and this OH, this is the 1, 2, 3, 4.0732

Let me number these so you can see them.0740

Oops, I wanted to do that in red, make sure we press it.0743

This is the no. 1 carbon, so 1, 2, 3, 4, and the 4 carbon is down; so the OH is here.0749

And now, over here, we have, of course, our CH2OH; and the others are just hydrogens.0757

That is OK, I can go ahead and put them in; it is not a problem.0765

Here, OK, that is one chair conformation.0770

Now, let me go ahead and put a little perspective on it.0772

I'm going to bold that out with a wedge.0777

I'm going to bold this out, and I'm going to bring this out and make it bold.0781

There you go.0788

This alpha-glucose, this is one of the conformations; this is the left side flipped up and the right side flipped down.0790

Now, we are going to be the left side flipped down, the right side flipped up.0797

Let me go ahead and draw that one out.0800

We have got this, that, that.0804

Again, oxygen is back there.0812

This is there; this is there, and this is there.0815

OK, now, our axials are here, here, here, here, here, and nothing over there; and now, I'll do our equatorials.0819

We have one there; we have got probably one like that, or it is probably not the best representation.0830

Actually, let me make it a little bit more angled.0837

These are always interesting to draw, aren't they?0841

OK, this one goes that way; this one goes that way, and this one goes that way.0844

OK, we said, we are still dealing with alpha-glucose.0851

OK, this is for alpha-glucose.0855

This is one conformation of alpha-glucose; this is the other chair conformation.0857

Alpha-glucose, this has no. 1 hydroxy, so let me go ahead and number.0861

This is 1, 2, 3, 4, 5, 6.0865

I'll go ahead and put the CH2 group up here; I'll just go ahead and write it in.0874

Now, let me go back to black.0878

It is still below the ring.0880

OK, this is below; this is above.0884

This is below.0889

OK, let me go ahead and put the Hs in just so we see them.0891

OK, so, we have something that looks like this; and now, this is the other.0896

Now, let me put a little perspective on it.0903

I just wanted you to see what this looks like.0912

These are the actual forms that they take.0916

Now, notice, in this particular case, in this one over here on the left, this CH2OH group, that is in equatorial position, not axial position.0917

It is actually pointing away from it; it's not vertical.0927

It is away from the ring.0929

Here, with a little bit of a flip, this flipping down, this flipping up, it takes on an axial position; it is vertical.0931

You remember from organic chemistry, the largest substituent on a 6-membered ring, because of steric reagent, it will actually arrange itself such that the largest substituent is in the equatorial position.0939

This particular conformation will probably be more abundant simply because the CH2OH in this conformation achieves an equatorial position.0955

Now, these 2, alpha and beta, are configurations.0966

The only way to go from 1 configuration to another configuration, bonds have to break.0971

In other words, this bond has to break and has to reform up here as a hydroxide.0975

These are conformations; bonds don't break.0981

The only thing that happens is the molecule flips around a little bit or bonds rotate.0984

That is the difference between conformation and configuration.0990

Alpha-glucose has 2 conformations, 2 chair conformations- this and this.0992

We don't give names to them; they are just the 2 conformations.0997

Beta-glucose also has 2 conformations- this and this.1000

They look exactly the same except for beta, the hydroxy would be up here, and the hydroxy would be up here.1004

That is the difference.1012

Let's go ahead and mark in blue just so we know the no. 1 carbon.1014

This is the anomeric carbon.1018

OK, and again, we still kept our conventional.1020

The oxygen is in the back right, oxygen is in the back right, but now, it is in actual position.1025

Now, this is what the molecule looks like in space, and here is the anomeric carbon for that one- there and there.1029

OK, now, let's talk about some hexose derivatives.1040

Derivatives just mean we have reacted them with something, and we have attached new groups to it.1046

That is all a derivative means; we start with something basic, and we derive something from it.1050

Let me go ahead and stick with blue; I like blue.1055

Hexose derivatives that play key roles in physiological processes.1061

I'm not sure about the extent to which your teachers are going to have you necessarily memorize these.1077

It is good to be introduced to them.1081

We will tell you a little bit about them; tell you their names just so if you run across them, you will have a good idea what it is that you are dealing with.1084

You are going to see these again, anyway.1090

Again, it is going to be up to your particular professor about the extent to which they want you to know the structure, the name, what it is, things like that; but it is good to see some, anyway.1093

OK, let's start off with our beta-D-glucose.1100

Again, it is always great to start with your basic structure, so you always know where you are coming from.1105

Beta, the hydroxy is up on the top; this one is down.1111

This one is up; this one is down, and we have CH2OH.1115

This is our beta-D-glucose.1120

OK, now, let's go ahead and make a little bit of a change here.1127

Let's go ahead and draw this same thing.1131

O, except, instead of a hydroxy - let's go ahead and do this one in red - let's go ahead and put an NH2 group there.1136

Let's go back to blue, and let's finish off the hydroxys and CH2OH.1147

OK, so, what I have done is I have taken this no. 1, no. 2.1155

The hydroxy on the no. 2 carbon, I've gone ahead and replaced that with an amino group and NH2.1158

This turns it into beta-glucosamine.1163

Beta-glucosamine or glucosamine, however you want to pronounce it- not a problem.1168

That is it; I've just replaced this with this.1177

The hydroxy on the no. 2 carbon is replaced with an amino group- nice and easy, OK, in other words an NH2.1187

OK, now, let's do another derivative.1208

Let's go ahead and draw it again; this time I'm going to do it.1210

You know what, let me stick with the blue, and let me go draw out my hexose ring.1214

And again, this is beta, except now, what we are going to do is we are going to put an N, and what we are going to put is C, and we are going to put a CH3, and we are going to put that there; and everything else is going to stay the same.1224

Oops, let me go back to blue.1239

Hydroxy, no, that is not right.1245

Hydroxy goes up; this is glucose and CH2OH.1249

And again, I hope that you are checking these structures with me because again, once you just sort of start writing them, mistakes are made.1258

We are all human.1265

OK, here, what we have is, notice, we have this amine part, which is the same.1266

So, we have the nitrogen, but we have stuck this acetyl group on it.1272

This is called N-acetyl-beta-glucosamine.1276

That is it; the acetyl is attached to the end.1289

I haven't attached it anywhere else; I could have.1293

So, this is N-acetyl-glucosamine.1296

That is it.1301

This is my new group.1304

This is a derivative.1306

That is it.1308

An acetyl group is attached to the N of the amino group.1310

OK, alright.1333

Now, let's do another derivative here.1342

Let's go ahead and go back to blue.1345

Let's draw our structure.1348

Actually, you know what, this one, I'm going to need a little bit more room up on top; so, let me try this again, draw it a little bit lower.1354

Let me draw it down here.1362

O and this is OH, and I'm going to go CH2O.1364

Let me actually finish the glucose part first.1375

This is up, and this is down.1377

There we go; and now, I have P, double bond O, single bond O-, single bond O-.1382

We have this phosphate group - OK - attached to the no. 6 carbon.1389

This is the no. 6 carbon, no. 1 carbon- anomeric.1401

This is, well, beta-glucose-6-phosphate.1405

This is beta-glucose-6-phosphate.1411

It just is that I have a beta-glucose, and on the no. 6 carbon, I have attached a phosphate.1418

OK, this is the first step of glycolysis, where glucose is converted to glucose-6-phosphate.1424

OK, now, let's draw another derivative here.1431

Let's see how these turn out.1435

I wonder if I should do...that's OK, I guess I can do it in this page, not a problem.1437

This one I'm going to do in black.1440

Let me go ahead and go this way.1444

Well, that is fine; I'll go ahead and just do, I'm going to draw this particular one out.1449

No, what am I doing?1456

Let me go OH, CH2OH, and this is going to be O-.1463

OK, that is up; that is up.1472

That is down; that is up, and this is down.1476

OK, now, what we have done here is the aldehyde, which is originally this thing, is now, the aldehyde has been oxidized to a carboxylic acid or a carboxylate.1479

In this case, it is carboxylate because the hydrogen is deprotonated, so it is a -1 charge.1508

If the hydrogen were attached, it would be the actual carboxylic acid.1513

OK, this is called, in this form, D-gluconate or just gluconate.1517

Again, we are dealing with the D configuration.1526

So, this is gluconate.1530

Now, this particular one, gluconic acid - that is the protonated form, OK - is called gluconic acid.1532

The deprotonated form is just gluconate.1545

Carboxylic acid, carboxylate, OK, the general term for this class- an aldonic acid.1550

Whenever you take the linear form of the sugar, and then when you oxidize the aldehyde end to a carboxylic acid but without oxidizing anything else, just this one, just the aldehyde end, you turn it into something called an aldonic acid or an aldonate.1560

In this particular case, since we used glucose, we turned it into gluconic acid; but it is deprotonated, so we call it a gluconate.1575

Again, it is just a question of protonation and deprotonation.1583

This is the general term; it is called an aldonic acid when you oxidize just the aldehyde, not anything else.1586

OK, now, here is what's interesting about this.1594

We have this carboxylate group, and we still have this hydrogen over here, and it still has these nucleophilic electrons.1600

So, the hydroxy on the no. 5 carbon - and again, this is the no. 5 carbon there - can still react with the no. 1 carbon - this is the no. 1 carbon, the no. 1 carbon hasn't changed - to form something called a lactone.1609

And, a lactone is just a fancy word for a cyclic ester.1647

It is an ester that is a part of a cycle.1651

We put the parentheses down here.1658

A lactone is nothing more than a cyclic ester.1662

OK, let's go ahead and draw this out.1666

What I have got is this, this, this, this, this, that.1671

I have that; I have OH.1678

This is down; this is up.1680

This is down, and this is CH2OH.1682

Nothing else has been oxidized, and this is called glucono-delta-lactone.1687

Remember what we have done here, that hydroxy?1698

It actually reacts with the carbonyl; it kicks off that other OH, that other O-.1703

Actually, let me go ahead; and let me draw it here, again, just off to this side.1711

So, we have something like this: COO, O-, this, and CH2OH, down, up, down.1715

Remember we had something that looked like this on the previous page?1728

This is our gluconic acid.1731

Well, this thing is actually going to form a bond with this thing, and this thing is going to end up going away.1737

We won't worry about the mechanism, but you end up with something like this.1741

This is a lactone; it is a cyclic ester.1744

An ester is a carbonyl with an oxygen attached to another carbon, but this carbon happens to be part of the ring.1747

So, a cyclic ester is called a lactone.1756

Now, it is called a delta-lactone because, remember what we said, the carbonyl is the no. 1 carbon, but the carbon next to that is called the alpha carbon.1758

This is alpha, well, this is the beta carbon.1766

This is the gamma carbon; this is the delta carbon.1770

Because the oxygen next to the carbonyl is attached to delta carbon, it is called a delta-lactone.1773

It will be a gamma-lactone, an epsilon-lactone.1780

That is all that's going on here.1785

OK, alright.1787

Let's go ahead and close this lesson off by talking a little bit about the idea of a reducing sugar.1790

Let me draw a little bit of a line here.1795

Now, monosaccharides - I'll just write monosacchs - can be oxidized by very mild reagents such as, in particular, I should say, Fe3+ and Cu2+.1798

So, the sugar is oxidized.1830

OK, they are reducing agents.1832

Reducing agents are the things that are oxidized.1843

They are reducing agents because they reduce the iron, and they reduce the copper; but they themselves are oxidized- very, very important.1847

OK, as such, they are called reducing sugars.1856

They are called reducing sugars.1862

And, the carbonyl carbon, the carbonyl C, is oxidized to a carboxyl group.1874

The general reaction is as follows.1897

You have RC.1900

This is the aldehyde.1904

OK, let's just go ahead and use 2Cu2+ to 2Cu1+, and what you end up with is RC.1907

What you end up with is that.1922

This aldehyde is actually converted to a carboxyl, OK, because that is available, the free end of the linear form of the sugar.1924

So, we call them reducing sugars because they are capable of being oxidized by iron or copper, iron ionic, copper ion; as such, we call them reducing sugars because they are reducing agents.1935

That whole oxidation-reduction thing, calling them reducing agent, it still confuses me.1949

I just think in terms of oxidation, but they are called reducing sugars.1953

OK, let's go ahead and draw out the reaction in full form.1956

Let's go ahead and start off with a nice hexose structure like this.1962

Let's go ahead and use our beta form.1970

OH, OH, OH, and then we have CH2OH there, so this is...actually you know what, I'm going to use...I'll make a little bit of a change.1974

Just for a little, slight variation, I'm going to use galactose.1993

Instead of glucose, I'm going to use beta-galactose just for a little bit of a variation.1998

This is beta-galactose.2007

Notice, in the glucose so, 1, 2, 3, 4.2008

Galactose is a c-4 epimer of glucose.2012

Everything is the same except at the fourth carbon, the hydroxy is on the other side.2016

It is above the ring instead of below the ring.2021

So, it is a different, actual molecule.2023

Now, the ring form is going to be in equilibrium with the linear form.2028

There is going to be some linear form of this sugar that is going to be available.2034

1, 2, 3, 4, 5, I have 6 carbons.2038

I open it up; I have the aldehyde on top, and of course, I have OH here.2041

This time, I have 2 OHs on the left.2047

I have this one - oops - and, of course, I have my non-chiral carbon.2050

CH2OH, there we go.2057

Now, once it actually opens, and this end is free, it can react with 2Cu2+ ions to release 2Cu1+ ions, and, of course, now, 1, 2, 3, 4, 5, 6.2061

This is H2OH; I have my carbonyl which has been ox by aldehyde, has been oxidized now, to a carboxylic acid.2086

This hydroxy is on the right; this hydroxy is on the left.2097

This hydroxy is on the left, and this hydroxy is on the right.2100

So, what I have here is beta-galactose ring form.2104

This is also beta-galactose linear form.2110

This one, galactonate, remember gluconate, galactonate.2115

This O-N, just drop the O-S-E from the galactose, glucose, mannose.2125

Add the O-N, and either put galactonic acid, if it is protonated, or galactone if it is deprotonated; and, that is it.2129

I'll go ahead and write that, in other words, if this is protonated right up here, if that is protonated.2139

OK, now, of course, the aldehyde group must be free in order for this reaction to take place.2157

In order to be oxidized, the aldehyde has to be free.2175

These are in equilibrium.2184

In solution, yes, it is going to exist mostly in this form, but there is always going to be some of this available; and once this is actually used up and converted to this, then more of this is going to open up.2185

This is just an application of Le Chatelier’s principle.2195

A reducing sugar is precisely this.2199

A reducing sugar is a sugar that can actually be oxidized by Cu2+ or Fe3+ to form the corresponding aldonic acid, which itself can actually form a ring again to form that delta-lactone that we just mentioned a little bit earlier.2201

OK, that should just about cover it as far as derivatives is concerned.2220

Thank you for joining us here at Educator.com2223

Take care, bye-bye.2224

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today, we are going to start talking about disaccharides, putting 2 monosaccharides together in something that is called a glycosidic bond.0004

Let's go ahead and get started.0013

Alright, let's see how can we do this.0016

A disacch is exactly what it sounds like.0020

CCH, I always forget how to write that.0028

Disacch is made of 2 monosacchs joined covalently- actually, you know what, let me just - joined by a covalent bond called an O-glycosidic bond- very, very, very important bond in biochemistry.0033

The hydroxy group of 1 sugar has reacted with the anomeric carbon of the other sugar.0083

Now, when we say the hydroxyl group of 1 sugar, the truth is, it can actually be any of the hydroxyl groups; but in general, it is going to be specific hydroxy groups that are going to react with the anomeric carbon of the other sugar.0127

Again, you will see from the bond, which particular hydroxy is going to react; but it actually can be any one of them.0143

OK, let's just go ahead and do examples of these.0151

We will just go through several examples to get a sense of how to put the structure together, how the bonds are drawn, the perspective drawings that we are going to be using, and how we are going to name them systematically.0155

That is what's important.0168

Let's do examples of disacchs and how to name them systematically.0172

OK, let's go ahead and start with a particular disaccharide.0189

Let's start with alpha-D-glucose, and connect it to beta-D-glucose in something called a 1→4 connection.0193

OK, let's go ahead and draw out our alpha-D-glucose.0200

Again, always proceed systematically.0206

Start with your basic unit.0209

Alpha-D-glucose, we are looking at this thing right here.0212

God, I really love the blue; I think it is my favorite.0215

OK.0218

CH2OH, this is alpha-D-glucose.0222

And now, we are going to be joining it with - I'll go ahead and put a little plus sign here - beta-D-glucose.0231

Let me draw out the beta-D-glucose first, here.0238

Beta is going to be up here, but everything else is the same because we are talking about glucose, so CH2O4.0244

OK, now, when we put this and this together - let me go ahead and do a red - I'm going to number my carbons.0255

This is no. 1, no. 2, no. 3, no. 4, no. 5.0263

And again, 1, 2, 3, 4, 5, I'll go ahead and put 6 too; it is not a problem0270

And again, standard position, oxygen is in the back right; the anomeric carbon is on the right.0276

That is the reducing end of the sugar; this, over here on the left, is the non-reducing end of the sugar.0281

In this particular case, let me go ahead and write, so this is beta-D-glucose.0287

Again, there is a bunch of hydroxys here that can actually react with this anomeric carbon.0296

In this particular case, we are going to form a 1→4 connection.0301

The anomeric carbon, the no.1 carbon, is going to be connected to the no. 4 carbon; and here is the reaction that is going to take place.0306

This is a condensation reaction.0316

In other words, the elements of water are going to be taken away from this, the elements of water that we are going to be taking away right there.0318

This hydroxy on the anomeric carbon is going to go away, and this hydrogen connected to the oxygen on the no.4 is going to go away.0329

So, the oxygen that is going to go between this unit and this unit, actually belongs to this carbon, the no.4 carbon.0338

What you end up with is the following.0347

Let me go ahead and draw a couple of arrows, one this way and this way; and if we go this way, we are actually losing water.0350

If we come this way, we are actually adding water, so this is a hydrolysis.0358

When you hydrolyze a disaccharide, you get 2 monosaccharides.0363

When you condense 2 monosaccharides, you are getting the disaccharide.0366

OK, this is how it is going to look.0370

I'll do this one in red, boom, boom, boom, boom, boom.0374

OK, I'll go like this, and up like that; and then I've got that, that, that, that, that, that.0380

OK, this is the beta, and now, I'll go ahead and fill in my rest.0389

This is up; this is down.0395

This is CH2OH; this is down.0400

This is up; this is involved in the bond.0404

This is CH2OH.0407

OK, this particular sugar is called maltose.0410

Maltose- this is the common name.0416

In the sugar, maltose, what has happened is that an alpha-D-glucose has reacted with the beta-D-glucose.0422

The anomeric carbon has reacted with the no. 4, with the hydroxy on the no. 4 carbon, to create this right here.0430

This right here, this is your O-glycoside bond.0440

OK, that is your O-glycoside.0446

O, because the O is involved; and notice, down, down.0447

That is why it is drawn this way.0453

This is how we actually represent the arrangement in space, but we keep this particular perspective, so that we see how the molecules, how the individual units are arranged.0456

This is how we do a disaccharide.0466

OK, the name for this is alpha-D - because it is a polymer - glucosyl, the first (1→4)-D-glucose.0469

Alpha-D-glucosyl, that is the first monomer arranged in a 1→4 pattern.0495

One on the left connected to the no. 4 carbon on the right- that is where this 1→4 in parentheses means, and a little arrow going from the 1 to the 4.0502

We go from left to right- D-glucose.0509

OK.0512

I should actually write this as...because we have actually specified the stereo chemistry on this particular monomer, it is the B.0517

This is beta-D-glucose.0525

Now, there is a slightly longer name, but we are actually going to be dealing with a shorter name.0529

I'm going to write the longer name, but then we are going to exclusively start dealing with the shorter name.0533

This is also called - and I'm not going to draw out the structure again, you can just flip back - alpha-D-glucopyranosyl.0539

Remember, pyranose, 6, gluco, pyro, glucose pyranosyl- it is a little redundant, but you will see it, pyranosyl(1→4)-beta-D-glucopyranose.0553

Now, obviously, you can't use something like this, so here is the shortened version.0574

All of the sugars just like the amino acids, they have 3-letter shortcuts for them, like Ala is alanine.0580

Well, in this particular case, glucose is Glc; and there is list in your book of the 3-letter shorthand notation for all of the individual sugars, galactose, glucose, ribose, deoxyribose, whatever it happens to be.0588

In this particular case, the name is going to be written this way: Glc for glucose.0605

We write the configuration at the carbons that are attached by the O-glycoside bond.0614

The 2 carbons that are involved in the bond, we give their configurations: alpha 1 to 4.0620

The fourth carbon, we don't do alpha-beta because the alpha-beta designation is only for the anomeric carbon.0635

Alpha(1→4)-beta-Glc, also written as Glc-alpha(1→4)Glc.0641

Often, the monomer on the right, the fact of the matter is, well, I'll tell you in just a second why it is I wrote beta here and not there.0654

Let me go ahead and tell you what is going on.0667

This does not mention the beta explicitly on the second monosacch because something called mutarotation, it often switches the configuration.0670

Even though, we know that we use the beta version of the second monosaccharide, the fact of the matter is, the alpha and beta forms, they actually often switch.0712

So, the stereo chemistry on the reducing end of the sugar, the one that has the free anomeric carbon, it is often unspecified.0722

Sometimes, you don't necessarily have to put the beta there; it is not a problem, unless, specifically, they want you to.0730

In this particular case, we put together an alpha-glucose with a beta-glucose, and what we ended up with is Glc, glucose, alpha-1 configuration at the anomeric carbon connected to the no. 4 carbon, the hydroxy on the no. 4 carbon, and the other disaccharide was a glucose.0736

That is all that happens- switches the alpha-beta-configurations.0755

Again, either one of these is fine.0764

If you want to specifically write the beta, that is fine; if not, you are not going to have any points taken away.0765

OK, now, again, as I said, if the anomeric carbon on the second monosacch, in other words, the one on the right, on the monosacch is free like it was with maltose, then this is called the reducing end of the sugar because now, you have a disaccharide, which happens to be a reducing sugar.0770

Fe3+ of Cu2+ will still oxidize that end.0815

It is available to be oxidized; it isn't always the case.0820

In a minute, you will see an example of something that is not a reducing sugar, a disaccharide; but this one, because the anomeric carbon, that hydroxy was not involved in the O-glycoside bond, it is still free, it is a reducing sugar.0823

It is called the reducing end.0839

And again, we have a reducing sugar.0845

Let me read that again.0855

If the anomeric carbon on the second monosaccharide is free, then this is called the reducing end; and again, we have a reducing sugar.0857

And, of course, the other end, the one on the left, the one that cannot be oxidized, that is the non-reducing end.0864

OK, maltose, what we just did, is a reducing sugar.0870

Maltose is a reducing sugar.0880

OK, now, as we said, any of the OH groups on the second monosacch can react with the anomeric carbon of the first monosacch - OK, excuse me - even the hydroxy on the anomeric monosacch, even the OH on the anomeric C.0885

Let's look at an example.0943

OK, now, let's go ahead and look at the sugar trehalose.0946

Let's look at the disacch trehalose.0958

OK, let's see what we have here.0964

Now, trehalose, we are going to have an alpha-D-glucose and an alpha-D-glucose that are going to be connected by their anomeric carbons.0966

Let's go ahead and draw the alpha-D-glucose.0974

We have, this alpha is right there; that is there.0978

That is there; that is there.0988

This is CH2OH.0990

This is alpha-D, and I will just put Glc for glucose.0993

Now, we are going to connect it to another alpha-D-glucose- down, up, down.0998

OK, this time, they are connected; the no. 1 carbon is connected to the no. 1 carbon.1018

How are we going to show that?1028

OK, but now, the OH on the...no...the OH on the no. 1 carbon of the second sugar, no. 1 carbon, the anomeric carbon of the second sugar, is going to react with the anomeric carbon of the first.1030

Sugar reacts with the no. 1 carbon of the first sugar.1070

OK, how are we going to deal with this, and how are we going to represent it?1083

We are going to do this in 3 different ways.1087

Well, let's see.1092

How do we deal with that?1095

How do we represent this schematically?1099

How do we actually show the bond?1103

How do we represent this?1105

OK, here is how we do it.1107

We either flip or spin the second sugar so that the Ns that are going to be reacting are close to each other.1111

When we do that, we are going to have to change the arrangement of the second sugar.1120

In other words, now, it is no longer going to have the conventional representation of the oxygen being on the back right.1124

Let's go ahead and how do we deal with this?1135

Well, we flip or spin the second monosacch for a new arrangement on the page.1136

OK, let's go ahead and do the first one; I'm going to do both.1163

I'm going to do a flip first; I'm going to do a spin first because you are going to probably see them both.1168

My guess is, more often than not, in most biochemistry books, you are actually going to see it flipped; but you will see the spin version, too.1173

This is really, really, really important.1180

There is lots of carbons and oxygens and hydrogens floating around here.1183

Do not go through these structures quickly.1189

Make sure you understand, make sure you pay very close attention to where each individual atom is, particularly that oxygen because that is what's going to give away the structure, and what it is that is going on.1192

OK, because all of these things look alike, the only thing that actually tells you what is different is the arrangement of this particular oxygen in the second monosaccharide.1205

If it happens to be back, if it happens to be forward, that tells you which carbons have reacted.1216

OK, let's go ahead and deal with the flips.1220

We are going to actually end up flipping this one; and the reason we are flipping it, in other words, I'm saying "flip it this way", because you want this carbon to be over on this side, because you want it to be close to the carbon that it is going to be reacting.1223

We have already told you that trehalose has connected the anomeric carbon; carbon no. 1 and carbon no.1 are the ones that are connected via the glycosidic bond.1236

So, we need a way to represent this on paper, so we want to flip this, so that this bond over here, we don't just want to draw the line connecting them.1245

OK, let's draw this one.1254

Again, let's do it in blue.1256

We have got this, this, this, that, that, that; and this is alpha.1260

This is going to be here.1269

OK, we've got CH2OH.1276

Now, what I'm going to do is I'm going to...wait, where am I?1279

OK, yes, now, I'm going to flip this.1286

OK, when I flip this, I'm just flipping it like this.1290

Everything that is on the right goes to lo the left; everything that is on the top goes to the bottom.1300

Thing move around.1304

I'm going to redraw it like this.1306

OK, I've taken it and I've flipped it.1313

Now, what used to be...let me number these.1315

Oops, I want to do this in red.1321

1, 2, 3, 4, 5, 6, now, what you have is, I've flipped it, so now, it is 1, 2, 3, 4, 5; and let me actually finish off the structure.1324

Let me do the structures here.1341

When I flip this, this is on the bottom; it goes over to the right, and now, it is up here.1344

This O has moved over here; this C has moved over here, this 5 carbon, but now, this is up.1352

Now, this is down- CH2OH.1359

OK, the no. 2 carbon, it is down; so this is going to be up.1363

No. 3 carbon, this is up; so it is going to be down.1369

No. 4 carbon is down, so now, it is going to be up.1373

This is the flipped arrangement.1376

Now, OK, does that make sense?1380

Notice where each group is; 1, 2, 3, 4, 5, I have flipped it.1382

It is not a mirror image.1387

OK, it is not a mirror image.1389

I have actually flipped it; everything is reversed, not mirror-wise, but this way.1392

That is what's going on.1401

What I need it to do was I need it to bring my no. 1 carbon to put it in close proximity with the no. 1 carbon of my first monosaccharide.1403

Now, this thing is going to react with this thing.1414

OK, 1, 1.1425

The elements of water that are going to disappear is the hydroxy there, the H here.1428

This oxygen is going to be attached to that carbon.1435

When I put those together, this is the arrangement that is going to take place.1438

What I end up with is the following.1442

This is the same; we haven't done anything to the first monosaccharide.1467

That is CH2OH, but this one, things are a little different.1475

We have flipped this other one.1478

So, this does not look the same.1481

You might think it does, but it does not.1485

Then it is very, very important that you realize where the oxygen is.1489

OK, here the oxygen is on the top right, back.1495

Let me draw the perspective.1498

OK, here the oxygen is on the back left.1508

Here, the CH2OH is on the upper left above the ring; here it is on the back left, but the CH2OH is below the ring.1511

OK, this 1, 2, 3, 4, 5, 6, 1, 2, 3, 4, 5, 6, I have flipped it, so that I could put these 2 carbons, the ones that are reacting in close proximity, so that I can represent this glycosidic bond in this fashion.1520

OK, when this is an alpha, the hydroxy is below the ring.1557

This is also an alpha; in conventional position, the hydroxy is below the ring, but because I flipped it, I now, have the hydroxy above the ring at that carbon.1562

That is why I draw it this way.1574

This is alpha-D-glucosil-1↔1 - a double arrow, when you are connecting anomeric carbons - alpha-D-glucose.1577

A shorthand notation, Glc for the first monosaccharide; alpha-1 configuration, that carbon, the no. 1 carbon, it is an alpha-configuration that is involved in the glycosidic bond.1598

The other one is also the no.1 carbon of the second monosaccharide in alpha-configuration, that is the other carbon involved in the glycosidic bond; and just go ahead and put that.1615

This is what you want to write: Glc-alpha-1↔alpha-1-Glc.1627

This is trehalose.1634

OK, now, you notice, the anomeric carbon, the reducing end of 1 carbon reacted with the reducing end of the other.1637

This end over here, it is not available for oxidation.1645

OK, it is not available for oxidation, so this particular sugar does not have a reducing end.1649

Therefore, this is a non-reducing sugar.1656

Trehalose is a non-reducing sugar.1659

This changes the chemistry entirely.1661

They behave in completely different ways.1664

Notice, if you were to just look at this really, really quickly without even, sort of, thinking about it, it would look almost exactly the same as the sugar that we just did, which was maltose.1668

I mean, yes, you might notice that this particular thing is different, but you might think to yourself "oh, maybe they just drew it differently, that's all".1677

No, there is nothing random here; everything is drawn with a specific purpose.1684

What you would notice on maltose, is that on both monosaccharides, the oxygen is on the back right.1688

Here, the first monosacch is on the back right; the second monosacch, the oxygen is on the back left.1694

We have specifically drawn it like this.1699

This is what is important.1702

So, you really, really have to pay a very close, detailed attention to these particular structures.1703

OK, I'm going to go ahead and do the spin version of this really quickly.1708

Again, this is just something that you may see.1719

I'm going to take my alpha-D-glucose, so let me draw that under standard.1723

Alpha-D-glucose, hydroxide, hydroxide, hydroxide there, hydroxide here, and CH2OH.1730

Now, instead of flipping that second monosaccharide, I'm going to go ahead and spin it.1739

When I spin it, I end up with the following.1745

Let me see.1749

Yes.1754

OK, when I spin it, it means I'm not flipping it, I'm just spinning it 180°.1756

I'm rotating it this way.1764

Flipping means like that; spinning or rotating means like this.1767

So, here is what it ends up looking like, boom, boom.1772

Now, the oxygen is on the front left.1776

OK, basically, just follow all of these things and just go to the opposite pole.1780

This goes here; this goes here.1785

This goes here; that is all you are doing.1788

It ends up like this; that OH goes there.1792

This ends up coming, CH2OH.1797

This is down, so it is going to stay down.1805

This is up; it is going to stay up, except now, it is going to be in the back, and I think, I have covered everything.1808

So, what you have got is 1, 2, 3, 4, 5, 6; now, what you have is 1, 2, 3, 4, 5, 6.1815

Now, notice, in this particular case with this spin arrangement, now, my oxygen is down below the ring.1831

So, when I draw my trehalose structure, here is what my structure is going to look like.1837

This stays the same; that, that, that, that, that and that.1842

That is going to go like that; and here, oxygen is there, so, OH, OH, OH, CO2OH.1854

And now, we have OH, OH, OH; and we have CH2OH.1871

OK, this is again, glucose, alpha-1, alpha-1, in parentheses, double arrow, because now, double arrow, I'm connecting the anomeric carbon with the anomeric carbon.1884

Each one has an alpha-configuration, and they are both glucose monomers.1898

Notice, this is the exact same thing as this.1904

Notice, here, the glycosidic bond is represented this way, below, above.1908

The oxygen on the second monosaccharide is in the back left, but this was based on the flip.1915

Here, I have decided to spin it; and now, the glycosidic bond is represented this way.1922

This is the same molecule; this is not a different molecular.1927

It is just that the second monosaccharide is arranged in a different way.1930

The first monosaccharide is exactly the same.1933

This is why it is really, really important to be able to distinguish, watch for where this oxygen is, watch for where the no. 1, no, 2, no. 3, and no, 4 carbons are.1937

Here, it is 1, 2, 3, 4, 5, 6; Here it is 1, 2, 3, 4, 5, 6.1947

This is the same molecule, different arrangements in space.1957

OK, so, you have seen the spin; you have seen the flip.1962

I am actually not going to go ahead and show you the third version.1965

I don't think your teacher actually wants you to do that anyway.1968

OK, now, let's see.1975

Trehalose, again, is a non-reducing sugar because there is no anomeric carbon that might open up to release a free aldehyde that can be oxidized, so it completely changes the biochemistry.1979

OK, let's go ahead and do an example.1990

Well, we have done a couple of examples; this is just some sort of a free example.1995

OK, sucrose, which is table sugar, is Glc-alpha-1-beta-2-Fru.2002

OK, sucrose is a disaccharide, and it is made up of a glucose unit and a fructose monomer.2016

So, glucose is a hexose; it is a 6-membered sugar.2025

Fructose, remember, is a 5-membered ring sugar.2027

The connection between the two, the glycoside bond that connects them, connects the anomeric carbon, which is alpha and the no. 2 carbon, which is beta-configuration on the fructose.2030

Let's go ahead and draw out the structure.2046

That is our assignment; this is what we want to do.2048

In this particular case, we haven't given you a structure.2050

What we have done is give you the name, in shortened form; and we want you to draw the structure- very, very typical question on an exam.2053

OK, in this particular case, well, let me just go ahead and draw the...should I go ahead and...well, that is fine.2065

In this particular case, I am joining the anomeric carbon of both.2078

Again, I am going to have to bring the carbons in close proximity.2084

I am going to have to flip or spin the second monosaccharide.2089

I am going to choose the flip version.2091

Let me go ahead and draw out my...I choose to flip the second monosaccharide.2094

OK, and again, when you see an alpha or a beta on this second carbon here, this second sugar, that is what's going to tell you that you are probably going to have to do some spinning or flipping.2108

If there was just a number here like 4 or 5 or 3 or something like that, then you can just leave them alone, and just connect the carbons.2119

OK, let's go ahead and draw out our alpha-glucose.2129

That is going to be like this, alpha which means the hydroxy is down here, and this is there, and this is there, and this is CH2OH.2133

OK, now, flip the beta.2149

Let me see.2155

I'm actually going to go through the process of putting this particular fructose together.2158

I'm going to start the process this way - I'm going to do this in blue - just so you see again, a little bit of review on how it is that we actually create this particular ring sugar.2163

Fructose, again, is a 6-membered ring, so we have 1, 2, 3, 4, 5, 6.2174

That is up; that is going to be down.2186

Yes, OK.2188

Except this time, C, this is H2OH.2190

So, this is a ketone; this is a ketose.2196

The no. 2 carbon actually has the carbonyl.2198

Here, OH, I'm drawing out the linear form, the 1, 2, 3, 4, 5...oops, I forgot.2201

This is OH, and this is CH2OH.2212

So, this is the linear form.2214

I'm going to go ahead and take this linear form, and draw it in such a way, in order to create my ring.2216

Let's see, I have got C, carbonyl.2224

This is CH2OH here.2232

This is C; this is C.2235

That is C, and then I have my OH, and I have my CH2OH here.2240

This is in freeform that I've taken and rotated; I have brought the other thing around.2245

Now, 1, 2, 3, 4, 5- that is right.2250

I am going to end up attacking that right there, and what I'm going to end up creating is my beta-fructose.2256

My fructose is going to look like this, and I have that.2266

So, beta, that means, so this is the no.1 carbon, right?2271

No. 1 carbon, no. 2 carbon, so CH2OH, let me go ahead and number these.2276

This is 1; this is 2,2284

This is 3; this is 4, and this is 5, and I will do no. 6 in just a minute.2286

Let me go ahead and put all of my substituents on here.2294

Oops, as you can see, things get very, very, very involved.2299

Let me go ahead and put my hydroxy there.2306

Let me put my hydroxy there.2309

Let me put my CH2OH over here, and I have my beta-configuration.2312

This is my - write these out - this is my alpha-glucose; and this is going to be my beta-fructose.2318

I started with my fructose, linear form; I created my regular beta-fructose.2329

This is the standard, conventional arrangement.2335

I am going to be reacting this with this, alpha-1, beta-2.2338

I am actually going to be connecting this carbon with this carbon, which means that I am going to have to flip this, so that I can bring this carbon in close proximity to this carbon.2343

Now, when I flip this, here is what it is going to look like.2353

I am going to do that in red.2358

The flipped fructose, I am going to flip it.2365

Well, the arrangement, that way, actually stays the same; but, of course, the substituents look different, this.2369

Now, what I have is, this is down, so it is going to be up.2376

This is going to be CH2OH.2384

This is up; it is going to be down.2387

This is CH2OH; this is down.2390

This is up, so it is going to be down over here; and this is going to be up.2393

This OH is up; over here on the right, it is going to flip around this way.2404

It is going to end up being down here.2408

It almost looks the same, except the carbons are numbered differently now.2410

Now, we have our no. 1 carbon, no.2, no. 3, no. 4, no. 5, and no. 6.2416

Oops, go ahead and do that.2427

Now, I am going to connect.2430

This is our first sugar; this is our flipped fructose.2437

I am going to connect that carbon with this carbon, the no. 2 carbon.2441

So, you notice, this hydroxy is down, and over here, this hydroxy is down, below the ring.2450

My final fructose structure - I'm sorry - my final sucrose structure is going to look like this.2457

I have got this; let me make this kind of big.2464

OK, I have got 1, 2, there, there, there.2471

Let me go ahead and draw my glycosidic bond, 1, 2, 3, 4, 5.2477

This is an oxygen, and now, I can put my substituents in, hydroxy down, hydroxy up, hydroxy down, CH2OH.2486

Now, I have got a hydroxy down here; I have got a hydroxy up there.2497

I have got a CH2OH here, and I have got a CH2OH - actually, let me draw it to the left because there is plenty of room on the left - CH2OH, and there we go.2504

Now, I have my alpha-1 configuration, alpha-1-carbon on my glucose.2519

This is my beta-2; you can write b2 2-beta.2526

It does not matter, the order, as long as the alpha-1 is on the left of the arrow.2531

OK, we have Glc-alpha-1↔beta-2-Fru- this is sucrose.2537

This is table sugar, and you notice, there is no anomeric carbon that is available.2553

This is a non-reducing sugar; the 2 anomeric carbons are connected by an O-glycosidic bond.2559

It is arranged this way.2566

This is not the standard, conventional arrangement; I had to flip this fructose monomer, in order to bring this carbon in close proximity with this carbon.2568

This is the process that you go through.2579

Again, glucose, glycosidic bond, glucose is connected to fructose; the glycosidic bond is the alpha-1, no. 1 carbon alpha-configuration connected to the no. 2 carbon beta-configuration.2580

That is it.2595

We will do more of these; don't worry about that.2597

We will do plenty of these because it is very, very important that we would be able to go back and forth, that you see a structure, be able to name it, that you'd be given a name, and be able to draw out a structure for it.2599

OK, thank you so much for joining us here at Educator.com.2609

We will see you next time, bye-bye.2611

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

So, the last lesson, we started a review of some general chemical principles that are going to lay the foundation for the work that we do in biochemistry. 0004

For the next couple of lessons, we are going to continue that.0012

Today, we're going to be discussing dilution and osmotic pressure.0015

The notion of osmosis is profoundly important in biological systems.0019

The cell is a semi-permeable membrane, and the relationship that a cell has with its surroundings outside the cell, inside the cell, is all based on the difference in concentration; so, osmosis is huge in biological systems.0025

OK.0042

Let's just go ahead and jump in.0043

Let's begin with dilution.0047

You all know from experience what dilution is.0051

Basically, you are changing a given concentration by the addition of solvent.0054

If you have some solution and it has a certain concentration, if you add more solvent to it, the concentration of the solute is going to diminish; because remember, molarity is moles of solute over liters of solution, right, mol/L.0081

If the number of moles of solute stays the same in there, but all of a sudden I up the liters of solution by adding more solvent to it, well, this is a fraction: as the denominator increases, the concentration decreases.0104

That is what dilution is.0117

OK.0121

Once again, you want to change a given concentration by the addition of solvent.0122

You are trying to dilute the solute.0126

Let's go ahead and do an example.0129

This is a very, very important example.0134

This is something that all of you are going to do at one time or another, if you haven't already.0137

A student is given a stock solution - let me erase this, actually - of hydrochloric acid which is 11.8mol.0142

How can he prepare 500mL of a 1.5M HCl solution from this stock?0176

Basically, he has this solution which is 11.8mol but what he needs is, he needs to create 500mL of 1.5M, so, how is he going to do this?0203

Let's think about what he is going to do.0217

Basically, what he needs to do is put in a certain amount of stock solution of a given molarity, and he needs to dilute that by adding water, bringing it up to 500mL, and then making sure that that 500mL is 1.5M.0220

The question becomes, "How much of the stock is he actually going to pull out of the stock solution to put into this beaker, and then on top of that, add the water to bring it to 500mL, to turn this into a 1.5M solution?".0244

That is what we are doing here.0258

OK.0260

The basic dilution equation is as follows.0261

The basic dilution equation says that the initial molarity of something times its initial volume is equal to its final molarity times its final volume.0271

This is also written as m1v1 = m2v2.0285

This M is molarity, not mass - very, very important.0304

The molarity times the volume that we start off with, in other words, of the stock solution once we've diluted it, the final molarity, the final volume- this is the equation that we want to work with.0306

Again, we're concerned with - let me go to red - we want to know how much stock do we have to pull from the stock solution to dilute that up to 500mL.0318

That is what we are trying to do.0330

OK.0332

Well, Let's take a look at what it is that we actually have.0333

We know that we need this, this, this and this.0336

Of course, with any equation that has four different parameters, you can be given any of the other three, in order to find the fourth.0341

The problem itself is going to dictate which parameter that you are actually looking for.0346

In this case, let's see what it is that we have, and then we'll find out what parameter we need.0352

Well, we know what the final volume is going to be so the final volume, that one equals 500mL or 0.5l depending on which units you're going to use, as long as the units are consistent.0357

We have the final volume, and we also have the final molarity.0372

We are looking for a solution which is 1.5M.0377

I'm just going to make my M the way that I'm comfortable doing it.0386

Here, how's that?0388

Our final molarity is going to be 1.5M.0389

OK.0395

What's our initial molarity?0397

We are using a stock solution.0400

The initial molarity is 11.8mol, so, that's going to be 11.8.0402

The only thing that we are missing is the initial volume.0408

In other words, I need to find how much of the stock solution I'm going to dilute to 500mL.0412

I'm going to put in a certain amount of stock solution that's this, my initial volume of my stock; and on top of that, I am going to add water to bring that final volume to 500.0419

So, that is what I do.0429

Well, This is nice and easy, very basic equation.0431

We will just go ahead and put it in.0433

So, we have initial volume times initial molarity, which is 11.8 equal - you know what I'm going to do, I'm actually going to use the units here and because I want you to see what cancels and what doesn't.0436

So we have Vi x 11.8mol/L, that is the initial volume; and molarity is equal to the final volume, which is going to be 0.500L because we are dealing with moles per liters, so the units have to match so that they can cancel, and the final volume, that's this one. 0450

The molarity is 1.5mol/L. 0478

I just switched them around.0483

Here, I have M first and V second; here I have V first and M second.0485

Sorry about that.0490

I hope that doesn't confuse you.0491

That is it, now, let me go ahead and solve for my initial volume.0492

Vi is going to equal 0.0636L or 63.6mL.0498

63.6mL is the amount of stock solution that I am going to pull, 0510

I'm going to add that to a beaker, and then I'm going to bring the volume up.0518

Let me go ahead and write out the procedure.0521

Take 63.6mL of stock solution, stock HCl and pour into a volumetric flask.0529

I'm sorry, a 500mL volumetric flask.0547

Now we want to be as precise as possible, I mean you can use a beaker if you want but again, you are trying to create a solution, so, you want to use a 500mL volumetric flask.0551

Those are flasks that are specifically calibrated to create a very, very accurate volume: 500mL, 250mL, 1000mL, 50mL.0560

They have a certain mark on them to tell you where to stop adding water; at that point, you are exactly where you should be.0571

So, you pour this 63.6mL of stock into a 500mL volumetric flask, then add solvent, add water until the mark, which is 500mL and then just mix thoroughly.0578

There you go.0606

That's it.0607

The dilution equation: the initial molarity times the initial volume equals the final molarity times the final volume; therefore, parameters here, in order to find the one that you want, you have to have the other three.0611

That's it.0626

OK.0629

Now, the idea behind this equation m1v - let me write the equation again, let me go back to black here- the idea behind this m1v1 = m2v2, is that when you're diluting something, when you're adding solvent to something, you're not changing the solute amount.0630

The amount of solute, whether it's floating around in 100mL of solution or floating around 1000mL of solution, it's still the same amount that is floating around in solution; there just happens to be a hell of a lot more solvent now in which it can float around, so the number of moles of solute actually stays the same.0654

That's what this equation is based on.0674

Let me write this as mol/L x L = mol/L x L.0677

What liter cancels, leaving you the initial number of moles, is going to equal the final number of moles.0687

That's the whole idea here.0692

That's what this equation is based on.0695

The amount of solute doesn't change.0696

The moles of solute floating around are the same.0698

The only thing that changes is the volume.0701

Given this, I am going to go ahead and do this problem again in an alternate way just for that sake of doing it in an alternate way.0704

I personally do it this second way.0713

I don't use the m1v1 equation, but I know a lot of kids actually prefer the M in equation to work with; but I think about it like this in terms of keeping the amount of solute constant.0715

This is going to be example two, and let me do this one in blue, and either procedure is fine.0727

Different people find different ways of looking at a particular problem, but it's nice to know, in this particular case, we can give you an equation and you can work with it.0735

But, if you know it's actually happening underneath, if you know that it's based on the fact that the moles are the same, then you have another way of thinking about it.0747

You can reason it out stoichiometrically.0753

Well, what is it that we wanted?0762

We want 500mL of a 1.5M HCl solution.0764

OK.0774

That means, we are going to concern ourselves with moles- 500mL of 1.5M solution, so that's 0.5L x 1.5mol/L.0776

That means, we need to have 0.75mol of HCl floating around in our solution.0791

Well, 0.75mol of HCl x 1L, our stock solution has 11.8M of HCl floating around per liter.0802

When I do this division, I get 0.0636L or 63.6mL of stock solution.0819

This tells me that I need 63.6mL of stock solution.0832

I put that in a beaker or volumetric flask.0840

I add enough water to bring it to 500mL.0842

Now, I've created my 1.5M HCl acid solution.0845

It's based on the fact that all I've added is solvent.0849

The solute concentration, the amount of HCl floating around, is constant; and I've just done it that way.0853

I've gone from volume into molarity to moles, and then from moles, using molarity, back to volume.0860

That's it, so it's just an alternate way of doing it.0865

OK.0869

That is dilution- very, very, very important.0870

Now, let's go on and discuss a new topic.0876

I'm going to discuss colligative properties.0882

Of the colligative properties, we're really only going to be concerned with one of those properties- osmotic pressure.0885

That is going to be the thing that is most important in biological systems.0888

Let me just start off with some basic definitions, and then we'll jump in to osmotic pressure and do some examples.0893

OK.0898

I think I am going to actually go forward one page here.0900

The colligative properties are properties of a solution that depend only - very important - only on the number of free particles of solute floating around in solution, not the identity of the particles. 0903

The colligative properties are certain properties that a solution has that is based strictly on the number of things that are floating around in there.0910

Let's say if I create a sugar solution and there's a certain amount of sugar molecules, let's say 500 sugar molecules floating around in a certain volume of solvent, that's just a sugar solution with 500 particles of sugar.0928

Well, let's say I take another molecule, whatever it is, some protein, and I have 500 protein particles floating around in that solution; well, that solution is going to behave the same way simply because there are 500 of each particle.0933

It's the number of particles that matters; it doesn't matter whether it's sugar or salt or glucose or protein or something else.0952

The colligative properties don't care about what the identity of the species is.0959

All they care about is how many of those particles are floating around freely in solution, running interference with that solvent, because now the solvent isn't pure anymore.0965

It isn't just solvent molecules interacting with each other, it's solvent molecules that have a bunch of things floating around in it; and those things, the fact that they are there, they change the property of the solvent.0973

OK.0984

I should say one thing, but recall, very, very important: covalent compounds and soluble ionic compounds do not dissolve the same way.0986

Covalent compounds, it's one for one; one covalent compound dissolved and becoming aqueous releases one free particle, but as salt dissociates it to free particles - so it just depends on what the salt is made of- when it dissociates one unit of salt, sodium chloride produces two particles.1079

One unit of magnesium chloride produces three particles.1099

One unit of aluminum chloride produces four particles.1103

It's the total number of particles that are floating around that affect the colligative particles.1108

Now, and again, I'll just repeat what I just said.1122

One molar glucose solution has 6.02 x 1023 free particles floating around in there; however, a 1M magnesium chloride solution has 3 x 6.02 x 1023 free particles.1129

This is what we have to remember: what is it actually that we're dissolving, what is the solute.1165

If it is an ionic compound that dissolves, we need to keep track of the number of things that it is actually producing, number of free particles that are floating around.1172

If it is covalent, it's just one; it's not a problem.1179

OK.1183

Pure water has certain properties, for example, its boiling point.1188

We know that water boils at 100°C - pure water.1203

It has a freezing point at sea level by the way, which again, we are here, we are not up in space or down under the earth or anything.1212

We know that water freezes at 0°C.1224

It has a vapor pressure of 24 Torr, Torricelli, or 24 mmHg - it's the unit of pressure.1230

I'll just go ahead and write Torr at 25°C.1245

If I take a glass of water and I cover it up, well, at 25°C, some of that water is actually going to escape into the gas phase because it's not going to be just pure liquid.1257

There is enough motion, there is enough heat, there is enough energy in the water to actually kick off some of the water molecules in the gas phase.1267

Here, let me draw this out.1276

This is the liquid water; however, some of the water molecules are going to escape in the gas phase.1279

The ones that are here in the gas phase, they're bouncing around the walls of the glass; they create a certain pressure- that's what the vapor pressure is.1288

At certain temperature, there are certain parts of the solvent that actually exist in the gas phase in equilibrium with the liquid phase.1298

It contributes a little bit of pressure- that's what vapor pressure is.1309

It's not going to concern us, but I just thought you should know where this comes from.1312

OK.1317

Now, here is the interesting part.1318

Adding a solute - in other words, free particles - changes the values of these properties.1323

So, just by virtue of dropping in some salt into water, all of a sudden I change its boiling point now, of the solution.1349

Now, the solution has a new freezing point and it has a new vapor pressure.1355

Now, the solution has a new boiling point, freezing point and vapor pressure.1363

OK.1380

Now we come to osmotic pressure.1381

Adding a solute to a pure solvent- in other words creating a solution - it creates a new property for the solution.1385

That property, we call osmotic pressure.1419

And, this is what we are going to talk about now, osmotic pressure- profoundly, profoundly important in biological systems.1426

Osmosis is ubiquitous in biological systems.1432

Well, I'll go ahead and talk about it now and we'll talk about it more biologically later.1439

Here is what happens.1443

I'm going to draw something here.1444

I'm going to take a little bit of something called a "U tube", something like that.1448

It's just a tube that is in a shape of a U, and down at the bottom, I'm going to have something called a semi-permeable membrane.1457

OK.1468

Let me go ahead and label that.1470

This is a semi-permeable membrane, and what that means is that solvent molecules can pass back and forth through that membrane, but solute molecules cannot pass.1472

It's like a filter is what it is: allows certain molecules to pass, others not to pass.1484

In this case, it allows the solvent itself to pass but nothing else.1489

So, this is a semi-permeable membrane- like a cell wall.1497

This is the model for a cell wall.1503

A cell wall is a semi-permeable membrane; certain things can pass naturally, other things cannot pass without being given permission.1508

OK.1516

On one side, I have pure solvent. 1517

This over here, this is going to be pure solvent.1521

Over here on this side, this is going to be the solution.1528

And of course, because it has solution, I'm going to use Xs to designate solute particles, so we have a bunch of solute particles floating around in that solution.1537

OK.1547

Here is what's interesting.1548

I was going to go ahead and write this out, but I think I'd rather just say it and describe it because I think it'll make more sense.1550

Now, with this semi-permeable membrane, when I create this situation, and now, here is what happens.1555

Intuitively, you have a sense on what's going to be going on.1562

These solute particles, there's a whole bunch of particles here in this solution and there's nothing over here.1565

By nature, these solute particles are going to try to distribute themselves evenly across all of these volume that they have available, but this is a semi-permeable membrane, these solute particles can't pass through this membrane, so what happens is that it actually induces water from the solvent side, that way, it actually pulls water into it in order to dilute this until there is an equal distribution.1571

However, as water passes into this compartment, well, this water level is going to drop because water is passing and it's going to reach a new level.1600

Actually, let me do this in red.1614

This water level is going to drop and it's going to come down here, and of course this water level is going to rise until it comes to about here.1623

OK.1632

Now, when a solution is separated from a pure solvent by a semi-permeable membrane, the solute particles want to naturally distribute themselves, but they're not going to so what they end up doing if they can't go to the water, they are going to pull the water to them so water ends up moving across this membrane into this compartment and the volume of water rises.1633

It's going to rise, it's going to rise, it's going to rise but at a certain point, it can only rise so far because now this water has weight, this extra water that is coming in it has weight so what it's doing is it's actually pushing down on this thing, trying to keep it from rising. 1657

So, there's this water moving in this way, pushing the water level up, but at some point there is the extra weight of water that's actually now pushing it down; there comes a point where it reaches an equilibrium.1676

Once this stops that level and that level, what we define is the osmotic pressure, is the pressure that's required to actually keep this water flow from actually happening.1687

OK.1700

In other words, the volume of the solution, of the extra water keeps more water from coming in.1702

At some point it reaches an equilibrium where this weight is pushing down, this is pushing this weight trying to move in, well, the amount of pressure that actually keeps water from moving in to begin with, we define that as the osmotic pressure.1708

It's a measure of the extent to which the solution is actually pulling water into it in order to dilute it.1725

That's what it is, and we can actually measure this, so it is an osmotic pressure.1734

It's as if this water is actually pushing this solution that's why we call it osmotic pressure but really what it is, it's the pressure that I have to maintain on this to make sure that no water passes across the membrane.1740

I hope that makes sense.1757

OK.1759

Now, like I said we have a way of actually representing this numerically and that's what's nice, we want some number that we can use.1761

So, let's go ahead and define our osmotic pressure.1771

It is equal to iMRT or osmotic pressure, we use a pi symbol, you'll see that, iMRT.1776

The variables that you use don't matter as long as you understand what it is that the variables represent.1792

Let's go ahead and use this one because that's probably the one you see in your book.1797

It's pretty standard now to have osmotic pressure represented by this symbol π.1800

OK.1807

So, let's go ahead and talk about what this means, so let me go to the next page, let me go to blue, actually let me go back to black.1808

We have the osmotic pressure is equal to i x M x R x T.1816

Let's talk about what these things mean, the i, the M, the R, the T.1822

OK.1826

M is the molarity of the solute.1827

R- that's the gas constant.1839

That is 0.08206 and the unit is Latm/molK.1845

Remember that from the ideal gas law?1857

T is the absolute temperature, in other words, the temperature in Kelvin - very, very important - has to be in Kelvin; it's what absolute temperature means.1860

Now, i is something called the van't Hoff factor and this is really, really easy.1873

Van 't Hoff factor is just the number of particles produced upon dissolving.1882

So, let's just do a couple of examples of i just to make sure that we understand that part.1898

OK.1909

So, glucose, C6H12O6 solid, when it dissolves in water, it produces C6H12O6 aqueous.1910

It doesn't come apart.1924

One molecule releases one free particle; one mole releases one mole.1925

i = 1, it's the number of free particles produced upon dissolving.1931

Magnesium chloride, solid, when it dissolves you produce 1 magnesium particle and you produce 2 chloride particles: i = 3, 1 + 2 = 3.1938

That's it, that's all that's going on here.1959

OK.1962

Let's go ahead and do an example.1963

Yes, it's fine, I'll go ahead and start here.1969

Example number 3: At 25°C, what would be the osmotic pressure of our 0.686M lactic acid solution?1975

Remember the solution from the previous lesson?2009

We want to know what kind of osmotic pressure it produces.2014

In other words, if I were to take this lactic acid solution and separate it via a semi-permeable membrane from pure water, what pressure do I need to apply to the lactic acid solution to prevent any water from actually flowing into it?2019

What is the extent to which it's actually pulling in water across the semi-permeable membrane to dilute its particles?2038

What would that pressure be?2044

Well, let's go ahead and work that out.2046

OK.2049

Let's see what we've got.2050

Well, again, in example it's is always nice to write out your equation so π = i x MRT or you'll also see it this way, iCRT.2051

C stands for concentration. 2065

Concentration has to be in mol/l as far as osmotic pressure is concerned but you're going to see C, you're going to see M.2067

Again, the letters, irrelevant, what's important is that you understand the properties.2074

OK.2079

Well, so lactic acid is a covalent compound where a covalent compound i = 1, the van 't Hoff factor.2081

So, the osmotic pressure equals 1 times its molarity, well the molarity is 0.686, that's mol/L.2089

I'm going to go ahead and use all the units so you see how they cancel.2101

R is 0.08206, that is Latm/molK, and then of course we're at 25°C so that is going to be 298K.2104

K cancels K, mole cancels mole, liter cancels liter, the unit we're left is atmosphere, perfect, it's a unit of pressure, everything is good,2123

When we go ahead and do this, we end up with the following: the osmotic pressure is 16.8 atm.2136

If I have a solution of lactic acid which is 0.686M, and if I actually separate that solution - let me go ahead and draw my other U tube here, OK - so here I have my lactic acid solution, here I have just pure water and this is semi-permeable membrane that allows water to pass and not solute particles.2145

So, I have the water level, I have a bunch of lactic acid particles.2170

Now, 16.8 atm, what that means is that I need to apply 16.8 atm of pressure on top of this solution to prevent water from actually flowing from the pure solvent into the solution side- that's the osmotic pressure.2177

Simply by virtue, the fact that I have a bunch of particles floating around here, a solution, versus no particles around here, it actually causes water to be pulled in to that solution.2197

The pressure that I have to apply, because again, I need something physical that I can measure, so that's why this is osmotic pressure, I need to apply - that's a hell of a lot of pull, that's very, very strong tendency to pull - I need to apply 16.8 atm of pressure to prevent that from happening.2208

16.8 atm of pressure is no joke, that's a hell of a lot of pressure.2225

There you go, that's osmotic pressure.2231

Now, a little bit of information: this is 16.8 atm, this is just a little more than twice the osmotic pressure of blood, and the osmotic pressure of blood happens to be 7.7 more or less, 7.7 atm.2234

Then again, we have other units of pressure but atmosphere, because of the R, so osmotic pressure will often be expressed in atmosphere.2277

We can convert later - it's not a problem - to Torricelli, or kilopascal or pascal or whatever we need, but it is expressed in atmospheres.2285

Blood has a lot of things dissolved in it, so if you separate blood from water by a semi-permeable membrane, it will actually pull the water across that membrane into it in order to dilute it.2293

7.7 atm, that's how much pressure I have to apply to the blood to keep the water from flowing in.2309

7.7 atm is a hell of a lot of pressure.2315

OK.2321

So, that's dilution and osmotic pressure.2322

In the next lesson, we are actually going to continue our discussion of osmosis because there is a little bit more to say about it.2324

Thank you so much for joining us here at Educator.com.2329

We'll see you next time, bye-bye.2333

Hello and welcome back to Educator.com and welcome back to Biochemistry.0000

Today's lesson, we're going to start talking about polysaccharides.0004

These are just long chains of individual sugar units, glucose, galactose, mannose any number of possible, so, it is just long chains of them- that is all.0008

Instead of a disaccharide, so now we have multiple monomers.0019

Let’s go ahead and get started.0023

Before we do though, I just want to do a quick recap, one more example for disaccharides just so we can get a little bit more practice with actually drawing the structures out by hand.0025

We need to be able to draw them out by hand, not just by recognize them passively.0033

Let's do a recap example here before we get started.0039

OK, so, a recap example.0047

We would like you to draw the structure of galactose - oops let me make this L a little bit better here - of Gal(alpha1)(beta1)Man.0051

Galactose, alpha-1, beta-1, so the glycosidic bond between the galactose and the mannose is going to be the alpha-1 carbon of the galactose, the beta-1 of configuration of mannose.0074

Let's go ahead and draw this out.0085

Let's see; the first thing we are going to do, I’m going to start off by drawing the linear structures and then the rings, and then I'll put the rings together.0090

It is always a great way to do it like this; this way, you are always nice and systematic.0097

Galactose and mannose are both hexoses, so we have 1, 2, 3, 4, 5 and 6.0102

Let's make them a little big here.0110

OK, we have got OH, OH, OH, OH and CH2OH0114

This is going to be our alpha-galactose.0123

Actually, not alpha yet, it is just galactose because the alpha and the beta configurations are when they are actually in a ring.0127

So, you notice we have right, left, left, right, 1, 2, 3, 4; galactose is the c-4 epimer of glucose.0134

OK, this is our galactose.0143

Now, we are going to put it together with our mannose.0146

Again, we have a 6-carbon sugar, 1, 2, 3, 4, 5, 6; there we go.0149

We start with our aldehyde; let’s go ahead and put our CH2OH of the other end, and now, we can build what's in between.0156

This is going to be...no, this is mannose, right?0162

OK, mannose is a c-2 epimer, so this is going to be OH, OH, OH, OH.0167

You notice the 2 on the left, 2 on there right; this is our mannose.0175

OK, now, let’s go ahead and draw the ring structure for them0181

And again, what you do is you take this linear structure which is vertical; you rotate it to the right, and then you bring this side around the back, and then you make a little bit of a rotation, so that the OH is actually pointing to the right and the CH2OH is pointing up, and you'll see what it looks like in just a minute, and when you put it together, you get the following.0183

Yes, so we have - let me, yes, that's fine - so this, this, this, like that, and we have the alpha-configuration, so this hydroxy is down.0209

This hydroxy is down; this hydroxy is up, and this hydroxy is up, and, of course, we have our CH2OH.0223

That is what we did; now, we have our alpha-galactose aGal.0231

And now, let's go ahead and do our mannose.0237

Again, rotate it to the right; bring this aldehyde down to the right.0240

Now, this CH2OH, bring it around back, and then rotate this carbon right here, the no. 5 carbon, rotate this one.0245

So, the hydroxy is pointing to the right, and the CH2 is pointing up.0254

That is the deconfiguration, and when you do that, what you get is the following.0258

I'll go ahead and do this one in red.0263

We will draw the general 6 carbons, and we said this is the beta-mannose, correct, beta-1.0266

Beta, this is going to point up, right?0273

And then, looks like this one is also going to point up.0278

That is this carbon, and then on this one is going to go - let me see, wait, now, I'm lost - this is going to be up; this is going to be up.0281

This is going to be up; OK, sorry about that.0293

This is no. 1 carbon, no. 2 carbon, no. 3 carbon, 1, 2, 3.0295

So, we have taken care of those, and now, we have this one; this hydroxy is going to be down, and this is CH2OH.0299

As you can see, it is very, very important to keep track of which carbon we are looking at; this thing can be very, very confusing, so all the more reason to do it nice and systematically0307

OK, this is going to be alpha-1, beta-1.0315

In blue, we are going to be connecting this carbon with this carbon.0320

What I'm going to do is, this mannose, in order to draw it out and bring this carbon in close proximity with this one, I'm going to have to flip this.0323

Now, there are 2 ways that I can flip this molecule; remember we talked about spin and flip?0330

I think flip is probably the best way to go.0336

It seems to be the one that you see more often in biochemistry text books as opposed to spin, but you realize, this is a flat molecule, right?0338

So, we have this right here; let me go ahead and draw, so you can see.0348

This is Haworth projection; this is a flat molecule.0352

You are looking at it like that; you can flip it two ways.0355

You can flip it that way, or you can flip it that way, right?0358

There are two ways you can flip something, either side to side or forward and back.0364

In this particular case, let me go ahead and do this in blue.0368

I'm going to go ahead and flip it, and what I'm going to do is, I'm going to flip it sideways.0373

In other words, I'm going to bring this carbon here, and I’m going to bring this carbon over there.0377

OK, so I'm going to flip it sideways, that way, not this way, forward back.0384

When I do that, it is the oxygen that tells me how the flip has happened.0390

This is the thing, wherever the oxygen is, that is what tells me where to put the other substituents- the hydroxys and the CH2OH.0394

So, when I do the flip, I end up with this ring structure.0402

Now, the oxygen is on the back left.0407

OK, and now, let’s see what it is that I've done.0411

This OH, now, this is the no. 1 carbon over here, 1, 2, 3, 4.0414

Now, the no. 1 carbon is over here, and this is the no. 4 carbon.0419

OK, let me go back to blue.0425

OH is down here; this OH is going to be up there.0429

These have been flipped, so now, these are both down- that hydroxide and that hydroxide.0433

This one has been flipped over to the other side, and it is also down, so this is going to be over here, CH2OH.0438

Now, that is the arrangement.0447

Now, we are going to put this thing and this thing, so this is beta mannose that has been flipped.0450

Now, we are going to put this together with this; we are going to connect this carbon with this carbon, and when we do that - let’s go ahead and just draw our little equilibrium arrows - our final product is going to look like this.0459

I am going to do this in black, actually.0478

We have that, and remember, we do our little arrangement like this except this time, the O is over here, like that.0481

We have this OH; we have this OH.0498

We have this OH; this is in standard configuration.0501

The oxygen is on the back right, and over here we have flipped it, so now, the oxygen is on the left.0504

That means this hydroxide is here; this hydroxide is here.0510

This hydroxide is here, and this CH2OH group.0514

It is always interesting to try to draw it; I will go ahead and put the H2 there and the OH, and there we go.0520

This is our Gal(alpha1)(beta1)Man.0526

There you go, nice and systematic.0534

Draw out the linear structure; rotate them.0535

Create the ring structures in standard configuration with the oxygen on the back right, and then decide which carbons you are going to have to connect, and then decide which one of those monomers you are going to have to flip.0538

OK, flip, it is up to you.0548

You can do a flip or spin, as long as the arrangement of the substituents is such that it is very, very clear what is where.0550

If you want, you can go ahead and add a little stereochemistry by doing that, darkening up some lines.0557

Let me do that; I personally do not.0564

I just sort of leave it like that, but, of course, your teacher might want you to actually demonstrate the projection by showing the darker lines.0568

So, there you go; that is it- nice, basic disaccharide0574

As long as you know the structures of the monomers, which I imagine your teacher is probably going to have you memorize, everything should be nice and straight forward.0578

OK, let’s start our discussion of polysaccharides.0586

I will go ahead and I am going to do this in blue.0591

Polysaccharides, now, we are just going to be adding a whole bunch of monomers, one after the other on a chain, just like we did with proteins, except those were amino acids instead of sugar units.0595

Polysacchs- they are also called glycans, and this glycan name will come up.0608

In case it does, it is not a different type of molecule; it is just another name for a polysaccharide.0623

Now, polysacchs, they differ from each other - there is a whole, whole, whole bunch of polysaccharides - with respect to 4 things.0628

The monosacch units that actually make up the polymer.0654

Which monosaccharide units are we using?0658

Are we using only glucose or are we using glucose and galactose and mannose and n-acetylgalactosamine?0659

Which monomers are we using?0668

Chain length, you might have a polysaccharide that is only 15 monomers long.0671

You might have one that is 150 thousand polymers long, so chain length.0675

And, if you have, let's say, a bunch of glucose that is 15 long and a bunch of glucose that is 1500 long, those molecules are going to behave differently, just because they are made of the same monomer, glucose, the length will actually change the chemistry.0682

Branching along the chain - I'm sorry, branching along the, well, yes, branching along the chain.0700

What you are going to have is something like this.0716

You are going to have some monomer going on, going on, and all of a sudden it is going to branch off like this, and maybe branch off again, and then maybe branch off again.0717

Polysaccharides will do that, and we will see some examples in just a minute.0725

And, of course, the last thing that they differ with respect to is the nature of the glycosidic bond connecting the monosaccharides.0729

In the example that we just did - monosaccharides, just let me go ahead and write this out - our connection here was alpha-1, beta-1.0751

This is the no. 1 carbon; this is the no. 1 carbon on the mannose.0762

So, and alpha-1, beta-1, well, maybe if I had an alpha-1, alpha-1 mannose, that is going to be an entirely different polysaccharide simply by virtue of the nature of the glycosidic bond.0765

Totally different, totally different chemistry, totally different folding- that is the whole idea.0775

Small subtle changes make huge differences because you are talking about big molecules, and when all of these things sort of add up, you get entirely different chemistry.0779

OK, define a couple of more terms.0780

A homopolysaccharide is exactly what you think it is.0798

It is a polysaccharide made of 1 type of monomer.0805

I'll just write "1 type of monomer makes up the chain".0810

In other words, it is just the same monomer one after the other, glucose, glucose, glucose, glucose, glucose makes up the chain.0819

That is a homopolysaccharide, and, of course, heteropolysaccharide, you have 2 or more; but for formality's sake, let's go ahead and write it down.0826

A heteropolysaccharide- 2 or more monomers make up the chain.0835

Maybe you have an alternating glucose, mannose, glucose, mannose, glucose, mannose.0854

That is a heteropolysaccharide; there happen to be 2 of them.0858

There can be more.0861

OK, now, polysaccharides serve lots of purposes.0863

What is really, really exciting is glycobiology, the study of carbohydrates, it is a fantastic, fantastic area of research right now because every single day, literally, every single day, some new polysaccharide, some new protein attached to a carbohydrate, is being discovered that serves a whole different purpose; and that is what's amazing about this.0869

Polysaccharides, they serve many purposes; and certainly, many of the purposes we haven't even discovered yet.0894

There is plenty of room for growth in this particular field, and polysacchs, interestingly enough, they have no specific molecular weight.0900

They have no specific molar mass.0919

In other words, we don't talk about a polysaccharide that has a molar mass of 50,346g/mol.0924

It is not that precise; it is not like proteins where you have a specific number of amino acids if you have 1 more or 1 less.0933

It is an entirely different protein for the most part.0940

Polysacchs, when they are synthesized, we talk about a polysaccharide that is roughly 25,000 monomers long, 300 monomers long, more or less.0943

There is no specific molecular weight.0954

OK, now, let's talk about some homopolysaccharides that serve as a fuel storage.0958

One of the purposes of polysaccharides is as a reserve fuel source if the organism is not actually taking fuel in.0965

In our case, we tend to store fuel as glycogen; that is our primary polysaccharide for animals.0974

For plants, it is starch, so homopolysacchs serving as fuel storage.0981

And, we are going to talk about starch, and we are going to talk about glycogen; and both occur inside the cell.0999

Now, starch and glycogen are actually the same thing; they are made of the same thing- glucose.1017

It is just the degree of branching, as you will see in a minute, that is going to differentiate the glycogen from the starch- an entirely different chemistry.1022

It is actually amazing.1028

Let's talk about starch first.1030

Starch has 2 types of glucose polymers.1035

Starch is made of 2 types of polysaccharide chain.1048

One of those is called amylose or amylose; however you want to pronounce it.1053

It is glucose monomers connected by alpha-(1,4) glycosidic bonds, just one glucose after the other with the connection as alpha-(1,4), alpha-(1,4), alpha-(1,4) all the way down the line in just a straight, single chain.1060

Now, the other particular polymer for starch is amylopectin, and it is also glucose connected by the alpha-(1,4).1085

It has that chain, but along that chain, there are branch points; and those branch points - and, alright - and branching by alpha-(1,6).1100

So, at the no. 6 carbon, that CH2OH sticking up, it actually branches off at that point, and it starts a whole new chain.1121

And then maybe along that chain, it branches again; and it starts a whole new chain, so that is the difference.1131

Amylose and amylopectin, and they are sort of intertwined; and again, you will see it in a minute.1136

We are going to do a detailed structure, and then, sort of a broader structure.1141

OK, now, let's see what else do we want to talk about before we actually start looking at some structure.1146

Ah, yes, so, we talked about reducing sugars, non-reducing sugars, there is a reducing end, in other words, a free anomeric carbon that can react, that can be oxidized by iron ion or copper ion.1154

There is a reducing end, so polysaccharides, polysacchs, have a reducing end and many non-reducing ends because of the branching.1172

And this idea of the non-reducing end, having many of them, is going to play a very, very important role in physiology as we will talk about at the end of this lesson.1198

OK, glycogen, just one quick word about glycogen, it is the same as starch.1208

OK, in other words, it has the amylose; it has the amylopectin, but it is more highly-branched, and more compact.1220

Again, totally different chemistry simply by virtue of the branching.1240

Amylopectin, it might branch off maybe every 30 to 35 glucose units, 30 units and then it will branch off, 30 units and it will branch off.1244

Glycogen might do 10 units branch, 10 units branch, 5 units branch, 7 units branch.1256

It branches more often, and it tends to be more compact, more dense.1262

That is it; that is the only difference between the 2, but the fundamental structure is the same.1267

Alpha-(1,4) glucose units and alpha-(1,6) glucose, glycosidic bond at the branch point.1271

OK, let's go ahead and take a look at some structures here.1281

In this particular case, I am going to be presenting them as illustrations instead of drawing them out by hand simply because we want to save a little bit of time, but now, I think we have a little bit of a sense of what the glucose looks like, what the monomers look like, alpha-1.1285

We just want to be able to identify what is what, what is connected to what.1297

Let's take a look at some pictures, and the first one we are going to look at is amylose.1302

OK, we have our amylose, and we said that we have alpha-(1,4) and its glucose.1310

So, we have 3 monomers of glucose, so it goes off in this direction.1319

It goes off in this direction.1321

Let me go ahead and use, yes, let me go ahead and stay with red here.1323

Here is our 1,4; this is our no. 1 carbon, alpha-configuration.1328

This is our no. 4 carbon on the other.1333

This is our no. 1 carbon, alpha-configuration, the hydroxy is down; and this is our 4 carbon from the other side.1336

This is it; this is amylose.1344

I don't think I'm going to have to...that is fine, I'll just go ahead and...not a problem.1348

This is our amylose chain, and it goes off this way, and it goes off this way.1351

Now, at the end, of course, this is your reducing end.1355

The polymer will usually go on in that direction connected to this hydroxy.1361

Let me go ahead and actually do that.1366

It will end up being connected to this hydroxy.1369

Here is the reducing end; this is the non-reducing end.1371

That is it- nice, basic structure.1375

Glucose units, down, up, down, CH2OH, CH2OH, oxygen is in the back right, oxygen back right, oxygen back right- this is a nice, good, very, very well-behaved polysaccharide.1378

We did not have to flip anything; we did not have to spin anything.1392

That is good; this is the Haworth projection that you see.1395

Now, of course, you know the hexose rings, they assume chair conformations.1401

They are not flat like this; they are not like benzene.1405

Benzene is a flat molecule, these are not.1408

I wanted you to see what this looks like in actual configuration, in actual conformation, I'm sorry, conformation.1410

These hexose rings actually assume chair conformations, and here is what the glycosidic bond looks like.1417

This right here is actually this right here.1425

So, we have our 1 and our 4 carbon, our 1 carbon, our 4 carbon, 1 carbon, 4 carbon, and, of course, it goes on like that.1429

You see this little stair step pattern, this is how it looks.1437

Now, again, it is going to be up to your teacher whether he or she wants you to draw it like this in this projection, or whether he or she actually wants to see at least 2 or 3 units in the chair conformation.1442

I will leave that up to your teacher, but again, what you want to notice is the arrangement, oxygen back right, oxygen back right; here is your CH2OH.1457

Notice, here, they actually wrote the C; here, it is just 2 lines coming together at a point, at a vertex.1466

So, that is a carbon; that is a carbon.1474

That is a carbon, nothing new here.1475

Equatorial, axial- that is what you have to watch out for.1478

OK, start with this projection, and then, go to this particular rendering, this particular representation.1482

OK, now, let's take a look at amylopectin.1490

I think that is going to be the next, yes; this is a little piece of amylopectin.1494

Let's go ahead and identify our 1,4.1499

So, we have our 1,4, 1,4 .1504

This is our glycosidic bond, glycosidic bond; and now, we have our 1 and 6.1507

There you go; that is your branch point.1514

At the no. 6 carbon of a particular glucose monomer, that oxygen has reacted with the anomeric carbon of another glucose, and it has started the chain, a second chain.1516

Now, this chain is going to go off in this direction, and then, this chain is going to go off in this direction; and they are going to parallel each other.1528

And, as you will see in a minute, they don't just parallel each other, they actually wind around each other in a helical pattern.1535

That is it; that is the only difference.1542

You have the 1,4 configuration, and in amylopectin, your branch points are at 1,6.1545

So, maybe a little further down the line, there is another 1,6.1550

That is it, alpha-(1,6).1555

This is alpha, because the hydroxy is pointing down.1558

There you go, and, of course, this is another representation of it with just a couple more.1562

Here, we have 3 and 1; here, we have 3 and 2.1569

You see our 1, our 4, our alpha-1, our 4.1572

This is alpha-1; this is our 6, and then, of course, it continues on.1577

This is alpha-(1,4), that is it.1581

This is amylopectin, nothing going on here; monomer, monosaccharide, disaccharide, now, it is polysaccharide.1585

What is important are the individual monomer units; if you understand those, you can build any polysaccharide you want.1592

That is the whole idea; that is what we want you to be able to do.1599

OK, so, now, let's take a look at...OK, here we go.1602

So, we talked about the actual amylopectin clusters.1610

This is amylopectin; this is a macroscopic.1615

We started over here; we have some chain, and then, this one branched off, and then it branched off again, and then it branched off again.1621

And then now, when the whole branching thing stopped, these 2 that were paralleling each other, now, they start to wind around each other in a coil.1628

What you end up with is, once everything is built, you end up with something that looks like this.1639

That is it; this is a nice cluster of amylopectin.1645

That is all.1649

This, right here, this is a detail of those 2 strands.1650

Once it branches off in the 1,6, they parallel each other, they actually start to wind around each other, so this is 1 strand, and this is another strand, and they intertwine.1653

That is it; they intertwine.1666

There is nothing in between them; they just intertwine.1668

It is like the backbone of the DNA, a nice helical pattern, and that is represented here.1671

Let me just write, this is an amylopectin, helix.1678

That is it, nothing strange happening here.1685

OK, let's actually, let's draw something by hand; let’s see.1690

Again, we want as much practice as possible.1698

Let's draw an alpha-(1,4) and an alpha-(1,6) of amylopectin just for a little practice.1705

Pretty much what we just saw, let's just draw it out by hand, so we know where to put everything.1718

OK, let's do our glucose down here.1728

Let's go 1,4.1733

This is going to be alpha-(1,4), so it is going to be like this, like that.1740

This is our 1,4 linkage, so this is alpha-1, and this is 4.1749

Let me go ahead and put that there, this, there, this, there, here.1755

I have got the CH2; this is going to be my 1,6.1763

Let me go ahead and put this as CH2OH.1767

OK, 1,6, this is the no. 6 carbon.1771

Let me use black.1776

This is our no. 6 carbon, and now, this is going to be connected to a - go back to red - another alpha-(1,6).1780

Alpha means that the hydroxy is below the ring.1794

It is going to look like this, and, of course, this is - nope, that is not there - this is glucose, so this is down.1798

This is up, and this is going to be off connecting that way, and chances are...you know what, I am going to go ahead and connect this one off too.1809

This is going to go that way, and that is going to be running off, and then, this is going to be CH2OH.1824

There you go; here, we have our nice amylopectin branch point, glucose, alpha-(1,4), glucose, alpha-(1,6).1831

This is the alpha-1; let me go ahead and erase these and write them in black.1843

This is the alpha-1, and this is the no. 4.1851

That is it; there should be no problem at this point.1855

Hopefully you have had a little bit of practice, and you should be able to just knock them out given the particular configuration at the glycosidic bond.1857

OK, let's see what else we can do.1866

OK, now, let's talk a little bit about glycogen.1871

Let me go ahead and...a page here.1874

Now, glycogen, let's see, you haven't eaten for a certain number of hours, and your body needs some energy.1878

Well, if your body doesn't have any energy in terms of the food that you put into it, it is going to go to its next readily available source of energy, and that is the glycogen that is stored in your cells, that is stored in your body, mostly in your liver.1892

That is the first place that it is going to go in order to break off glucose units and send those glucose units into the bloodstream and out to the other parts of the body, so that your body and brain can function.1907

That is it; glycogen is essentially just a readily available, very quick storage for available fuel that gets it into your body right away.1920

Glycogen is hydrolyzed - in other words, it is broken up - from the non-reducing ends, the left ends.1934

Remember we had that little cluster; well, I will draw it out in just a minute.1957

OK, now, since - excuse me - there are so many branches and we said the glycogen is more heavily branched than starch, several glucose monomers can be cut off simultaneously in order to supply the body with glucose, with individual glucose monomers.1962

Now, let's go ahead and draw what that looks like.2016

Let's do a little bit like a main chain, something like this.2019

So, I have got another branch here, maybe another branch, another branch, another branch, another branch.2024

I'm just going to draw a whole bunch of branches like this, like that, like that, like that.2030

Each one of those, at some point, it terminates; and what you have are these, you have glucose at the ends.2036

These are the non-reducing ends right here; of course, it is a lot more compact than this.2044

This is the reducing end right here; this is the reducing end.2049

Now, when your body needs glucose, glycogen has evolved to take on this particular structure because what it can do is, once it needs the glucose, it can just go and cut off these individual ones.2054

Instead of one long chain, where it has to go cut this one, then this one, then this one, then this one, then this one, then this one, it is going to take a certain amount of time.2070

Yes, enzymatic reactions are very, very quick, but still, it is going to take a certain amount of time; but if it can take a whole bunch of these off simultaneously, hundred of thousands of them, millions of them, and just deliver them into the body, for the time it takes to cut off one, well, it can cut off several hundred thousand, and deliver them into the body.2078

That is why glycogen has the structure that it has, highly branched, so it has a whole bunch of glucose monomers available to it all at once because you are going to need that fuel all at once.2100

That is what's happening.2113

OK, now, let's talk a little bit more about glycogen and concentration, glycogen, which is stored as insoluble.2115

That is the key word here, insoluble granules, and there is probably a picture of it in your book.2135

You will see a cell with the little dots, the little black dots.2141

Those are granules of actual glycogen, granules in the cytosol.2146

Cytosol is the intracellular fluid.2154

OK, so, glycogen which is stored as insoluble granules in the cytosol - excuse me - contributes nothing to the osmolarity of the cytosol because it is insoluble, it is solid.2158

In other words, it doesn't have, there aren't free particles of glycogen floating around in an aqueous environment; it is not dissolved.2182

Now, liver cells store glycogen equivalent to about 0.4M free glucose.2193

In other words, if I were to take all of the glucose, all of the glycogen, and break it up into individual monomers, the amount of glucose that is there actually accounts for about 0.4M because glucose is soluble; glycogen is not soluble.2218

Now, if glucose were stored as monomers, just sort of floating around in the cytosol, the osmolarity of the cytosol would go through the roof.2234

The osmolarity of the cell, well, 0.4M, the osmolarity, there are going to be so many more particles inside the cell than outside the cell.2265

It is going to cause the liquid to flow from outside the cell into the cell causing the cell to burst open.2281

OK, it caused fluid to flow into the cell and rupture it.2289

The body definitely knows what it is doing; it needs to have a supply of glucose, but it can't just have free glucose floating around in the cell, in the cytosol because then, the osmolarity of the cell would be so high that it would cause a difference in osmotic pressure inside and outside.2309

Osmosis would pull water into the cell, and the cell would just explode.2326

So, by storing it as insoluble glycogen, it is there.2332

It is insoluble, so it does not contribute anything to the osmolarity, but it is readily available; and the particular form of glycogen as such that all of those glucose monomers are available very, very quickly- fantastic, absolutely fantastic, extraordinary molecule, extraordinary molecule.2337

That is what's going on with glycogen and starch.2356

OK, thank you for joining us here at Educator.com2360

We will see you next time, bye-bye.2362

Hello, and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today, we are going to continue our discussion of polysaccharides, and talk a little bit about cellulose, another polysaccharide; and we are also going to introduce these things called the glycosaminoglycans.0003

Let's get started.0015

OK, as far as cellulose is concerned, I thought we would introduce that with an example, get a little practice, a little more practice with drawing structures.0018

I hope I am not making you crazy with drawing all these structures.0028

It is very, very important to be able to handle them without any problems.0031

OK, let's start with, cellulose is a structure - remember we said that polysaccharides have many different things that they do.0035

One of the things was fuel storage with starch and glycogen.0054

Well, structurally, some polysaccharides …cellulose is one of the structural polysaccharides, homo, and it is a homopolysacch consisting of glucose, and this time, we have a beta-(1,4)configuration, beta-(1,4) glycosidic bonds.0058

We had alpha-(1,4) for starch and glycogen; now, we have beta-(1,4).0100

Just by changing the configuration- totally different molecule, totally different chemistry.0105

OK, what we want you to do is to draw a trimer - 3 individual monomers - in a Haworth projection.0110

OK, let's go ahead and draw our monomers.0126

We know we are dealing with a homopolysaccharide.0130

Let's go ahead and do this in blue.0132

So, it is just glucose, so let's draw out 3 glucose monomers.0134

We should be all pros at this already.0140

Beta-configuration, beta means we have the hydroxy up here.0146

Let's go ahead and draw each glucose before we move on to the next glucose.0148

And then, we have another one.0155

That is there, and beta-configuration, and this.0159

Oops, let me work from right to left.0164

This is down; this is up.0168

This is down, and this is CH2OH; and, of course, we have one more.0169

Here we go, and this is beta.0176

This is down; this is up.0184

This is down, and we have CH2OH.0186

And again, I tend to not draw the thickened lines simply because that is just sort of a habit that I have gotten into, but if your teacher wants them, they are there, if not, that is fine.0191

Here is going to be the connections.0204

Beta-(1,4)...let me go ahead and do it in black.0208

I have got my 1 carbon, my 4 carbon, my 1 carbon, my 4 carbon.0212

What is happening is the elements of water - this is a condensation reaction - the elements of water are going away.0217

This oxygen is going to connect to that carbon.0230

This oxygen is going to connect to that carbon.0234

This is going to be our beta-(1,4) glycoside bond.0237

Now, let's go ahead and draw that; I am going to draw this one in black.0241

I will go boom, boom, boom.0248

I am going to go ahead and draw in the trimer, and then I will go back and put on the individual substituents.0251

This is O, like that.0258

I should probably draw it a little bit bigger than that; sorry about that.0267

And then, of course, we have this, and this is going to go on that way; and this is going to go on that way.0272

Now, we can go ahead and fill it in: OH, OH, CH2OH.0283

We have OH, OH, CH2OH.0292

Let's make this one a little bit better, and we have OH, OH, and CH2OH.0297

There we go; this is our basic trimer arrangement.0308

Notice they are all like this, and this is the glycosidic bond.0311

We have our beta-(1,4), beta-(1,4).0316

This is the beta right here, this and this.0321

We have drawn it, the bond this way, the bond this way, to show that this oxygen is actually up, beta; and this one is a down.0324

Now, I am going to go ahead and draw this not in another configuration, just a different representation of this actually showing the bonds a little bit more directly.0330

This is just another way of drawing it, and you are welcome to do it like this.0344

It is not a problem.0347

OK, so, we are going to have something like this.0349

I am going to start on this side, and let me do this in blue.0352

I have got, that is that; I have got O.0357

This is O; here, this is O.0370

This is there; this is there, something like that.0375

And, of course, it just sort of goes on like this.0380

That goes that way; that goes on that way.0384

And then, of course, we have our OH, OH, CH2OH, OH, OH, CH2OH.0389

All I have done is I have actually represented.0402

Instead of drawing them all straight like this, I have actually shown them in a sort of a stair step pattern just to show that more directly to the eye that this is below, this is above.0405

Let me go ahead and finish my substituents here, and I will go ahead and label some carbons.0418

Let's do that in black.0424

This is our alpha-1, no, not alpha-1; this is beta-1.0426

This is beta-(1,4); this is beta-(1,4).0432

The hydroxy is above the 4; the oxygen is below.0437

So, this is another way that you can draw it if you want to.0442

OK, now, I will do 1 final representation in order to show the geometry at the oxygen of the glycosidic bond, oxygen of the glycoside bond.0444

In other words, in order to show the geometry here, it is often drawn like this.0480

This is probably how you are going to see it in your book, at least one of the pictures...drawn like this as follows.0488

And again, you know oxygen, water, this is not a linear molecule; it is bent because we have these electrons here.0498

So, the geometry is a bent geometry; this is a linear geometry.0505

The picture before this, we had these curvy lines showing the arrangement, but now, in order to show the geometry, we are going to have to flip some of these monomers around.0509

Let's go ahead and draw what that looks like.0521

I am going to draw this one in black.0525

I am going to draw a central 1 first.0529

I have got that, that, that.0533

OK, I have got O, and I have got O.0537

Let me go ahead and draw my central 1 in here, CH2OH.0541

Now, a little bit different, this is going to be there.0548

This time, I have got my O here, and I have got CH2OH.0554

I have got OH; yes, that is correct, and of course, this one is going to be up.0566

And, of course, this one is going to be...let's just go ahead and put that there.0574

Let's put that there, and now, we will go ahead and put our O here, and we have our CH2OH.0579

We have OH, and we have this O there, and that goes on.0593

I am missing an OH; yes, that goes right there.0599

So, notice what I have done; I have taken this one.0602

I have left it the same; let me do this in red.0606

I have taken this thing, and I have left it the same.0610

I have flipped this one down, in order that this bond, now, shows the normal geometry of oxygen.0614

By flipping it down this way, what I have done is I have brought the oxygen which is in the back right; now, it is in the front right.0623

Same thing, this one, I have flipped up.0634

I have just flipped it in order to show the geometry at that oxygen and that oxygen.0637

And again, same thing, what used to be an oxygen on the back right, is now, an oxygen on the front right.0643

In order that we can show the geometry, this is probably how you will see it.0649

And again, when you look at these things in your book, when you are looking through multiple structures, you want to make sure...a polymer of glucose can be any different kind of molecule, what is important is the nature of the glycosidic bond, alpha or beta, what carbon it is attached to.0656

And, what you are looking for in these structures if you are just looking at it as opposed to drawing it with your hand, you are looking for where this oxygen is in the ring.0675

That is what is going to tell you what molecule you are dealing with.0683

So, this one might happen to look like some other polysaccharide, a glucose, but you need to be very, very careful, and identify where the oxygen is and what the glycosidic bond is.0687

Here, you have your beta-1, and this is 4; and this is 4, and this is beta-1.0703

This is beta-1, and the reason it is beta-1 is, under normal circumstances where the oxygen is in the back right, the standard way we actually arrange things, the oxygen is up.0715

That is the beta-configuration, but by flipping it, now, the oxygen is below the ring; but the oxygen here is in the front right.0727

So, that confirms the fact that this is a beta-1 configuration.0734

Just by looking at the hexose, it looks like it is an alpha because the oxygen is below; but the oxygen is here.0737

The oxygen in the ring is down here, not back there, so this is a beta-configuration- very, very important.0743

OK, let's see what we have got.0751

Let's take a look here at some actual illustrations.0755

We have our standard here, beta right?0760

This is the beta-carbon; this is the beta-1, and over here, this is our no. 4 carbon.0767

But notice, this time, we have left this particular one on the left in the beta-configuration under normal circumstances, the standard Haworth projection with the oxygen on the back right.0775

This time what we have done is we have taken the other one, and we have flipped that.0789

Notice where the oxygen is; the oxygen is in the front right.0793

These are the things that we have to recognize.0796

We know that glucose 1, 2, normally 1, 2, 3, 4, the oxygen is below the ring.0799

Here, the oxygen is above the ring.0806

Well, it is above the ring because I flipped the molecule, flipped it like this.0808

Now, this no. 4 carbon, the oxygen goes up.0812

Again, this shows the geometry at this particular oxygen.0816

These are the things that we have to watch out for.0820

They are going to be drawn in any number of ways.0824

What we want to do is look and see what is where, and that will tell us what is happening.0827

OK, this particular vision right there, this is the same thing, except now, it is going to show some of the hydrogen bonding that takes place among the different monomers in a polysaccharide or an oligosaccharide.0832

So, in this particular case, we have some hydrogen bonding taking place here.0847

We have some hydrogen bonding taking place here.0851

This is hydrogen bonding within the chain itself, and the hydrogen bonding, in addition to the geometry in the oxygen, is going to dictate how this molecule looks in 3 dimensions, how it folds, how it spins this way, how it turns this way, how it bends this way; and that is going to have an effect on the chemistry.0855

That is the whole idea behind the organic chemistry.0878

Structure is function- that is the whole idea.0882

What this thing looks like in 3-dimensional space - the final shape that it takes - is going to dictate what function it serves.0885

Now, this is an extended network of hydrogen bonding.0893

So, we have hydrogen bonding within an actual chain, but in another chain, there is also some hydrogen bonding going on here and here and here and here.0897

Again, all of this, the net effect is that all of this has an effect on the final structure of a cellulose molecule, if you want.0908

I mean it is just an extended polymer- it is what it is.0922

It is not an actual molecule, but there you go.0925

You have the glycosidic bonds; you have the particular arrangements.0928

You have the hydrogen bonds within, among the individual monomers in a chain; and you also have a hydrogen bonding between the chains, and that is the whole idea.0933

Again, you have got hydroxys all over these carbohydrates, so clearly, there is going to be a lot of hydrogen bonding.0942

It plays very, very important role in the structure of carbohydrates.0948

OK, let's talk about a different family of polysaccharides.0953

These are called the glycosaminoglycans- very, very important.0960

OK, now, let's go ahead and this is going to be a bit long, but it should not be too bad.0977

Now, these are heteropolysacchs; up until now, we have been talking about homopolysaccharides.0984

We have been using just glucose.0989

These are heteropolysaccharides of the extracellular matrix.0996

Now, the extracellular matrix is just a fancy word for that jelly-like substance that is outside of the cells, that tends to hold cells in place.1006

You cannot just have cells wandering around everywhere.1015

Certain tissues are cells that are there; they are held in place.1019

They don't move around; they are held together by this thing called the extracellular matrix.1025

It is basically just a scaffold for these cells to be; it keeps them in place.1029

That is all it is; that is all you want to think about it as.1034

They are heteropolysaccharides of the extracellular matrix, which is a gel-like substance - probably the best way to think about it, just a gel-like substance - that provides support for cells - in animal tissues, anyway - in animal tissue, as well as, provides a porous network for the movement of oxygen and nutrients, OK, oxygen and nutrients to individual cells.1038

That is it.1110

OK, now, the glycosaminoglycans, I am going to actually abbreviate this as Gag.1113

You will also see this in your book.1127

The glycosaminoglycans, they form a family - linear heteropolysaccharides, heteropolysacchs - composed of repeating disaccharide units.1129

So, what you have is, it is a heteropolysaccharide, and that is has, in this particular case, it is made up of 2 monomers.1166

Those 2 monomers are going to alternate, so A, B, A, B, A, B, A, B.1173

You can think of a disaccharide, that A-B, as one unit.1176

You have A-B, A-B, A-B, A-B; that is what we mean by a heteropolysaccharide composed of repeating disaccharide units.1180

That is all it means.1189

OK, now, one of the disaccharide units, one of the disacchs, one of the monomers of the disacch, is either N-acetylglucosamine or N-acetylgalactosamine.1191

I am having a hard time writing today; sorry about that.1241

And, in a minute, we are going to start to use the abbreviations.1250

OK, the other monomer of this disaccharide unit, the other monomer, is most often a uronic acid; and uronic acid, for your quick recollection, if you go back a lesson or two, it is where the no. 6 carbon has been oxidized to a carboxyl group, COO-.1254

And, the two that you will probably see are D-glucuronic acid - and don't worry, we are going to be going over the structures in just a minute, D-glucuronic acid or D-glucuronate for the one that has been deprotonated, which under physiological conditions, it actually shows up as COO-, not COOH, so D-glucuronic acid and interestingly enough, the L-isomer of iduronic acid.1299

More often than not, these glycosaminoglycans, they consist of N-acetylglucosamine as one of the monomers, and some uronic acid as the other monomer; and those 2 units will alternate, and they will keep repeating.1332

Instead of the N-acetylglucosamine, you might have N-acetylgalactosamine.1350

Again, just another hexose, the hydroxy has just changed.1357

OK, one last thing before we start looking at some structures.1360

OK, in some of these Gags, in some of these glycosaminoglycans, one or more of the hydroxys have sulfates attached.1369

In other words, let's say you just have something like, let's say this one, CH2, instead of the OH that has a sulfate attached- that is all.1403

It could be at this one; it could be at this one, this down, up, down.1423

It can be on this one; it can be this one.1431

It can be this one; it could be any two.1432

It could be any three; the different arrangement of the sulfates along this linear polymer actually becomes a site of recognition for different proteins.1434

So, the arrangement of sulfates, the number of sulfates, the density of them has different recognition, it serves recognition, function for proteins that need to bind to them electrostatically.1444

Obviously, if you have a bunch of sulfates, you have a high degree of negative charge, so there is going to be a lot of electrostatic interaction.1455

I just wanted you to know that in some of these, one of more of the hydroxys has a sulfate attached to it- that's it.1460

OK, let's take a look at some of these structures first.1468

Let's do this in black; let's look at the monomers.1473

Let's look at the monomers.1479

Now, we said N-acetylglucosamine, so that is going to look like this, this, that, that, boom.1490

Let's go ahead and do the beta version.1500

We have N; we have C.1503

We have CH3; we have OH, OH, and we have CH2OH.1506

This is N-acetylglucosamine.1513

This is the beta-D-N-acetylglucosamine.1518

Its shorthand is Glc - no, I need my N-GlcNAc.1529

That is N-acetylglucosamine.1539

OK, now, let's do the N-acetylgalactosamine.1541

We have got this, make it a little bit broader here.1546

Let's do the beta form, and again, we have an N.1551

We have a C, and a CH3.1556

This is our N; this is our acetyl group.1560

This one is up, and galactose is a 1, 2, 3, c-4 epimer, so CH2OH.1563

So, here, we have beta-D-N-acetyl…wooh, this is tiring, makes me crazy having to write all these stuff out.1573

This one is GalNAc.1586

That is the shorthand for that one.1590

OK, now, let's do our glucuronic acid and our iduronic acid.1592

Do I want to do them on this page or the next page?1598

You know what, I think I will go ahead and stay on this page.1600

Hopefully, there is enough room here.1604

I have got this; I have got that.1605

Alright, there is that; I will go ahead and do the beta, and this is there.1610

This is there, and this is there; and we said that the no. 6 carbon has been oxidized.1618

So, this one is our - that is what makes it - so, this is beta-configuration D.1625

D- hat is the configuration here and glucuronate.1640

Now, I did the glucuronate instead of the glucuronic acid.1645

The glucuronic acid would just be this protonated- that's it.1649

That is the only difference, so glucuronate.1651

OK, acetic acid, acetate, propanoic acid, propanoate, the A-T-E just tells me that I am deprotonated; and at physiological pH, I am going to be deprotonated.1654

This is beta-D-glucuronate glucuronic acid.1666

Now, let's do the beta-D-iduronic acid, the other particular monomer.1670

Let's see; let's go ahead and go here.1675

Let's see if I can do this one.1679

Alright, let's do this as a beta-configuration.1682

Now, this one is going to be OH.1686

This is a little different, and this is going to be OH; and here, we are going to have the COO-.1690

Here, this is the L; remember, we said it is L.1700

This is beta-L-iduronic, iduronate or iduronic acid.1704

The L-configuration, remember, if we said any 2 substituents, we change configurations.1714

The D in the L is based on the chiral carbon that is farthest from the carbonyl.1723

The carbonyl carbon is this one; that is the anomeric carbon.1732

It is the no. 5 carbon - 1, 2, 3, 4, - that decides D or L.1735

D the CH2OH, which is now carboxyl, is above the ring.1740

The L-configuration just switched the H and the CH2OH.1745

Now, the CH2OH, or which is now the COO-, that is below the ring.1749

OK, this is L; this is D.1754

A galactosamine, in general, has one of these and one of these alternating.1757

Let's say we have N-acetylglucosamine and we have beta-D-glucuronic acid, A, B, A, B, A, B, that is going to be a particular glycosaminoglycan.1763

OK, let's see what we have got.1777

Now, and let me just write down one thing regarding the sulfate attachments.1783

Yes, that is fine; I will go ahead and write it down.1790

OK, regarding the sulfate attachments, I am just going to reiterate what it is that I said before.1793

The pattern of attachment provides for recognition by protein molecules which can bind electrostatically.1807

Now, protein molecules can also bind covalently, but in this particular case, it tend to bind electrostatically.1849

Oligosaccharides, polysaccharides, sugars, carbohydrates, on the cell surface, are how cells recognize each other.1858

The whole idea of recognition is all based on the arrangement of sugars on the cell surface- a particular configuration, a particular arrangement, 15 monomers, 27 monomers.1866

That is how individual body, individual cells recognize each other and communicate with what is happening inside the cell.1877

Glycobiology- profoundly important, and it is a fantastic, fantastic area of research that is only just beginning.1885

It is really only just beginning.1892

There is so much work to be done and so many wonderful new things to be discovered in this absolutely amazing, amazing field of biochemistry.1894

OK, let's take a look at some of these glycosaminoglycans.1903

Let's go ahead and go to blue; there we go.1907

Let's take a look at some glycosaminoglycans, some Gags.1913

OK, the first one we are going to look at is hyaluronic acid.1920

This is hyaluronic acid or hyaluronate, and this particular Gag is made up alternating monomers of GlcA and GlcNAc.1925

Oh, you know what, I think in the last page, when I did the glucuronic acid and the L-iduronic acid, I forgot to put the symbols.1957

So, this GlcA, that stands for the glucuronic acid.1969

That is the shorthand for glucuronic, and then, of course, we have the IdoA.1977

That is the iduronic acid; sorry about that.1985

OK, in this particular case, this particular glycosaminoglycan called hyaluronic acid, it has alternating monomers of glucuronic acid and N-acetylglucosamine- A, B, A, B, A, B.1992

OK, and here is the pattern of the binding for the glycosidic bond.2005

I am going to do this with 3 monomers.2010

Let me go ahead and do this in black.2015

This is going to be GlcA.2020

It is going to be beta-(1,3) - very unusual, very unusual - beta-(1,3) GlcNAc.2025

This one is going to be beta-(1,4); this is a little bit more normal.2033

And then, we have GlcA again, and then it goes on like that; and, it is going to be somewhere in the neighborhood of about 50,000 monomers.2037

It is a pretty long molecule.2049

Left to right, the glucuronic acid connected to the N-acetylglucosamine is connected by a beta-(1,3) glycosidic bond; and the N-acetylglucosamine connected to the next glucuronic acid is connected by a beta-(1,4) bond.2053

So, we have everything that we need right here in order to draw out the structure.2070

Now, let's go ahead and draw out the structure.2074

OK, let me start with a GlcA on this side.2076

Again, glucuronic acid, let me do this in black.2082

I have got this, that, that.2087

Now, let me go ahead and draw those two.2093

This is going to be the glucuronic acid.2102

This is going to be OH; this is going to be OH.2106

This is going to be OH, and this is going to be the COO-.2110

That is the carboxylate.2114

Now, we said that it is connected in a beta-(1,3).2116

Well, this is the anomeric carbon.2119

Let me number these; this is the no. 1 carbon.2124

This is no. 2; this is no. 3.2126

Over here, what we have is...let me go back to black.2133

This is going to be the N-acetylglucosamine, so this is NH.2137

I am thinking in the last structure, I think I forgot the H on the nitrogen; sorry about that.2141

OK, this is going to be C.2145

This is going to be CH3; this is down.2149

Now, here, on the no. 3 carbon, let me actually say the numbers until afterward; I think it is a little bit better.2153

So, here, this is going to be O, like that; and, of course, here, we have the OH, and we have CH2OH.2160

Let me draw a couple of more of these, and then I will go ahead and discuss these particular glycosidic bonds on this molecule.2179

Let me see; let me go ahead and do this.2189

Let me go ahead and do that.2191

I will just make it a little bit quicker here.2194

COO- and I’ve got an OH on top; I’ve got OH on the bottom.2197

This is glucuronic acid, OK.2203

I have got this one here, and I’ve got this.2205

It is going to be O there; this is going to be a CH2OH.2214

This is going to be glucose, and this is going to be, this one is up, and this one is down; and let’s go ahead and leave it as beta.2226

Let’s just write 4 of those units right there.2237

OK, let’s take a look at what we have got here.2239

This is our GlcA, and this is our GlcNAc.2243

This is our GlcA, and this is our GlcNAc- glucuronic acid, N-acetylglucosamine, glucuronic acid, N-acetylglucosamine.2252

A-B, A-B- just keeps going in this direction and this direction; and the GlcA to the N-acetylglucosamine is beta-(1,3).2263

Well, here is our beta-1, and here is our no. 3.2271

Notice, beta-oxygen above 3 glucose, the no. 3 carbon, the oxygen is above the ring.2275

The hydroxy is above the ring, that is why we drew it this way; it is very, very unusual.2283

Now, N-acetylglucosamine connected to GlcA, with N-acetylglucosamine on the left, GlcA on the right, is connected with a beta-(1,4).2288

Well, here is our beta-1; this is our beta-1 carbon, and here is our no. 4 carbon.2299

And then again, GlcA to N-acetylglucosamine, it is going to be beta-(1,3).2304

This is our beta-1; this is our no. 3 carbon- that is it.2312

If you have this and if you know what the individual monomers look like, that is there, this is there, and that is there, and this is...oops N-acetylglucosamine, that is not right.2319

This is NH; sorry about that, COOCH3.2334

Yes, I know; a whole bunch of carbons, oxygens, and things floating around.2341

It is very easy to lose your way as you can see.2346

OK, that is it.2348

Once we have this arrangement, once we know what is connected to what, we can draw up out structure.2350

If we are given the structure, we should be able to go backward; we should be able to recognize this is a uronic acid.2356

This is N-acetylglucosamine; this is a beta-1 configuration no. 3.2362

We should be able to write this out, you have to be able to go both ways.2366

OK, now, let’s talk about these hyaluronates or these hyaluronic acids.2370

Not only do they form part of the extracellular matrix, they actually form the lubricants for your joints, lubricants in your joints, and they also happen to give your eye that jelly-like consistency.2386

So, I can do that because of these hyaluronic acids.2407

Now, I am just going to go ahead and list some other glycosaminoglycans, and I am going to list them; and again, I am going to encourage you to use your book because it is your primary resource.2410

It is a fantastic resource with wonderful pictures and further discussion of what these individual glycosaminoglycans happen to do.2421

And again, we are just learning what these things do very, very recently.2431

Glycobiology, it is a brand; it is your definitely ground level if you want to get into glycobiological research- fantastic area.2438

Let me do this in blue here.2445

Other glycosaminoglycans, and I encourage you to take a look, maybe do a little bit of look on the web, look on your book- whatever it is that you need to do.2449

I am not going to talk about them again.2461

The only difference is you have different monomers, but it is always going to be a disaccharide unit.2463

It is always going to be an alternating A-B, A-B, A-B.2468

You are not going to have a C, a D, an F.2471

It is going to be, it is hetero, but there is only 2 monomers that make up this linear chain; and there is not going to be any branching.2473

Glycosaminoglycans, at least, not that we have discovered yet; I could be wrong.2482

That is the wonderful thing about biochemistry- you will never know what is going to happen tomorrow.2488

OK, some important ones, some of these you have actually heard off.2493

Chondroitin-4-sulfate, this chondroitin-4-sulfate, this just means that the hydroxy on the no. 4 carbon...so, if I just take some random 1, 2, 3, 4, either there or there.2498

I will not specify the stereochemistry.2516

OK, this could be glucose; it could be galactose, so I will just put OH here.2518

OK, it could be any stereochemistry, above or below.2521

This has been sulfated; that is all this means- 4-sulfate, chondroitin-4-sulfate.2525

Maybe it is 3 sulfate; maybe it is 6 sulfate that tells me the carbon that has been sulfated, the carbon that has the oxygen attached, that has the sulfate attached to it.2536

OK, chondroitin-4-sulfate generally tends to be in the range of about 20 to 60 monomers- very, very short.2548

These are a lot shorter than the hyaluronic acid; hyaluronic- they are huge.2555

These tend to be very, very short.2560

Keratin sulfate, somewhere in the range of maybe 25 monomers.2564

Heparin- definitely a molecule that you want to get to know well.2574

And, those of you that are going to be going on into medicine, you will get to know it very, very well.2577

Heparin, somewhere in the range of 20 to 90- a lot of variation.2581

OK, clearly these are much shorter.2588

You know what, I don’t need to write that; I mean, clearly, you know that these are much shorter, obviously.2600

We said that 50,000 versus let’s say 20 unit; yes, it is a lot shorter.2604

OK, these tend to be covalently linked to proteins.2610

So, if you run across a keratin sulfate, heparin, chondroitin-4-sulfate, any number of things, these will tend to be covalently linked to some type of a protein.2631

OK, that finishes our discussion of polysaccharides- almost, actually.2644

We have a little bit more to discuss, but that certainly finishes today’s lesson.2648

Thank you for joining us here at Educator.com.2652

We will see you next time, bye-bye.2653

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

In the last lesson, we talked about polysaccharides.0004

We are going to continue our discussion of polysaccharides, and talk about something called glycoconjugates.0007

These are polysaccharides that are attached to proteins and lipids.0013

Let's just go ahead and jump in, get started.0022

OK, let's make sure we have...let's start off with black here.0026

In addition to their structural and fuel storage roles - actually, I do not think I will use the word "roles", I think I will use the word "capacities" - oligo and polysacchs - excuse me - they also carry information.0031

OK, let's go over here; they also carry information.0087

In other words, in this particular case, they are involved - as we mentioned in the previous lesson - in cell to cell and cell extracellular matrix interactions.0093

In other words, they are involved in recognition.0122

OK, in these cases, the carbohydrate, oligopolysaccharide, the carbohydrate is often joined to a protein or a lipid.0125

Lipid is just a fancy word for fat, and we are going to be discussing lipids in great detail very, very soon.0160

OK, let's talk about these things called proteoglycans.0167

Let me go to blue here, and let me see.0171

Is proteoglycan like a protein?0175

Yes, that is fine; alright, proteoglycan.0177

Our first class of glycoconjugates are these things called proteoglycans.0185

OK, they are cell surface and extracellular.0192

Extracellular means that they are just proteins that are not attached to the cell; they are actually proteins that are floating around in the cytosol.0202

That is all that means- proteins with one or more glycosaminoglycans, one or more Gags attached.0207

OK, now, these things called proteoglycans, they are the carbohydrates, but they are attached to cell surface proteins or they are attached to free proteins, secreted proteins, proteins that have been created and sent outside of the cell to sort of wander around in the matrix or wherever it is that they wander around, proteins with one or more glycosaminoglycans attached.0227

What is important here is the attachment to the protein is an actual glycosaminoglycan, and you remember from the last lesson that the glycosaminoglycan is this particular polysaccharide that has alternating, it is a linear polymer.0254

It is not branched; it is linear, but it alternates A-B, A-B, A-B with a certain collection of monomers, those monomers generally being N-acetylglucosamine or N-acetylgalactosamine and the other monomer being, more often than not, a glucuronic acid or an iduronic acid.0269

Now, other things can show up; it is not a problem, but the majority of the glycosaminoglycans are going to be those.0289

So, when that particular arrangement of polysaccharide is attached to a protein, we call that glycoconjugate; the whole thing, it is called a proteoglycan- that is all this is.0296

The carbohydrate in a proteoglycan tends to dominate the structure, and that is where the biological activity is.0306

The protein just happens to be a point of attachment.0316

OK, another family of glycoconjugate is something called a glycoprotein.0320

I know, it is kind of interesting, isn't it?0328

Glycoprotein, notice, here, proteo is first, glycan is second; here, glyco is first, protein is second.0330

Now, these are proteins with one or more oligosaccharides covalently attached, and they are also covalently attached here.0337

They can be covalently or electrostatically, but here, they are covalently attached.0364

Now, the carbohydrate portion is generally more varied in the sense that there is a greater collection of monomers to choose from, that the oligosaccharide is made of, is more varied and complex than the glycosaminoglycans on the proteoglycan, the glycosaminoglycan chains on the proteoglycans.0370

If you see a protein, and it has some carbohydrate attached, if the carbohydrate attached happens to be a glycosaminoglycan, we are talking about a proteoglycan- the whole thing.0422

If it tends to have some oligosaccharide or polysaccharide attached, but it is a lot more complex and it has branching, that is a glycoprotein.0430

That is the difference between the two; I will actually write that out in just a second.0440

Let me go ahead and just run through the list here of glycoconjugates.0445

We have proteoglycans, glycoproteins, and now, we have something called a glycolipid.0450

Now, this is just lipids in the cell membrane that have oligosaccharides attached- that is it.0461

In this particular case, the carbohydrate, the oligosaccharide, the polysaccharide, is attached to a fat, not a protein - that is it - covalently linked.0486

OK, now, proteoglycan versus glycoprotein.0496

I know this is good.0505

Now, what is the difference, proteoglycan, glycoprotein?0508

Here is the difference.0510

Proteoglycan, it has a linear glycosaminoglycan attached- that is the difference.0512

The attachment to the protein is some linear glycosaminoglycan, and we know that a glycosaminoglycan is a heteropolysaccharide.0524

It is linear, has no branching; and it consists of alternating monomers, A-B, A-B, A-B- whatever those monomers happen to be.0533

OK, linear Gag attached, and, of course, the Gags are repeating disacch units - that is how you tell - whereas a glycoprotein, it has various oligos attached, different monomers, linear, branched, all kinds of crazy things, any arrangement, high binding specificity.0542

Now, we said that these oligosaccharides attached to proteins that are on the cell surface are floating around, they serve recognition purposes.0594

Well, yes, proteoglycans are involved in recognition; glycoproteins are involved in recognition.0606

Proteoglycans are involved in sort of a global recognition - attached here, attached there.0612

Because these tend to be smaller, more complex, they have a higher binding specificity.0620

They bind specific things at specific points, whereas a proteoglycan might have some glycosaminoglycan attached; and maybe, it is sort of attached at 15 or 16 different points, just sort of like, it is attached here, attached here, attached here, sort of like a claw, whereas these glycoproteins are attached in one location.0626

So, these oligosaccharides tend to be much smaller, and they have very high binding specificity- that is the thing.0648

These glycoproteins are the ones that are involved in your body's immune response.0656

The immune cells that your body sends out to attack invaders recognize these glycoproteins on the cell surface of the bacteria and viruses and things like that.0664

OK, let's take a look at a drawing here, and OK.0675

This is a cell surface; a cell is a lipid bilayer, so that is this thing right here in blue.0681

OK, we are not going to worry about that; let's just sort of take a look and see some of these things.0690

Now, notice this particular protein here; this particular protein looks like it has a couple of things attached, so it looks linear.0697

This could be a proteoglycan; I will just say PG.0708

This could be a proteoglycan.0711

This over here, this is a membrane bound protein; part of it is in the membrane.0714

Part of it is inside the cell; part of it is outside the cell.0720

This is inside the cell here; this is outside the cell, and it has this oligosaccharide attached.0723

Notice, this is not a linear oligosaccharide; this is, it branches here.0730

It branches there, and it branches there.0735

This one is a glycoprotein- that is it.0737

That is how you tell; I mean unless you do a particular analysis, there is no way to actually tell, but this pretty much gives it away.0741

If you have some extensive branching, you are pretty much looking at something which is a glycoprotein.0749

It is going to be a very specific binding site.0754

Let's see, over here, there is another in an integral protein.0757

Integral means it is part of the cell membrane.0761

There is a part of the protein that extends inside; there is a part that extends outside, and again, we have a linear.0765

Now, this could be a glycoprotein; it just depends on what the identity of this oligosaccharide is.0770

If it happens to be a glycosaminoglycan, let's say something like chondroitin-4-sulfate, we know that this whole thing, the protein and the oligosaccharide, is a proteoglycan.0775

If it happens to be just some collection of monomers - N-acetylglucosamine, N-acetylgalactosamine, maybe a mannose, maybe a galactose, something like that - then we know we are looking at a glycoprotein.0786

It serves a more specific type of recognition site- that is all it is, and this is what it is going to look like.0800

You see, all of these proteins, they have little oligosaccharides.0807

Your cells are covered in this stuff; bacteria is covered in this stuff.0811

That is how things get recognized; in this particular case, glycoprotein, proteoglycan, I just wanted you to see what this looks like.0815

Now, over here, we have this glycolipid.0825

Here, we have a lipid molecule, some fat; and again, we will be talking about lipids specifically, but here, you have an oligosaccharide that is attached to a fat.0829

It is not attached to a protein, not a proteoglycan, not a glycoprotein, but it is in the third class; it is a glycolipid- that is it.0838

It just means it has a sugar that is attached to some fat- that is it.0846

We will return to this a little bit later in the lesson when we get into a little bit more detail about proteoglycans and glycoproteins.0851

OK, let's talk about proteoglycans first, and let's talk about their common structure.0857

Proteoglycans are alright, proteoglycan common structure, and by common structure in this particular case, what I am going to discuss is the point of attachment to a protein.0869

In this case, we said that this was our proteoglycan, so we are going to be talking about - you know, I should do this in black, this right there - how it is actually attached to the protein.0894

Well, here is how it is attached to the protein; let's go ahead, for our example.0905

I wonder if I should do this on the next page.0912

Yes, that is fine; I should have enough room here.0916

Let's go ahead and actually do a chondroitin-4-sulfate.0917

OK, chondroitin-4-sulfate, it is a glycosaminoglycan; it is linear, and its particular monomers are glucuronic acid and N-acetylgalactosamine, and I am going to write that down in just a second.0930

So, I am not going to draw these structures; I am just going to write their names connected.0949

I have got a GlcA; I have got it connected to a GalNAc- that is the N-acetylgalactosamine - and I am going to write one more GlcA, right?0953

So, we have alternating A-B, A-B, A-B, and this is going to be connected to a galactose, which is going to be connected to a galactose, which is going to be connected to a xylose sugar.0967

Remember, xylose is a 5-carbon sugar; galactose is a 6-carbon sugar, and this happens to be connected to a serine residue, which is part of the polypeptide, which makes up the protein.0980

And, that is the term Gly, Hex, Gly; and, of course, the polypeptide continues on in this direction.0999

This is going to be the N-terminus.1008

Now, when it is attached to a serine residue, what tends to be attached to the serine is another glycine, some other amino acid and some other glycine.1011

In general, this is what we tend to find more often at this point of attachment, and, of course, this polymer goes on that way, the polypeptide, and this is the C-terminus.1023

OK, this tends to be the arrangement.1033

Right there, at the protein's surface, there is some serine, glycine, some other amino acid and glycine, this arrangement, and then attached to the serine, you will have a xylose sugar, a galactose, a galactose, and then, of course, you will have your molecule.1035

Here is your molecule of your chondroitin-4-sulfate.1049

This is our chondroitin-4-sulfate, and this is our trisaccharide linker that links the protein, the amino acid serine, to our glycosaminoglycan, which is the chondroitin-4-sulfate.1058

This is our trisaccharide connector.1080

OK, let's see; all these crazy words floating around.1087

OK, and let's do this one in red.1097

This right here, this glycosidic bond is going to be beta-(1,3).1102

The anomeric carbon of the glucuronic acid is connected to galactose at its no. 3 hydroxy on the no. 3 carbon.1107

This connection right here is a beta-(1,3) glycosidic bond.1118

Now, let's go ahead and draw out serine just so you see what serine looks like as a reminder.1124

We have NCC; serine has a CH2, and it has that, and, of course, this is H.1132

This is that, and this is going to go on that way.1143

This is going to go on that way; that is serine, just as a reminder of what serine actually looks like, so that is it.1145

This is the point of attachment for a proteoglycan.1153

This is the part that is different.1160

There are going to be different glycosaminoglycans attached through a trisaccharide connector to a serine residue on a proteoglycan.1163

That is what's happening right there.1172

OK, now, let's talk about glycoproteins.1176

You know what, I think I am going to go back to blue; for some reason, I just thought I really, really like blue, and I don't know why, but there it is.1186

Now, again, the oligos are smaller and more complex, smaller and more diverse and complex than the glycosaminoglycans.1200

Now, smaller, you might think "Well, wait a minute, smaller and more complex, that doesn't make sense".1224

It does, it is referring to the branching.1229

Yes, they are smaller; you have fewer of them, but there is more complexity and there is more variation because now, you are not talking about just 2 monomers alternating A-B, A-B, A-B.1232

You are talking about maybe 5 or 6 to choose from, in general, on most of these glycoproteins; and, of course, they can have all kinds of different branching on them.1242

That is what we mean by more complex.1251

Complexity is a measure of quality; length is a measure of quantity.1254

They are smaller but they are more complex; the quality of them is different.1258

Now, OK, the first monosacch is attached to the protein by its anomeric carbon.1263

The no. 1 carbon, well, I won't say the no. 1 carbon because for ketoses, it could be the no. 2 carbon, so we will just call it the anomeric carbon, the one that originally had the carbonyl.1290

Now, the sugar is in the ring structure; now, there is a hydroxy attached to it, so that is the anomeric carbon- the one that was originally the carbonyl by its anomeric carbon.1302

OK, and now, it is attached to its anomeric carbon in 2 ways: through the hydroxy - I will draw it this way - through the hydroxy group on either serine of threonine.1314

We call this O-linked; in other words, it is an O-glycosidic bond - no worries, we will be drawing it out in just a minute - or, it could be attached through the amide nitrogen which is on asparagine, on Asn.1341

This is called N-linked because this is going to be an N-glycosidic bond.1387

So, it is either an O-linked glycoprotein or it is an N-linked glycoprotein.1393

In other words, the oligosaccharide is either attached to a serine threonine residue that is an O-linked glycoprotein, or it is attached to an asparagine residue that is going to be an N-linked.1398

So, this is an N-glycosidic bond.1409

OK, let's go ahead and draw what these things look like; let's go ahead and do an O-linked first.1417

Let me go ahead and do this in black; I think I will do it.1423

Let's use N-acetylglucosamine as our monomer that is attached to a serine residue.1429

Let's go ahead and draw our sugar unit first.1436

That is there, and let's go ahead and make it an alpha; and let's go ahead and make this, yes, NH, COO, CH3.1439

This is OH; this is OH, and this is CH2OH.1459

And now, I have got my CH2; actually, you know what, I am going to do this in 2 colors here.1466

I really want you to see this in 2 colors; I am going to do this second one in, you know what, I will do it in red.1472

O, CH2, C, this is NH; this is COO, and, of course, the polymer, the protein goes that way.1480

The protein goes that way; this right here is our serine residue.1493

OK, now, the O that is connected to the sugar that comes from the serine that is not from the hydroxy on the original sugar- that is the whole idea.1498

This is the nucleophile; it is going to get rid of that hydroxy, so it definitely comes from this serine.1509

That is important to know; that is why I did it in 2 colors.1513

I hope that is not confusing; let me see, let me write out what this is over here.1517

Let me go back to black; this is the alpha-GlcNAc.1524

Alpha, that is that; the anomeric carbon, this is alpha-1, N-acetylglucosamine, and it is attached to a serine residue.1533

This is O-linked.1542

OK, now, let's do an N-linked structure, so you see what that looks like; and this time, I am going to use the beta-N-acetylglucosamine.1546

Let me go ahead and go back to black here; let me draw my sugar that way.1556

OK, you know what, I am going to draw it a little bit lower here.1565

I need a little bit more room; excuse me.1569

I will go ahead and draw it like that, so that is that; and we said beta.1574

Let me go ahead and put in this first; this is NH.1581

I am always forgetting that H; I don't know why.1584

Well, old habits- they die hard.1587

OH, OH, and we said N-acetylglucosamine, so this is CH2OH.1591

This is a beta-GlcNAc, right?1600

Yes, and we said, now, we will go ahead and go; we have N.1607

We have COO, CH2, C, NH, COO.1614

The protein, the polypeptide is this thing right here.1628

This is the asparagine residue; this is the R-group.1632

Well, the whole thing is the asparagine residue; this is the R-group on the asparagine.1635

We have got CO, and there is also, let's go ahead and put an H on here too because there is an H there.1640

this right here is our Asn residue, and this happens to be our beta-1-carbon, and the nitrogen, this is an N-linked, right?1646

So, we have, this is N-linked, and the nitrogen comes from the protein, from the amino acid.1662

OK, so it is N-linked; this is a nucleophile.1674

This is what is nucleophilic; it is what is going to displace the hydroxy.1677

OK, there you go.1681

Now, let's talk about the monomers that commonly show up in these glycoproteins, in these oligosaccharides, monomers that commonly show up in the oligo portion of glycoproteins.1683

I wonder if I am going to have enough room here to write out all of them; yes, it is fine.1725

OK, let's do this in black.1729

Now, it is not exclusively these; these just tend to show up more often than any others.1736

GlcNAc- that is N-acetylglucosamine.1741

GlcNAc- this is N-acetylglucosamine, and we have Man.1750

This is mannose; it is a hexose.1762

We have Gal; that is galactose.1766

It is another hexose.1772

We have Neu5Ac; this is called N-acetylneuraminic acid, otherwise known as sialic acid.1774

It is actually A-sialic acid; it is a class of molecule, but we tend to call it sialic acid.1788

And, no worries, I will be drawing out the structure in just a minute, or you can look in your book- either way.1795

And, we have Fuc, which is fucose; it is just another sugar, and this one usually shows up as the L-isomer, L-fucose, instead of the D-isomer.1805

And, our last one - which I should have left room but that is OK, I will go ahead and write it over here - is GalNAc.1823

This is N-acetylgalactosamine; this is N-acetylgalactose.1832

Wooh, I mean this is exhausting.1842

You can see why biochemistry is drilled with acronyms - Gag, Gal, Man, PG, GP - all over the place, simply because we can't write everything out.1845

OK, let's go ahead and draw out a couple of the structures.1858

We have talked about most of these, but I am going to go ahead and draw out the sialic acid, the N-acetylneuraminic; and I am going to go ahead and draw out the L-fucose just so you see what the structures look like, just for the heck of it.1865

OK, let's go ahead and do this in red.1878

Actually, no, let's go ahead and do this in blue.1882

This is going to be, that is that, here, here, here.1885

Now, let me see, we have got COO-, OH.1896

This one is deoxy; we have the hydroxy here, and here, we have NCOCH3, and we have an R-group, and R happens to be equivalent to C.1904

This is OH; this is O - yes, that is right, I always forget that - CH2OH.1923

This R-group right here, that is just this thing, just not enough room; we just do that.1929

This is our N-acetylneuraminic acid.1933

This is actually the deprotonated, so it is N-acetylneuraminate.1944

This is our Neu5Ac.1951

OK, now, let's go ahead and do our other one.1957

OK, I am going to leave the stereochemistry on this one unspecified.1964

I am going to go OH; I am going to go OH, and I am going to go OH, and there is going to be a CH3 on this one, and this is L-fucose.1970

L that is the carbon, 1, 2, 3, 4; that carbon is what specifies the L.1983

OK, let me see what it is I have got here- example of an N-linked oligosaccharide.1988

Let me just give you a quick example of an N-linked oligosaccharide.1997

I will do this in red just to show you what it looks like, and I am not going to actually draw out the structure; I am just going to draw out little hexagons and put numbers in them.2004

We have Asn; that is going to be our Asn residue, and, of course, the peptide goes in that direction.2024

It goes in that direction, and it is going to have attached, let's say, I will just draw them out as hexagons.2033

Now, of course, these are sugar rings, so there is definitely an oxygen in the ring somewhere; but I do not know where that oxygen is going to be depending on what the connection is.2040

So, I am just going to draw them as little individual hexagons.2049

You will often see it like this.2053

Let's see, boom, boom, boom, boom; and let's go ahead and go here, boom, boom, boom, boom, boom, boom, and up here like this.2058

These are all in original units, so this is no. 1, 1.2071

Let's go ahead and do something like that and maybe something like that.2074

Maybe I will just go ahead and put one more for the hell of it.2081

1, 1, 1, 2, 3, 4, 2, 3, so again, 1, 2, 3, these 1, 2, 3, 4s, they are different monomers.2084

This can be N-acetylgalactosamine; this can be fucose.2096

This can be N-acetylglucosamine; this can be mannose.2099

It could be anything- that is it.2102

This is just sort of what it looks like, and again, these are all sugar hexoses, but we have not put the oxygen in there because we do not know exactly where the oxygen is in terms of connection.2105

Now, what is interesting about this, the degree of complexity that we talked about earlier has to also do what the glycosidic linkages.2117

In order to fully specify what this oligosaccharide arrangement is, I have to give the connection, that one, that one, that one, I have to specify each connection; and what is interesting about these glycoproteins, the oligosaccharide portion of these proteins, is you can have 1,4 glycosidic bond.2125

You can have 1,2 glycosidic bond, 1,3; you can have 1,6.2146

You can have 2,3- any combination.2152

As long as there is a hydroxy available to react with something else, you can have those connections.2155

That is why in order to fully specify, you have to specify the connection at each glycosidic bond.2160

We are just generally going to talk about it like this.2167

If we happen to need a specific glycoprotein that has this connection to this connection, we will deal with it like that, but this is what we mean by complexity- certainly a hell of lot more complex than a proteoglycan, which is just alternating monomers.2169

OK, let's take a look at the picture one more time- the cell membrane picture.2186

Here we go, so, again, let's say this one is a glycoprotein.2195

Let's say there is another glycoprotein; let's say this is a glycoprotein.2205

This one is a glycoprotein; this looks like a proteoglycan.2213

This one here, probably a proteoglycan attached.2217

Actually, you know what, that one looks like a glycolipid.2221

This one right here, a glycolipid- that is it.2224

You are just sort of identifying and taking a look at what the carbohydrate looks like and what it is attached to.2227

OK, now, let's go ahead and see; let's talk about glycolipids a little bit.2236

Glycolipids are lipid molecules, fat molecules, in the cell membrane.2247

I mean all of these are fat molecules.2267

These are fat molecules that are not necessarily, that make up, the things that make up the lipid bilayer of a cell membrane, it could be attached to that, but more often than not, it is attached to some lipid that is in there like a cholesterol or some other lipid that happens to be in that lipid bilayer, and the oligosaccharide is attached to that.2276

OK, are lipid molecules in the cell membrane with oligosacchs attached, and, of course, we said that already, but there is no harm in repeating ourselves.2302

OK, now, we are going to, of course, be discussing lipids, and glycolipids in much more detail when we specifically get to that chapter discussing the lipids and all the different types of lipids and certain oligosaccharides that are attached to those things.2318

We are definitely going to get more into detail about this, but just to get an idea because we are talking about glycoconjugates, and a glycolipid happens to be a glycoconjugate.2338

OK, now, I am going to talk about one particular type of glycolipid.2348

It is called a lipopolysaccharide.2353

And again, this is mostly just for your edification at this point; we will be discussing it in more detail later- lipopolysaccharide.2359

OK, a lipopolysaccharide, now the dominant feature - and when we say dominant, we definitely mean dominant - the dominant feature on the outer membrane of gram-negative bacteria for example E. coli is a gram-negative bacteria and certain salmonellas or salmonella.2370

OK, the lipopolysaccharide is the dominant feature on the outer membrane of gram-negative bacteria.2415

I mean, it is just covered with this stuff.2419

OK, it is a complex oligosaccharide, complex oligosacch units covalently attached to multiple lipids in the outer cell membrane.2423

In this particular case, a lipopolysaccharide is a specific example of a glycolipid, and it is peculiar to gram-negative bacteria; and it is where this particular oligosaccharide are, these things are attached to not just one lipid molecule in the membrane, but several different lipid molecules that are anchored in the membrane.2463

And now, we want to go ahead and take a look at one of these and what it looks like.2488

OK, this is what it looks like; let's take a look.2494

Here, we are talking about the cell interior; this bottom portion right here, this is inside the cell.2498

Inside the cell membrane, you have these lipids, OK, these long carbon chains, these are fats, these lipids that are inside the cell membrane.2507

Now, notice, these are attached to sugar units, and, of course, on the no. 6 carbon, it looks like one of these, that through and O-glycosidic bond, is attached to several other sugars.2520

So, this is going to be outside the cell; this part is the core oligosaccharide.2540

This part is always the same; this particular arrangement Kdd, Kdd, Kdo - I'm sorry, yes - Kdo, Hep, Hep, Glc, Gal, Gal, Glc, Ngc, this particular arrangement of oligosaccharide, this oligosaccharide right here is always going to be the same.2549

Do not worry, these are just different monomers: Kdo, Kdd, Hep.2566

These are just different types of sugars with different things attached to them.2569

What is different from bacteria to bacteria or different places along the bacteria, is this thing right here.2574

This is the thing that changes; this is the same.2582

This is the same, but this is the thing that changes; and depending on what this is, what collection of monomers and what glycosidic bonds are actually connecting them, that is going to be the point of recognition.2586

We call this the O-antigen; this is what your immune cells recognize when they run across the bacteria in your body.2599

That is what they attach to in order to do what the immune cells do, which is destroy these things or whatever else they plan on doing to it- that is it.2608

This is just an example of a lipopolysaccharide; it is a glycolipid.2618

It has a lipid core that is in the membrane.2623

In this particular case, certain portion of it contains an oligosaccharide, which is in variant; and then, of course, at the end, it is variant.2629

Different things happen up here; this is going to be the same.2642

This whole thing is the oligosaccharide portion plus these two, and this is the lipid portion- that is it.2644

We just wanted you to get an idea of what something like this can actually look like.2653

OK, thank you for joining us here at Educator.com2659

We will see you next time, bye-bye.2663

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today we are going to round out our discussion of carbohydrates by just doing some more example problems with them.0004

If you feel comfortable with it, it is not a problem; you can move on to the next lesson, but I thought it would be nice to sort of recap, and maybe towards the end do something a little bit more complex.0010

Let’s get started.0019

Pretty basic stuff, again, we are mostly concerned with structure.0022

The idea behind carbohydrates is you want to be able to recognize the monomers, recognize the connections of the glycosidic bonds and be able to sort of put, not sort of, be able to put them together; or given a structure, be able to say that "this is this connection, this is this connection".0025

That is really the main purpose; we want you to have a good structural understanding of what is happening with the carbohydrates.0043

The rest of the information is information, the extent to which the level of detail that you need regarding the function of a particular family of carbohydrates over a specific carbohydrate, that is going to depend on what your teacher wants, how much he or she wants you to know.0049

It is structure that we are ultimately concerned with.0066

OK, example no. 1.0069

Let’s go to blue; I like blue.0073

So, example no. 1: draw the linear form of D-ribose.0080

OK, linear form; we are not putting it in a ring.0093

Ribose is going to be a 5 carbon sugar, and it isn't aldose.0098

It has an aldehyde on the no. 1 carbon, not the ketose, which would be a ketone on the no. 2 carbon.0103

Let’s just go ahead and draw our carbon chain; this is 1, 2, 3, 4, 5.0109

Let’s go ahead and put our aldehyde group up there.0116

Let’s go ahead and put the H2OH, the non-chiral carbon and as far as ribose is concerned, ribose happens to have all hydroxys on the right- that is it.0119

This is D-ribose, and notice there is no alpha-beta here because it is not in a ring formation.0130

That alpha-beta only refers to when a particular sugar is in a ring formation and whether the hydroxy is pointing below the ring, alpha or above the ring, beta when you are looking at it in a standard projection with the oxygen in the back right or in the case of a pentose ring, with the oxygen straight back- that is it, nice and simple.0135

OK, example no. 2.0155

Oops, go ahead and do this, example 2.0160

Xylose is a c2-epimer or epimer depending on how you want to pronounce it.0167

It is completely up to you; pronunciation is completely irrelevant unless people really just absolutely don not understand what you are saying- epimer of D-ribose.0175

Oh, and again, the D part, this D versus L, that comes from the chiral carbon that is farthest from the aldehyde; that is this one.0189

D, the hydroxy is on the right in linear form like this, in Fischer projection.0200

If it were an L-ribose, the hydroxy would be on the left, so this specifies the D.0205

OK, xylose is a c2-epimer of D-ribose.0212

Draw its linear structure; draw its linear structure.0216

Well, nice and easy.0222

We know that an epimer just means that at that particular carbon, in this case, the no. 2 carbon, the configuration is switched.0225

So, when we want to switch a configuration, we switch to the substituents.0232

In other words, we just move the OH from one side to the other.0236

I mean obviously over here we have hydrogens; we have not put them in here for the...but they are there.0240

In general, I will leave the hydrogens off.0244

OK, let’s go ahead and draw the structure again, 5 carbons, 1, 2, 3, 4 and 5; and we will go ahead and put our aldehyde.0249

Oops, excuse me; let’s make this a little bit better.0260

That is that; that is the hydrogen.0263

That is our aldehyde group; let’s go ahead and put our H2OH, the non-chiral carbon, so c2-epimer.0265

This is 1; this is 2.0271

This is 3; this is 4.0273

This is 5, so 1, 2...wait, I am sorry.0275

Xylose is a c3-epimer, my apologies.0282

I was going to say something is going on here, so 1, 2.0287

The 2 hydroxy stays the same; that is on the right.0289

c3-epimer, this is the one that goes on the left, and this is that, so there you go.0294

That is xylose, the c3-epimer of D-ribose.0300

No. 3 carbon just switched the configuration- that is all.0305

OK, let’s see; let’s draw some rings here.0308

Example no.3: draw the furanose and pyranose forms of D-xylose.0316

We just had the linear form of D-xylose.0338

Now, what we want to do is they want us to draw the ring configuration, but they are not specifying, well, they are telling me that they want both ring configurations.0340

They want the 5-membered ring, the furanose; let me go ahead and do this in red.0349

The furanose is the 5-membered ring, and pyranose is the 6-membered ring.0353

We are going to take a look at the structure to see which hydroxys we are going to attach to the aldehyde carbon to create the 5 and 6-membered ring.0358

And again, sugars, they can form 5-membered rings, 6-memembered rings, any of the hydroxys because sugars have hydroxys in all of the carbons- any of them can react.0369

So, if they can form a stable ring, they will, sometimes 5-membered, sometimes 6-membered.0377

Now, obviously, there are going to be certain rings that are going to predominate, but in this particular case, you can actually have both.0382

Let’s go ahead and draw our linear structure again; let’s go through our systematic procedure, and it is always best to do this.0390

We have 1, 2, 3, 4 and 5.0396

This is our aldehyde, and we had the hydroxy on the right, hydroxy on the left, hydroxy on the right and our achiral carbon, so that is that.0402

Now, let’s go ahead and do the furanose form first; furanose means 5-membered ring.0412

Well this is 1 carbon; Let’s do this in black.0419

This is no. 1; this is no. 2.0424

This is no. 3; this is no. 4.0428

We got an O to react with that, so I am going to go over here.0431

This is going to be no. 5.0434

OK, in our ring, this oxygen is what is going to attack that.0436

We have to know which oxygen is going to happen.0444

When we do the pyranose in a minute, it is actually going to be this oxygen right here, down below.0447

It is going to look completely different, but it is going to be the same sugar but in a completely different form.0452

OK, what we do next is we rotate this to the right, the top; we bring it down to the right.0459

Let’s go ahead and draw that configuration.0465

Let me see, should I do it...I’ll do it over here, a little higher up, so we have plenty of rooms.0470

1, 2, 3, 4 and 5, now, I have my aldehyde here, and this drops down.0473

This hydroxy goes up; this goes down, and here I will just write it as CH2OH.0483

That is this one right here.0490

Now that I have it in a horizontal fashion, I take the left hand side and I pull it.0492

I push it away from me, and I bring it around.0497

Now, let me draw this; when I do that, I have this.0500

Let me see, make sure I have enough room here.0506

C, C, this is my aldehyde; I have a C here.0509

I have a C here, and I have a CH2OH.0515

So, make sure 1, 2, 3, 4, 5- that is correct; and let me see, on the 2, 1, 2, this hydroxy is down.0522

This hydroxy is up, and this hydroxy is down.0530

OK, we said that this hydroxy on the no. 4 carbon, this one right here, that is the one that is going to react.0535

That is the one that is going to attack the carbonyl, so what we want to do is rotate 90° upward just like this.0540

In the back, this group right here in the back is like that.0552

This is the OH; we want to rotate it like this to put the OH horizontal just so we know where this thing ends up.0556

Let’s go ahead and do that; this is going to go, let me go back to black.0565

OK, now, I am going to redraw this structure with this back group rotated 90° up and just to make the OH horizontal, just to be consistent.0572

We are going to have C, C, C, C; actually you know what, let me make it a little bit lower here.0582

I have got C, C, C, C, and now, I have my OH there; and now, I have my CH2OH here.0595

This is the oxygen; let me finish by drawing in my carbonyl.0606

This is the oxygen that is going to actually attack; this is what is going to attack from above or from below in order to get the alpha-beta.0611

Here is what happens, when that happens, and now, I will go ahead and put equilibrium arrows because the ring structure and the linear structure, there is going to be an equilibrium between those two and here is how it works.0623

We have 1, 2, 3, 4, 5; let me confirm, and, of course, a 5 carbon.0635

We put the oxygen in the back, and I decided not to put the carbons in.0640

I hope you don not mind; I will do...actually you know what, maybe I will.0646

This is O; that goes there.0655

That goes there; this goes there.0657

This goes there, something like that.0659

Now, over here, we are going to have the...I am going to not specify the stereochemistry on that.0662

It could be alpha-beta depending on whether its attack from above or attack from below, but here, no. 2 carbon, the hydroxy is down below the ring.0667

The no. 3 carbon, the hydroxy is above the ring, and, of course, here we have our CH2OH group.0676

There you go; this is D-xylofuranose.0683

Take a good look at this because it is going to look nothing like the D-xylopyranose.0691

Unspecified stereochemistry could be alpha or beta, but these are...they are definitely specified.0698

This is below the ring; this is above the ring, and you have that.0703

This is the no. 1 carbon, just a number 1, 2, 3, 4, 5 carbon 1, 2, 3, 4.0708

This is the fifth member of the ring- that is it.0718

We have to know which oxygen we are using, attached to which carbon, is actually going to form it.0721

This is why it is important to go through the systematic procedure- do this, do this, decide which is what.0727

In a minute, we will see we do not actually have to go through the rotation here when we do the pyranose form- this is it.0733

This is D-xylofuranose; This is the 5-membered ring.0739

OK, now, let’s go ahead and do our D-xylopyranose form.0743

OK, let me go back to blue, and let me start with the third structure that I drew.0749

I had the linear; I turned it around, and I brought that back.0759

Let me start with that; I have got, let me see...a little bit lower here.0761

I have got C, C; now, let me put the carbon backbone in first, C and CH2OH.0770

Let me put my aldehyde in; I have got down below.0779

I have got above the ring, and I have got this.0784

Now, I do not have to do anything to this; now, I need a 6-membered ring, the pyranose form.0788

That means that 3, 4, 5, my sixth member of the ring is this oxygen.0794

It is this oxygen that is going to attack this carbon either from above or from below.0806

Now, nothing changes over here; this hydroxy is going to stay down below, and you are going to get a 6-membered ring.0813

Now, for a 6-membered ring, of course - excuse me - the standard position is oxygen in the back right.0819

Let me go ahead and draw that; so, we have this, this, this, that, that and that.0827

Now, let’s go ahead and see what it is that we have; again, we are going to not specify the stereochemistry that can be alpha or beta, depending on attack above or below.0834

Remember, that is the no. 1 carbon; OK, the no.1 carbon.0844

On the 2 carbon - let me go back to red - the hydroxy is down.0847

On the no. 3 carbon, the hydroxy is up; on the no. 4 carbon, the hydroxy is...wait, where am I?0852

1, 2, 3, 4, 1, 2, see, now, I am getting confused, alright.0864

We have got, on the second carbon, that is down; that is up, and this is down.0871

There we go; that is the no. 4 carbon, and over here, there is nothing at all.0880

As you can see, it is easy to lose your way here.0885

So 1, down the 2, 3 and 4, that is...let me see, this is blue.0888

This is 2; this is 3 and this is 4- good, everything is good.0894

On the 5 carbon, you just have 2 hydrogens; I will leave those hydrogens off- that is it.0898

This is the D-xylopyranose; it looks nothing like the D-xylofuranose.0903

It is just a question of keeping track of which hydroxys go where, what the arrangement is, and that is it- just nice, systematic, but as you can see, you have to be really careful because it is easy to lose your way.0912

OK, let’s see.0925

I have anything, and once again, take note of the fact that this no. 5 carbon, because it is a 5 carbon sugar in the pyranose form, it is forming a 6-membered ring.0932

This carbon, the no. 5 carbon, it does not have anything on it.0943

You are used to seeing something on it either a CH2OH, either above or below, because you are so used to seeing galactose.0948

Well, you are used to seeing glucose, galactose, mannose, things like that, but again, different sugars have different things that are attached to them.0955

In this particular case, you have not missed anything; it is just 2 hydrogens that are attached there, so you are good.0962

OK, well, let’s see; let’s move on to example no. 4.0969

OK, this one is a little longer, so instead of writing it out by hand, I thought I would just present it like this.0980

A biochemist wants to synthesize a new branched polysaccharide.0986

It is an amylose chain with branching at the no. 6 carbon.0992

So, you remember, amylose is the glucose monomers; it is a homopolysaccharide.0997

It has a bunch of glucose monomers that are attached by alpha-(1,4) linkages.1002

There is going to be branching at the no. 6 of one of those glucose.1009

OK, now, the first monomer at the branch point will be the furanose form of alpha-D-xylose- we just did that.1013

The rest of the branch alternates between monomers of alpha-D-xylose and N-acetylglucosamine in the following configuration: GlcNAc-alpha-(1,3)-xylose.1021

The Glc, the N-acetylglucosamine, is connected via alpha-1, and it is connected to the no. 3 carbon of xylose in the glycosidic bond.1033

We want you to draw the structure.1044

OK, basically, real quickly, just to get a sense of what is going on, so we are going to have this amylose chain, just these glucose monomers, again I am just doing a quick schematic before I...so, that, that, that.1046

Let me just draw 4 of them; that is going to be the alpha-(1,4) of the glucose- not a problem.1064

And, let me just actually draw one more, and at the no. 6 carbon of one of those, it is going to be something like that; and it is going to be O, and it is going to be connected.1069

The first monomer of the branch point will be the furanose form of alpha-D-xylose, furanose form, so this is going to be a 5-membered ring, here.1081

Let me just go ahead and just do a little square like that.1092

It is going to be a 5-membered ring, and it is going to alternate between monomers of alpha-D-xylose and N-acetylglucosamine.1095

Let me do a little triangle for N-acetylglucosamine, a square for xylose, a triangle for N-acetylglucosamine.1102

This is what is going to happen; we have glucose monomers making up this chain.1110

At a branch point no. 6, we are going to have in alpha-D-xylose; and then we are going to alternate alpha-D-xylose, GlcNAc, Xyl, GlcNAc.1115

That is what we want to draw.1126

OK, hopefully, we either know; we have either memorized the monomer’s structure, or we just open up our books or look on the internet to get the particular monomer’s structure, and then we just construct our molecule and just remembering that it is alpha-(1,3) in the GlcNAc-xyl glycosidic bond.1127

OK, let me do this one in red; let me see here.1148

I think I will do it on - oops - I think I will do it on the next page.1153

OK, OK, so let’s go ahead and draw our alpha-(1,4)-glucose.1161

Let’s draw out 3 of those; let’s go like that.1166

Let me just go ahead and draw my...so, this is alpha-(1,4)- there we go.1174

Now, let me go ahead and put in my CH2OH.1190

Let me go ahead and draw everything in, and then I will go ahead and talk about it.1202

OH, OH, and this is down below already; that is connected on a glycosidic bond, so we have got CH2.1208

OK, now, I am going to go ahead and connect this one, so I will do a CH2 here.1219

This is my no. 6 carbon; 1, 2, 3, 4, 5, 6, my no. 6 carbon.1224

This is the one that is going to be connected, oxygen; and this is what is going to be connected here.1235

Now, I am going to have a 5-membered ring; well, my 5-membered ring, let me make this...that is OK.1243

I do not have to make it too long; my 5-membered ring is going to look like this.1251

Bam, bam, bam, bam, and, of course, here, we have our oxygen; and then this is xylose, so that is OH there.1255

Let me go ahead and put my CH2OH here.1265

Now they said that it is going to be a 1,3 glycosidic bond with the next monomer, which is the N-acetylglucosamine.1270

I’m going to go ahead and draw this way, O; and now, I will draw in the N-acetylglucosamine.1277

This is N; this is COO.1291

This is CH3; this is OH, and, of course, this is going to be O, and this is going to be alternate with that.1295

So, we have another 5-membered ring.1307

This is OH; this is going to be CH2OH, and this is going to be another O connected to something else.1312

Let’s make sure that I have got everything that I am supposed to have here.1322

This right here...I apologize; I lost my colors.1327

This right here is our Glc-alpha-(1,4)-Glc.1334

This is our main; here is a branching point at carbon no. 6.1344

The first monomer is alpha-D-xylose; this is the alpha-1.1350

This is the alpha connection; this is the xylose ring.1355

And, we said that we have a GlcNAc, N-acetylglucosamine in alpha-(1,3) to xylose-alpha-(1,3).1361

This is the no. 3 carbon; OK, this is connected that way.1380

Then, of course, this goes to another; OK, this connection right here, that is going to be xylose-alpha-(1,4)-GlcNAc.1386

So, if you want you can sort of connect this to this one; actually, let me write it right next to it.1405

Because we are alternating monomers, we have to specify at least 3 of them.1410

So, GlcNAc, this is going to be 4; this is going to be alpha-1, and this is going to be Xyl.1415

Xylose-alpha-(1,4)-glucoseNAc, at the other end of the glucose, the reducing end, that is going to be alpha-(1,3) to the xylose and so on, so xyl-GlcNAc, xyl-GlcNAc this way- that is it.1423

That is our structure; everything is taken care of.1437

All the stereochemistry is represented, the connection, 1,6 branch point, 1,3, alpha-(1,4), here- there you go.1441

That is our polysaccharide.1452

OK, good.1456

OK, now, let’s do example 5.1459

OK, example 5 is a little long in terms of just actually writing it out as far as what is going on, but it is not altogether that difficult.1464

This is a great practice in structures, and it is a great practice on actually handling a carbohydrate, how one deals with finding out certain things about it.1473

OK, now, a biochemist wants to determine the extent of branching in a sample of glycogen.1482

So, we remember glycogen molecule, it has a whole bunch of branching.1489

It is just like the starch, except it is more heavily branched and it is more compact, consists of amylose and amylopectin with 1,4 connections and then 1,6 branching at the no. 6 carbon like we just did.1492

OK, a biochemist wants to determine the extent of branching in a sample of glycogen.1509

The branching takes place on those glucose monomers that have their no. 6 carbon and hydrogen attached.1524

A certain number of those glucose monomers have branching at the alpha-(1,6).1531

We want to know what percentage of those, so mind you, we are not saying what percentage by mass.1538

We are saying what percentage, which means we are talking about number.1543

So, of let’s say, 5,000 monomers that make up some glycogen molecule, how many of those 5,000 are actually monomers of glucose that have a 1,6 branch- that is what we are asking.1548

OK, here is what he does.1560

He takes the glycogen and he treats the sample with methyl iodide in order to methylate any free hydroxy groups.1563

That means, I am going to turn the OH groups, the free hydroxy groups in glycogen into OCH3 groups.1569

I am just converting them into, I am just adding a methyl group; I am replacing this hydrogen with a CH3.1578

OK, he then completely hydrolyzes the glycogen to release the free monomers.1584

So, once I convert the free OHs to OCH3, I split up every single glycosidic bond, and I have a bunch of monomers floating around.1589

He measures the amount of (2,3)-Di-O-methyl-glucose recovered, and he makes his computation.1598

OK, the structure of (2,3)-Di-O-methyl-glucose is as follows.1608

We will draw our regular hexose ring, Di-(2,3), (2,3)-Di-O-methyl.1614

Let’s go ahead and specify; let’s just not specify it.1625

We do not have to do that; that is not important.1628

Normally, we have OH, OH, OH and CH2OH, right?1631

Di-O-methyl, (2,3)-Di-O-methyl, well, this is the no. 1 carbon, no. 2, no. 3.1641

All we have done is replace this with CH3 groups.1647

This is the molecule (2,3)-Di-O-methyl-glucose.1654

So, he methylates the glycogen; he completely hydrolyzes it to release all of the free monomers.1657

Now, remember hydrolysis, elements of water, you are splitting it up while you are adding the elements of water, splitting it up the glycosidic bonds.1663

And, what he recovers, he recovers this molecule, and he measures this molecule, the amount that he has of this molecule to determine what percentage are actually branched.1673

OK, now, let’s go ahead and see what we can do, explaining detail, what is happening in this procedure using structures.1688

OK, let’s go ahead and draw a structure, and let’s talk about what it is that is actually going on.1698

OK, let’s draw a linear; let’s draw a part of this molecule.1704

I will draw it a little bit smaller than usual just so I have room on this same page.1716

OK, boom, boom, boom.1724

OK, I will go ahead and do that, and now, let me go ahead and draw my OHs in.1728

So this goes on that way, and this goes on that way; now, there is going to be a branch point.1733

This is going to be CH2, and that is going to be O, and that is going to be like that; and, of course, it is going to go on, it is going to repeat that way.1739

Oops, there is an O there, then that.1759

OK, now, let’s go ahead and draw in what it is that we have got.1763

We have OH, OH, OH, OH, CH2OH, lots of hydroxys.1767

Carbohydrates are just full of hydroxys.1780

OK, OH, OH, CH2OH, and the last one, OH, OH; and, of course, this goes on that way, and, of course, we have a CH2OH.1784

OK, now, this is what we started off with; this is the piece of our glycogen.1801

What we do to this is we methylate it, so every free hydroxy is going to be methylated.1805

Everywhere there is a free hydroxy, you are going to end up with OCH3 instead of OH.1810

Let’s go ahead and see where those are.1816

Those are going to be, well there, there, there, there, there, there, there, there, here, here and here, here, here and here.1820

Notice something here, on the free monomers, the ones that actually do not have a 1,6, that are not branched at the no. 6 carbon, those are going to end up once you actually methylate this and once you break all of these bonds, once you break them up into individual monomers, you are going to end up with something that is methylated at 3 points- 1, 2, 3, 1, 2, 3, 1, 2, 3, 1, 2, 3.1836

But, the free monomers that end up here, there is no hydroxy attached to this.1869

This no. 6 carbon and the oxygen attached to it is involved in a glycosidic bond.1877

So, when you break that bond, what you end up with is the (2,3)-Di-methyl-glucose, and what you end up with is also down up.1882

This is the (2,3)-Di-O-methyl-glucose.1908

Once you break that, these, the ones that have the branch points, this is not available for methylation.1917

Only 2 of them are available for methylation, so you get (2,3)-Di-O-methyl-glucose.1924

All of the others that are not involved in branching, that have no branching, those are going to end up having 3 methyl groups.1928

What you are going to end up with there is this.1937

You are going to end up with an OCH3 on the no. 2.1941

You are going to end up with an OCH3 on the no. 3.1945

There is an OH here because that is just a glycosidic bond.1948

The glycosidic bonds, they just end up back as hydroxys. but these OHs, you are going to end up CH2, OCH3.1953

You are going to have (2,3,6)-Tri-O-methyl-glucose.1960

That is what is going to happen when you hydrolyze it after you methylate it.1971

When you methylate it and then hydrolyze it, you are going to end up collecting 2 types of monomers: (2,3)-Di-O-methyl-glucose and (2,3,6)-Tri-O-methyl-glucose.1975

Well, if I can measure the amount of this, that will tell me how many of these are actually involved in branching.1985

Well, if I can take the total number of moles of monomers, or if I can take the number of these that are actually involved in branching by counting this derivative of it, divide it by the total number of monomers, I have my percentage of branching.1993

That is what I am doing here; I hope that made sense.2010

OK, now, let’s go ahead and actually run the calculations.2013

OK, let me see, comes from branching one of the hydroxys, yes, branching.2018

OK, 225mg of the glycogen is treated as above.2027

24.5mg of the (2,3)-Di-O-methyl-glucose is recovered.2032

What percentage of the glucose residues in glycogen are involved in branching?2036

Assume a glucose residue is a 162g/mol.2042

The hint here is recall what we mean when we use the word residue.2047

So, just as a recollection, when we talk about a residue, we are not just not talking about an amino acid residue.2053

We can talk about a glucose residue, an amino acid residue.2059

A residue is a general term for a molecule that has the elements of water removed from it, right?2062

Hydrolysis, you take off the OH, you take of the H, that is what you are doing; that is what a residue is.2070

When we talk about an amino acid residue, it means we have taken off an OH from the carboxyl end.2077

We have taken off an H from the amino end, and we have that residue.2083

So, we have actually lost 18g/mol for individual molecule.2086

When we talk about a residue of glucose, that means it is the glucose molecule that we know; but it is missing a hydroxy, and it is missing an H.2091

In other words 18g is missing, per mole.2100

That is what residue means, and that is going to be important in just a minute.2105

OK, what we want to do is the following.2107

The amount of the (2,3)-Di-O-methyl-glucose divided by the total amount of monomer of glucose times a hundred, that is going to give us our percentage.2113

And again, we did that by derivatizing it in such a way, so that when we finally hydrolyze it, you are going to end with 2 types of monomers.2139

The ones that have the branching points only have 2 methyls- the 2,3.2145

All of the other glucose monomers have the 2,3,6- those we do not care about.2150

We care about the 2,3, not the 2,3,6.2154

OK, let’s go ahead and do the math on this one.2157

Once again, we are not doing percent by mass, so we cannot just take the 24.5, and divide it by the - what is it - 225.2164

That is the whole idea; we have to be very, very careful.2179

It did not say percent by mass or percent by volume or something else; it actually said just what is the percentage, so we are talking about numbers.2182

We cannot use the masses directly; we have to go to moles because a mole is a measure of the amount in chemistry.2190

OK, let’s go ahead and calculate what it is that we have got.2197

We have, what did we say, 225?2202

OK, we have 225mg of glycogen, the total molecule; and we said that the average glucose residue, again, glucose is involved in this polymer.2205

That means that it has undergone a condensation reaction, so the elements of water have been removed from each glucose monomer in the actual molecule.2224

When I talk about 225mg of glucose, the 162g/mol, that is the weight of the residue.2233

Now, I am going to go ahead, since it was given to me in milligrams, I am just going to go ahead and write 1mmol is 162mg.2245

You can use these prefixes milli, centi, deci, kilo, as long as you change both of them, so millimole, milligram.2258

Do not change one of them, so 162g/mol is 162mg/mmol, 162kg/kmol.2265

As long as both, then you can just use the numbers as written.2277

You don’t have to write 225mg as 0.225g- that is it.2280

It is a personal thing; I just prefer to work like this by using the numbers that they gave me.2285

You could have written 0.225g x 1mol / 162g.2290

OK, now, I am not exactly sure, but I think my arithmetic was actually wrong on this.2295

So, I am just going to write down what it is that I have on a piece of paper, but I hope that you will verify this for me.2301

I think I used the wrong number here when I did this original division, but I ended up with 1.5mmol, which I’m sure is the wrong number; but again, the number is irrelevant.2306

It is the process that is important.2317

OK, now, let’s go ahead and do our number of moles of the (2,3)-Di-O-methyl-glucose.2320

We said that we recovered 24.5mg of the 2,3 derivative; I will just put 2,3 like that.2327

Now, 1mmol of that, when you calculate the molecular weight of that, it is going to end up being 208mg.2336

So, I end up with a total of 0.1178mmol of that.2347

OK, good.2356

Now, let’s go ahead, and we are done.2360

We have that; we have that.2365

We will take 0.1178mmol divided by 1.59mmol, which I think is not the right number.2368

225 divided by 162, I do not think it is 1.59, but I hope you will double check with me; and if it is not, just use a different number- it is not a problem.2378

Times a hundred, you end up with 7.5%.2387

7.5%, of all of the individual glucose monomers in this glycogen molecule, have a branching at the no. 6 carbon.2392

The amount of branching, the extent of branching in glycogen, this particular case based on this measurement is 7.5%, which makes sense.2401

It is going to be somewhere in the range of about 7-10% branching.2411

There you go; I hope that made sense.2415

Thank you for joining us here at Educator.com.2419

We will see you next time, bye-bye.2421

Hello and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today, we are going to start our discussion of lipids, another major class of biological macromolecule.0004

Remember we have lipids; we have carbohydrates.0012

We have proteins, and we have nucleic acids- four major classes.0015

Today, we are going to start talking about lipids, and we are going to start our discussion with the simplest of them- fatty acids and the triacylglycerols.0019

Let’s go ahead and get started.0029

OK, lipids are a profoundly, profoundly diverse group of biological macromolecule.0033

Lipids are a profoundly diverse group of biological macromolecule.0047

Their common feature - but they do have one common feature - their common feature among all of these, they are not soluble in water.0072

They are not soluble in water; that is their common defining feature as far as lipid is concerned.0087

They are going to be soluble in organic solvents as far as working in the lab is concerned, or they are going to soluble in other fats.0100

They are not soluble in water.0110

OK, now, their biological functions are as diverse as their structures.0113

The particular function that we are going to talk about today, as far as fatty acids and triacylglycerols, is fuel storage.0139

That is one function that fats serve; it is a primary function that they serve, is the storage of, it is a reserve.0146

It is a fuel reserve- is what it is, lipids as fuel storage.0154

Remember when we are talking about carbohydrates, we said that glycogen was actually a form of fuel storage.0169

It is a short term form of fuel storage; fat, triacylglycerols, fatty acids, lipids- they are long term storage.0175

And, we have a bunch of fatty acid derivatives.0187

We are going to talk about fatty acids first, and then we are going to talk about its simplest derivative- the triacylglycerol.0192

OK, a fatty acid- very, very simple.0198

I do not know why they have all these different names for things that we already know.0202

A fatty acid is just a carboxylic acid; it is a carboxylic acid whose hydrocarbon portion or chain runs from 4 to about 36 carbons- that is it.0207

And, this actually includes, this number, this includes the carbonyl carbon.0247

This includes the COO carbon.0253

OK, let’s just do a quick example.0261

I will do this one in a line form because it is often best to represent these in line representations.0265

That is 2, boom, boom, boom, boom, boom, boom; we just have to keep track of how many we have.0271

So, 1, 2, 3, 4, 5, 6, 7, 8 so 1, 2, 3, 4, 5, 6, 7, 8- yes, that is correct.0277

I will put the final one there, something like that.0283

This is a C8 fatty acid; this is a C8 fatty acid- that is it.0287

It is just a carboxylic acid with a certain length of chain.0295

In this particular case 1, 2, 3, 4, 5, 6, 7, 8- that is it.0299

We can write it as...let me go to blue here.0307

We can write it as, oops, you know what, that is fine; I will just keep it as black.0312

CH3, (CH2)6 and then COOH- you will often see it like that.0317

CH3- that is the last one; we have these CH2 groups.0325

We have 6 of them, 1, 2, 3, 4, 5, 6; and then, of course, we have our carboxyl group- the COOH.0329

That is this thing right here, so you will often see them in a shorthand written like that.0337

This is n-Octanoic acid, right?0342

That is how we name the carboxylic acids: propanoic acid, butynoic acid, pentanoic acid.0350

We take the pentane, hexane, heptane, octane, and we just add oic acid- that is it; that is all it is.0356

It is a carboxylic acid with just a really long carbon chain, generally from 4 to 36- that is all, nice and simple.0363

OK, a couple of more definitions here, let’s go to blue.0371

Now, a saturated fatty acid, which you hear about all the time, fatty acid, and I am going to start writing FA for that.0379

The hydrocarbon chain, it is just a fatty acid where the hydrocarbon chain has no double bonds.0392

In other words, it is saturated with as many hydrogens as you can stick on to the carbons- that is it; that is all a saturated means.0404

You put on as many hydrogens as you can; the molecule that we just saw, it is fully saturated.0411

There is no double bond; it is all carbon, carbon single bonds, and when the carbon is not bonded to another carbon, it is bonded to hydrogens- that is all.0417

So, an unsaturated is exactly what you think; unsaturated or polyunsaturated - you will often hear that polyunsaturated - it is a fatty acid with one or more double bonds along the chain.0427

You will often hear of a monounsaturated fatty acid; it has 1 double bond.0455

You will hear of a polyunsaturated fatty acid that has 2, 3, 4, 6, 9 double bonds, however many.0460

Let’s do a saturated example, which we actually just did, but what the heck; we will do it again.0467

We will have, let’s say CH3; let’s have (CH2)18, and let’s have COOH.0479

We have 1 carbon, 18 carbons and 1 more carbon, so this is n-Eicosanoic acid; Eicosa is Greek for 20.0487

Now, the n here, the n part, n-Eicosanoic acid, that just means there is no branching in the chain.0502

It is just straight chain, there is no strange branching going on.0512

So, n just means there is no branching in the chain- that is all.0516

And, what I mean by branching is, let’s say if you have something like this, let’s say you have this, branching would mean something like that.0527

There is no branching; it is just a straight chain of 20 carbons.0537

OK, now, let’s do an unsaturated example.0545

Let’s actually draw this one out, so let’s see how many carbons have I got 1, 2, 3, 4, 5, 6, 7, 8.0559

Let’s go 2, 3, 4, 5, 6, 7, 8, 2, 3, 4, 5, 6, 7, 8, and then I will go, how about going down like that, double bond; and then 1, boom, boom.0570

That is always interesting when you have to start counting these carbons 1, 2, 3, 4, 5, 6, 7, 8, 1, 2, 3, 4, 5, 6, 7 and 8, so we have this.0595

OK, let’s make sure we have got 1, 2, 3, 4, 5, 6, 7, 8.0609

This is our 8 carbon; this is our number 1.0614

This is 9; this is 10, 11, 12, 13, 14, 15, 16, 17, 18.0616

OK, we have 18 carbons on this one.0622

This happens to be cis-9-octadecenoic acid.0628

Cis tells me - this is 9 - tells me that at the 9 carbon starting with the no.1 carbon of the carbonyl, at the 9 carbon is where the double bond starts, so between 9 and 10; that is where the double bond is.0640

Cis tells me it is the cis configuration, not the trans configuration.0654

Octadec is 18; that means it has 18 carbons total.0659

Decenoic acid- that means there is an alkene which is this thing.0664

There you go; that is all.0670

This is the systematic name, the common name.0672

All of these have systematic names, and all of these have common names.0678

This is oleic acid.; it is one of the primary components of olive oil, the fatty acid in the triacylglycerol that actually makes up olive oil- oleic acid.0681

OK, and again, take notice of the E in there because it is not octadecenoic acid.0693

That would be just the saturated version, 18 carbons, decenoic acid.0700

That is actually telling me that I have an alkene in there.0706

It is a little redundant because you have this as 9, but now, there it is.0711

OK, now, there is a notation; there are several notations, actually, but there is a notation that we will be using.0715

There is a notation for fatty acids or FAs, which gives you all the information that you need about that fatty acid, which specifies chain length.0726

It specifies the degree of unsaturation - in other words, how many double bonds there are - the degree of unsaturation and the location of those double bonds, so pretty much everything we need.0748

I think it is best to just do some examples and it will make sense that way.0772

Let’s run some examples here, so if I write 20:0.0781

OK, this first number, that is the number of carbons.0787

That is the chain length; it is the chain length.0793

That is the number of carbons you have, so in this particular case, we have a 20 carbon.0802

This second number here, the one after the colon, that is the number of double bonds there are along that chain.0807

OK, the number of double bonds along the chain.0813

In this particular case, we have a 20 carbon, nothing in there.0826

This is the eicosenoic acid- that is it, nice, straightforward, 20 carbons, no double bonds, boom; you are done.0831

At one end is the carboxylate; the rest is just a bunch of carbons and hydrogens.0838

OK, how about another one; let’s do this one in red.0842

Let’s try 18:1 and delta-9.0846

This is called the delta notation or delta x notation, and here is what it means.0852

We know what these 2 numbers mean; this is an 18 carbon, so this is 18 carbons long- that much I know.0857

I know it has 1 double bond because of the 1, and the delta-9 tells me that it is on the no. 9.0867

It starts at the no. 9 carbon, so it is between 9 and the 10, starting with the no. 1 carbon being the carbonyl carbon- that is it.0877

The delta-9, the delta part, it means the double bond starts at the no. 9 carbon, starting your count from the carbonyl carbon.0884

There is another convention that starts from the other end, that starts from the end of the chain.0922

That is called the omega notation; omega carbon is your last carbon.0930

Let me talk about an omega-3 fatty acid; that means starting from the other end of the chain, if I count 3 carbons, that is where the double bond is.0934

We are using delta notation, delta; carbonyl carbon is the no. 1 carbon.0943

Count to no. 9, that is where the double bond starts, so 18:1 delta-9- that is all that means.0948

This happens to be the oleic acid.0954

Actually, I do not need to write that; I will just write it down here.0959

This is the oleic acid; the one, the structure for which we just drew in the previous page- 18 carbons long, 1 double bond.0962

The double bond is at the no. 9 location, counting from the carbonyl being no. 1- nice and simple, really, really nice.0970

OK, let’s do just a quick example here for more practice.0979

You want as much practice as possible, although this stuff, I think, is pretty straightforward.0985

Draw the structure, so in this case, they are going to ask you for a structure of 18:3 delta 9, 12, 15- 18 carbons long.0990

OK, now, I am going to go ahead and start with my carboxylic acid on the left, instead of on the right, because I just prefer to go from left to right.1007

It does not matter; you can draw it vertically.1013

You can draw it sideways, diagonally, however you want, as long as everything is there, so totally personal choice.1016

We have 18 carbons long; we have 3 double bonds, and the double bonds take place at the 9 carbon, the 12 carbon, the 15 carbon, so between 9 and 10, 12 and 13 and 15 and 16.1024

Let’s go ahead and draw it out; I am going to draw out my carbonyl first.1036

I will go ahead and do this and that is 1, 2, 3, 4, 5, 6, 7, 8, 9.1042

So I like to...10, 11, 12, wait, 10, 10, 11, 12, 12 is up here.1053

OK, delta 9, 12, I have got another bond that is 12; this is 13, 14, 15, 15, 16, 17, 18- there we go.1067

I have 18 carbons; this is my no. 1 carbon.1083

This is my no. 18 carbon; I have 3 double bonds, 1, 2, 3, delta 9, 12, 15- there we go, nice and straight forward.1086

Now, let’s talk about some names- the systematic name.1098

This is cis, cis, and do not worry about the dashes and comas; put them wherever you want.1107

It does not really matter, 9, 12, 15-octadecatrienoic acid.1115

This actually should be one word; I just, I tend to separate them, but it is up to you- trienoic acid.1132

Again, I think it just depends on your teacher and how rigid they are about do they want it as one word, do they want it as 2 words; I separate them.1139

It is the information that is important not the aesthetics.1147

This is the systematic name, cis, cis, cis; I have actually specified the stereochemistry at each of the locations of the double bonds 9, 12, 15-octadecatrienoic acid.1151

So, it is a little redundant, trien; I have 3 alkenes.1162

I know that because I have 9, 12, 15 cis, cis, cis, so it is a bit redundant; but that is the systematic IUPAC procedure.1166

Common name, let’s see, common name, this one happens to be alpha-linolenic acid.1176

I will just write this down, so clearly the 18-3 delta 9, 12, 15 is the best representation- nice and quick.1192

It tells you everything that you need to know about it.1209

OK, now, let’s do another example; let’s go back to red.1212

We have example no. 2; this time we will give you a structure and ask for the short hand.1219

so give the short hand - in other words, give the delta notation - for cis, cis, cis.1228

No, I only have 2, 5, 8 - see again, commas, dashes, it is enough to make you crazy - hexadecadienoic acid.1244

OK, well, hexadeca, so deca is 10, undeca, dodeca, trideca, tetradeca, pentadeca, hexadeca.1266

This is 16; hexadeca means 16.1276

I have got...go to blue, let me get my blue here; there we go.1279

I have got 16 carbons; I have cis, cis, 5, 8, so I have 2 double bonds delta 5, 8- that is it.1288

I am done, nice and simple, just start counting and putting it together, no worries.1299

OK, a little bit more information here.1307

So, most naturally occurring - I hope I spelled that right - most naturally occurring unsaturated fatty acids have the cis configuration.1311

I will go ahead and put Z because those of you in organic chemistry, we have the ZE, the zusammen-entgegen notation also.1337

So cis is Z; a trans is E, so most of these fatty acids, the naturally occurring ones, they have the cis configuration.1346

Now, if the trans or E configuration shows up, which it does, occasionally, it shows up, well, the notation reflects that, then the symbol reflects that- that is it.1356

Instead of writing cis, we just write trans; or instead of writing Z, we just write E.1386

OK, example no. 3.1393

We have something like, I will do it this way; I will do the (9Z, 12E)-tetradecadienoic acid.1399

In this particular case, I decided to use the ZE, so 9Z, 12E.1418

This is going to be cis trans, probably be something like this, cis trans 9, 12; and then the rest is the same- tetradecadienoic acid.1424

You can write it as 9cis, 12trans.1441

Again, there is no hard and fast; I mean, there is a lot of information here, and these things get really, really long, so I would not lose any sleep about what it actually looks like, as long as the information is there.1448

Your teacher will tell you how they want it, and you give it to them the way that they want it.1459

If you are just learning this on your own, as long as long as you understand it and are able to communicate it, that is all that matters.1465

Again, we want to make sure that we are concentrating on what is important, not on incidentals.1471

OK, we can write this as 14; in terms of symbolism, we can write it as 14:2.1479

We can write is as cis trans, delta 9, 12; that is one way of doing it.1488

14 carbons, that is the tetradeca; 2 double bonds, that is the dien.1494

Cis trans, cis delta 9, 12, the 9 is cis; the 12 is trans.1499

You can write it as 14:2, ZE, delta 9, 12.1505

You can specify it a little bit differently, if you want.1512

You can actually go something like cis delta 9, trans delta 12.1518

If you want go ahead be a little redundant with the delta, that is fine- you can.1525

Again, these are incidentals, as long as the information is there.1528

OK, that pretty much takes care of representing it symbolically and what it is that these things are.1533

Fatty acid, it is just a long chain carboxylic acid- that is all.1540

Now, let’s go to red here.1545

No, let's write this down- the length of the hydrocarbon chain and its degree of unsaturation.1559

Saturation accounts for the properties and chemistry of these fatty acids and lipids, so how long it is, how many double bonds it has or does not have.1579

That is what is going to affect all of the physical properties and all of the ultimate chemistry.1620

OK, let’s see a little bit more.1622

Now, the longer the chain - which makes sense, so the longer the chain, the hydrocarbon chain, let’s specify this - so the longer the hydrocarbon chain and the higher the degree of saturation, the less soluble it is in water.1628

Longer chain hydrocarbons like a C24 is going to be less soluble in water than, let’s say, a C7.1670

A saturated C24 is going to be less soluble than an unsaturated C24- that is all that means.1677

Now, at physiological pH, at physio pH, 7-7.4, somewhere in that range, the carboxylic acid group, the COOH, is actually ionized, is actually COOO-.1687

So, it is deprotonated at physiological pH.1708

Therefore - little triangle of three dots means therefore - the shorter fatty acids do display some degree of solubility in water, some fair degree of solubility in water.1713

I mean, they are not going to be like sugars that just dissolve and dissolve and dissolve or almost infinitely soluble in water, but they do display some degree of solubility.1733

And again, because now at one end, you have this charged thing that can interact with water, with the hydrogen bonding, electrostatically, not just hydrogen bonding, some degree of solubility in water.1744

OK, again, let me just draw a quick structure so you might have something like that.1762

So, you have this end, and if this is not too long, you are going to actually see a fair amount of solubility- that is it.1773

OK, also, the longer the chain and the fewer the double bonds, the higher the melting point.1781

This has very, very, important, important application in all kinds of things in our modern world.1809

OK, let’s take a look at this one, and talk about why this is the case.1822

When you have a certain compound, the measure of a melting point that the compound is going to come together, it is going to interact with itself.1828

The molecules are going to interact with themselves.1834

So, the melting point is a measure of how much energy I need to put in to it, in order to turn the solid into a liquid, to separate them out- that is all that is.1838

Well, one that is fully saturated is going to look something like this.1847

Let’s just have this end, so we have one end, and, of course, we have this long straight chain, no double bonds.1853

Well, I am going to represent this this way; I am going to represent it as that and just a little tail like that.1861

OK, so this is the polar end, and this is the nonpolar chain.1867

Well, in the case of something that does have a double bond, so actually does have some degree of unsaturation, you are going to end up with something like this.1871

It is going to bend, so what you are going to end up with is something like that.1885

There is going to be a kink in it; and, of course, the greater the degree of unsaturation, the more double bonds, the more kinky it is going to be.1889

Well, when these things aggregate, which they tend to do, you end up with something like this.1895

In these saturated compounds, because there is no kink, you will end up with something...they interact very, very tightly.1907

The hydrocarbon portions, they basically just lay on top of each other, and there is a lot of Van der Waals interactions, so they are pretty tightly bound.1924

It takes a hell of a lot of energy to actually separate them out.1931

In the case of the unsaturated fatty acids, when they aggregate, again, they are bent now.1935

They cannot really lay on top of each other all that much.1944

They are not as tightly bound, so the interaction among the hydrocarbon chains is not as strong.1952

Because it is not as strong, it takes less energy to convert them from a solid to a liquid- that is all that means.1959

And it is strictly a structural feature simply because they are bent and they are kinky- that is what happens.1967

They are not going to interact as much as the hydrocarbon portions.1972

There is going to be less Van Der Waals interaction.1977

So, you do not need as much energy to break them apart and turn them into a liquid, so they can slide all over each other- that is all that is happening.1980

A little bit of information, in the C12 to C24 range, saturated fatty acids are solids at room temperature - just think of lard - whereas unsaturated fatty acids are liquid oils, olive oil - that is it.1990

On the C12 to C24 range, saturated, they tend to be solid room temperature, 25°C, somewhere around there; and the unsaturated fatty acids are going to be liquid oils.2042

OK. now, let’s talk about triacylglycerols.2054

Now, triacylglycerols- very, very, very important group.2061

These are the simplest of the lipids, which are constructed from the fatty acids.2071

In some sense, the fatty acids are sort of analogy; well, you know what, no, there is no analogy here, never mind.2099

Sorry, sorry I brought it up.2107

OK, a triacylglycerol, which is just ALA ,not going to keep writing that, so I am just going to write as TAG.2108

TAGs or esters are esters of glycerol, and glycerol is the molecule OH, OH, OH, H2, H; and I will put an H2.2116

Glycerol- it is a 3 carbon molecule; each of those carbons has a hydroxy attached to it, so basically, it is just a triple alcohol, glycerol.2142

A triacylglycerol are esters of these - I should just have done this on a one page; sorry about that - where fatty acids are attached, where the OHs were before; and I will draw it out in just a minute.2153

Do not worry.2181

In other words, let’s go ahead and draw out glycerol again.2186

We have C, C, C; this is OH.2194

This is OH; this is OH, and it is very, very important to remember that this is a schematic.2197

Carbon is a tetrahedral molecule, so you are looking at something in free space.2206

When we draw it like this, we are draw it for convenience, so that we can see what the connections are.2211

In space it looks totally different than how I am drawing it.2215

This is glycerol, just to have it on this page.2219

And now, a triacylglycerol, it is just where these hydroxys are replaced with some fatty acid.2223

Let's go ahead and draw this again; let me do this one in black, actually.2228

I have C, C and C; I have O.2234

I have COO, and some R1; R1 is some hydrocarbon portion.2241

I have O, and I have COO, and I have R2; that is another one, and I will go ahead and put this one here.2247

This is O, and CO, and R3; so this is triacylglycerol.2263

You have the glycerol molecule, the 3 carbons; and then, you have these fatty acids that are attached.2272

Now, the fatty acids can all be the same; they can all be different.2282

You are going to have 2 of 1 and 1 of the other; all kinds of combinations are possible.2286

Now, these are called the triacylglycerols, and it makes perfect sense- triacylglycerol.2293

OK, now, they are also called tryglycerides.2307

You will often hear them talked about that way- triglycerides or just fats.2321

That is it, general term of what we know of as fat.2327

Now, of course, you have heard of mono and diglyceride.2331

A monoglyceride is just glycerol where there is 1 fatty acid attached.2333

A diglyceride is glycerol with 2 fatty acids attached, and the alcohol is the third one; it is a hydroxy.2338

A triacylglycerol or a triglyceride, it just means all of the alcohols have been replaced by fatty acid residues- that is it.2346

OK, again, R1, R2 and R3 can be the same.2355

If they are the same, if R1 is the same as R2, is the same as R3, we just call it a simple triacylglycerol; and if not, then, we call it a mixed- nothing strange about that.2359

OK, now, let's go ahead and do an example of this.2387

However, before I do that, what is important to remember, here, this glycerol, this has the hydroxys attached.2393

When glycerol actually reacts with fatty acids, it is going to go through 3 reactions.2402

Each one of the alcohol groups is going to react with a fatty acid.2406

When it does so, this oxygen right here, these oxygens, they actually belong to the glycerol; and I will talk a little bit more about that in just a minute, but let’s just do a quick example.2411

Let’s do C and C and C, and let’s go O, C, O.2434

Let’s go CH2.2448

I do not know, however many, and then, of course, CH3.2454

Let’s see, 16, I think this is going to be 14, 15, 16, so this is 12.2460

No, this is going to be 14, I think.2465

So, let’s go ahead and use palmitic acid, and for this one, let’s go ahead and do O.2470

We have CO, and then let’s do CH2.2476

Let’s have 7 of these; let’s have a CH, a double bond, a CH; and let’s have another 7 of these, and over here, let’s go ahead and do an O and a C, and let’s go ahead and do 10 methylene groups.2482

This is CH2; there is 10 of those, and we have a CH3.2502

Here, on this side, this particular fatty acid that is attached, this is going to be palmitic acid.2507

I will do this in a different color here, go ahead and go back to black.2515

This is palmitic acid that is attached to the no. 1.2520

This is a 16:0 fatty acid; this one happens to be oleic.2528

This is our 18:1 delta-9, and this is a 10, 13, 14, this is myristic.2534

This is myristic acid, and it is a 14:0 acid; and that is it.2544

If I wanted to name this, here is how I would name it.2556

This is no.1 carbon, 2 carbon, 3 carbon; I have to specify which fatty acid is attached to which carbon.2558

Just go ahead and number them, so this becomes 1. Palmitoyl - I drop the I-C and I add O-Y-L - 2. Oleoyl, 3. Myristoyl glycerol - that is it.2564

If you have to name it, it is probably not going to be too much of an issue, but that is it.2594

Just drop the I-C on the common name, and just go ahead and add O-Y-L to it, and there you go, so O-Y-L, O-Y-L. O-Y-L.2597

Now, let me go ahead and just go back here and do...so, this is oxygen, that is right here, OK.2612

These oxygens, that oxygen, that oxygen, these Os, they come from glycerol.2628

OK, they do not come from the fatty acid; they come from glycerol.2632

And just as a quick recap, let us recall ester formation.2637

OK, I am going to go ahead and draw out my gly...I will do this one in black.2652

I have C, C, C; I have CHOH.2654

I have CH2OH, and I have CH2OH.2662

I am going to go ahead and draw a couple of the electrons on those just to know which one is attacking which, and let me go ahead and draw in my fatty acid.2668

I am going to do CH3, CH2.2675

Let’s just do 6 of them, and then let’s go COO; and then, O and then, let’s go PO32-- there.2680

So, basically what I have done is instead of O-, I just have a little bit of phosphate in the body.2695

This thing right here is going to act as a leaving group, so it needs to be activated first because I cannot just attack it.2702

The O is not going to go anywhere; I have to convert it into a good leaving group - which is what the body does - by reacting this thing with adenosine triphosphate, and then it puts the phosphate on here, so now, this is a really, really good leaving group.2708

Here is how it happens; this is a nucleophile.2722

This carbon is an electrophile; it kicks the electrons up.2726

Tetrahedral intermediate kicks the electrons back down.2730

This goes away, and then, of course, what you end up with, once you actually take away the H that is attached right there, you end up with the following.2734

You end up with...I am going to do it in a reverse way.2745

Actually you know what, no, I will go ahead and just keep it like this.2751

C, C, C, I have an O; let me go ahead and write the H.2755

This is OH; this is H2.2765

This is OH; this is H2.2769

And now, let me go to a different color.2773

O, I have C, O, and I have (CH2)6; and I have CH3- that is what is going on.2778

When this reacts with that, so you notice, this oxygen, this ester linkage is actually, it comes from the glycerol; it is not that oxygen.2780

This oxygen that is originally attached to the carbon, that goes away with the phosphate as part of the leaving group.2782

And, of course, now, these are free to react with other fatty acids to actually form our triacylglycerol, so just a quick recap on ester formation.2795

This oxygen, the ester linkage on the fatty acid, this thing right here, it comes from the glycerol.2805

OK, now, let’s see; let’s talk a little bit about storage.2813

Now, we said - we are almost done, no worries - we said the body stores fuel.2840

Actually, it stores glycogen as a fuel source, glycogen as a reserve fuel source, and it does.2857

That is true; we did not lie to you.2867

The reserve fuel source, OK, reserve fuel source, forgetting how to spell today, I do not know what is going on.2874

Fuel, is it F-E-U or F-U?2884

It is F-E-U, yes.2886

OK, now, the body’s primary fuel reserve is not glycogen.2887

Fuel reserve, reserves, are stored as triacylglycerols in fat cells.2902

Fat cells are called adipocytes or adipocytes.2919

Again, it just depends on where you want to put your stress; it does not really matter.2923

Yes, it is true that the body does store glycogen as a fuel reserve, but that will last maybe a day, if you are lucky.2928

Probably not even that, maybe just 5 or 6 hours.2935

The body’s primary fuel reserves, the one that goes to when you really are not ingesting any food, it is fat.2938

Fat is how it actually stores most of its fuel reserves; That is triacylglycerols.2947

So, glycogen is just if you need something quickly, but if you need something over an extended period of time, it is going to go to the fat stores.2953

OK, now, let’s see, 2.2962

I will go ahead and do this in red.2968

There are 2 primary advantages, so you are probably thinking to yourself "Wait a minute, why not just store it as glycogen, why does it have to store anything that is fat at all?".2974

There are 2 primary advantages to storing energy as triacylglycerols instead of glycogen.2985

Now, the first one, since the...do I have another page?3005

Yes, I do.3014

Now, since the hydrocarbon portion of fatty acids on the triacylglycerols - wow, we have got a whole bunch of acronyms here - so since the hydrocarbons of the fatty acids on the triacylglycerols are more reduced than sugars - and more reduced means they have more hydrogens on them, more oxidized means they have more oxygen attached to them - sugars, they have a whole bunch of hydroxys attached to every single carbon.3015

Those carbons are reasonably oxidized, not fully oxidized yet; they are reasonably oxidized.3053

The fatty acids, they have no oxygens attached to them at all; they are all hydrogens, so they are completely reduced.3060

There is more energy available- that is the whole point.3065

Since the hydrocarbon portions of the fatty acids on the triacylglycerols are more reduced than the sugars, oxidation of fatty acids releases about 2 times the energy of sugar oxidation, so that is one reason.3068

If I store my energy as glycogen, there is a certain amount of energy that I am going to get out of it.3101

If I store the same amount of fat, I end up getting twice as much energy, gram per gram - that is the reason why, one of the reasons why.3105

OK, the second reason, and this is an interesting one.3114

Now, triacylglycerols are completely nonpolar; they are completely hydrophobic.3120

The fatty acids, we said, have a little bit of a polar end; but that is tied up now, in an ester linkage in a triacylglycerol, so it is all hydrocarbon all over the place.3129

There is nothing, there is no polar part for it to actually interact with water at all, so triacylglycerols are hydrophobic.3138

They do not want to be anywhere near water.3150

They do not bind water unlike carbohydrates, which have a bunch of highly polar molecules, bunch of hydrogen bonding going on.3158

Each sugar molecule is just surrounded with water, unlike carbohydrates.3173

They do not have the extra weight of the water attached that comes with hydration, the extra water weight associated with stored glycogen, which runs at about a 2:1 ratio, 2:1 grams of water to grams of carbohydrate.3187

So, when the body stores glycogen, because glycogen is a carbohydrate, there is a whole bunch of water that is associated with that carbohydrate.3236

It is heavily hydrated; it is very, very, very hydrophilic.3243

There is a whole bunch of water attached to it.3247

For every gram of carbohydrate that is stored as glycogen, there is about 2g of water attached to that.3250

So, you can imagine, if you, let’s say, have a kilogram of glycogen that you are storing, that is going to be 2kg of extra water weight that you are carrying because triacylglycerols are nonpolar.3256

They do not associate with water.3270

So storing, you can store a whole bunch of fat and not have to store all of the water that comes with it simply because it is hydrophobic, and that is it - 2 major advantages for using triacylglycerols and fats as long term fuel storage, that is it.3272

Thank you for joining us here at Educator.com.3289

We will see you next time for a further discussion of lipids, bye-bye.3291

Hello, and welcome back to Educator.com, and welcome back to Biochemistry.0000

Today, we are going to continue our discussion of lipid biochemistry by talking about membrane lipids; these are structural lipids.0004

On the last lesson, we talked about the storage lipids, the triacylglycerols.0011

Today, we are going to be talking about the structural lipids, the lipids, the fats that actually show up in biological membranes, in the cell membranes.0016

Let’s get started.0025

OK, let’s go ahead and write down a couple of things.0028

Yes, that is fine; I guess we can stick with black here.0033

Your membrane lipids are lipids that make up the biological membranes of cells, the bilayer - and we are going to be looking at some illustrations a little bit later on - in cells or reside in the membrane.0038

In the case of something like cholesterol, it is not so much that it makes up the fats in the membrane.0078

It just happens to reside there, or reside in the membrane among the other types of lipids.0085

Again, it is just sort of the other types of lipids that actually make up the membrane.0092

It is just a question of perspective, whether one considers it to be something that actually makes up the membrane or happens to just be there.0101

In another cell, it is not that important; what is important is the lipid, the structure and the function.0108

OK, they have a hydrophilic end.0114

Let’s call it polar end and a hydrophobic tail.0126

Again, it looks something like this; we generally tend to draw it like that, either a single or usually a double.0140

In the case of the lipids that we are going to be talking about today, this is the hydrophobic tail; this is the polar end.0146

And again, we will be looking at some illustrations a little bit later, after we talk about some structures.0151

OK, we are going to be talking about five major classes of membrane lipids.0157

We will discuss five major classes of the membrane lipids, and we are going to be taking about the glycerophospholipids.0166

OK, I have to warn you here.0188

As you have probably already noticed with biochemistry, the nomenclature, the number of the names, they tend to not only get very, very long, but we tend to have multiple names for the same thing.0192

For a student, it is very, very daunting because often we will be talking about, let's say we will mention 3 or 4 different names of something but we are actually talking about the same class of molecule; and I understand it is an annoyance.0203

So, sometimes we are going to be calling them glycerophospholipids; you will hear them talked about as phosphoglycerides.0216

Sometimes, the name is so completely different that you are wondering, you really think that they are different molecules and they are not.0223

I have to apologize for that; that is just the nature of biochemistry.0230

Different people from all around the world call them different things; people within a certain specific research community refer to them differently.0234

These are just names that you are going to get accustomed to hearing over and over and over again.0242

I will try my best to be consistent, but in all honesty, I think it is also a good idea to hear the multiple different names and to know that we are talking about the same class of molecule or the same molecule itself, so just a little warning; but you already figured that out.0247

OK, glycerophospholipids, and then the second major class we are going to talk about is going to be the galactolipids.0261

Let’s see, the third class, something called the tetraether lipids, and then, we have something called the sphingolipids, and then, we are going to talk about the steriles.0270

OK, let’s go ahead and start with our glycerophospholipids, and I think I will go ahead and go to blue here, so glycerophospholipids, glycerophospholipid.0293

Now, these are the ones that are also called the phosphoglycerides.0314

Now, the name pretty much says it all, glycerophospholipids; you have a glycerol backbone.0327

It contains a phosphate somewhere, and its glyceride part, it basically has some fatty acids that are attached to it.0332

Let’s go ahead and look at the general structure; I think I will go ahead and do it this way.0340

We have carbon, carbon, carbon; that is going to be our glycerol backbone.0346

We have O; we have O, and let me go ahead and put this O a little bit down here.0351

We have one fatty acid; I will go ahead and call it R1.0358

It could be any particular length that could have some double bonds.0361

It could have no double bonds, the saturated or unsaturated.0366

We have a second one attached; I will go ahead and call this one R2.0370

Rather than choosing a specific example of a fatty acid, I will just go ahead and leave them as R1 and R2.0374

And here, here is where we have our phosphate; and then, of course, we have...so, I am going to go ahead and put a little bit of an X there, this X.0380

This is the general structure of a glycerophospholipid; let me go ahead and finish with my hydrogens here, make sure I have all of those.0391

There is an H2 here, and we have an H here.0399

We have the glycerol backbone; this is your glycerol right here.0403

And then, of course, we have a fatty acid attached to one of them, a fatty attached, another one, in an ester linkage - right - C double bond O, single bond O.0410

And then, of course, to this third oxygen, we have a phosphate group attached; and then we have this thing right here, this XO.0420

I am going to put a little square around this, and the reason I am going to do that is the following.0428

We are going to be doing some specific examples of what this X group is, but it is really, really important to know that this oxygen, over here, that is attached to the phosphate and attached to the X actually comes from the X group.0433

The X is some kind of an alcohol, something that has a hydroxy group on it.0444

This oxygen actually comes from that alcohol.0448

Later on, when you study the actual biosynthesis of lipids, then you will actually see where each individual atom comes from; but for now, it is good to know that this X group, it is actually an XOH.0452

It is actually some alcohol that is attached to this phosphate- there you go.0469

And also, this oxygen right here, it actually comes from the glycerol.0475

All right, now, and, of course, here is our phosphate group right there.0479

A couple of things, now, the X group is what changes.0486

So, this is what is variable; the rest of the molecule is pretty much fixed.0493

I mean, it is true; R1 and R2 can be different.0496

Generally, there will not be a huge variety; they can be different, but what actually characterizes that particular glycerophospholipid is this XO group, the alcohol that is attached.0500

Group is what changes and gives a particular glycerophospholipid its name.0512

OK, X is the polar group, so this is the polar group; and these R1 and R2, these long hydrocarbon chains, that is going to end up being the nonpolar tail.0531

X is the polar group and is an alcohol like we said.0549

OK, now, when X = H, when X is equivalent, when it is just a hydrogen, then the molecule is called a phosphatidic acid.0566

We often refer to these as derivatives of phosphatidic acid because the X is going to change.0594

When it is just H, when it is just POOOH, it is phosphatidic acid or A phosphatidic acid.0599

OK, R1 and R2 are variable.0608

R...well, I do not need to write that; you know that already, that is why we call them R1 and R2.0613

OK, and at physiological pH, at physio pH, the phosphate group carries a -1 charge.0616

That is very, very important.0630

Charge on these lipids is actually very, very important; it is going to affect the biochemistry.0636

OK, that is why I have this -1 here; and in general, I am going to be putting circles around my charges, so that I can actually see them clearly and add them up.0642

OK, let’s do some examples here.0651

We have examples; let’s go this way.0655

I am going to draw this a little bit different; I am going to put the tails on one end, then I am going to the polar head group on the other.0662

Let me go ahead and put C, C and C.0668

I am going to put the esters on this side.0673

COO, this is R1; this is going to be one of the tail ends in ester linkage, and this is going to be that.0676

This is going to be R2, and over here, and I hope you will forgive me if I leave off my hydrogens.0684

I tend to leave off my hydrogens; if you see a carbon that has two bonds attached to it, the other 2 are going to be hydrogens.0689

That is just how it is.0695

O phosphate, O, there is - oops, let me go ahead and make that negative sign a little bit clearer, a little circle around it - O, and then if we have C, C, NH3+, this is called, so this right here - I do not know which one I should...yes, it is fine - I will go ahead and call it the whole thing.0699

This is ethanolamine; this molecule is called ethanolamine.0726

The name of this whole thing - OK - of the glycerophospholipid is, let me write it out, lamine, M-ethanolamine; the regular molecule without the attachment, is exactly what it sounds like.0734

It is ethanol, 2 carbons and a hydroxy and an amine group attached right there.0760

This is ethanolamine before it is actually attached.0765

Again, this oxygen actually comes from the alcohol.0772

Ethanolamine, this is called - depending on where you want to put the stress again with pronunciation - phosphatidylethanolamine or phosphatidylethanolamine.0776

It is up to you; again, pronunciation is unimportant.0784

You will often see this written as one word; I tend to write it as two words.0787

Again, it is going to be up to your teacher, about how strict they are with things like that.0791

What is important is the chemistry and the structure.0797

This part, the phosphatidyl, that is this basic structure; and then this other name right here, depends on what it is that is attached.0801

Let’s do another molecule; let’s do a C, a C, a C.0815

OK, let’s go O, C, R1, O, C.0821

This is R2, and then, we have O, P, our phosphate with a negative charge.0830

We have O, and this time, we have C, C; we have the nitrogen again, but this time, we have a CH3, a CH3 and a CH3.0837

And again, there is a positive charge on there, negative charge here.0848

There is a positive; there is a negative charge.0853

Notice the net charge on the ethanolamine derivative is 0 because the nitrogen is carrying a positive.0856

The phosphate is carrying a -1, so this is a net charge of 0; so this is a neutral glycerophospholipid.0865

This one also, it is a neutral glycerophospholipid; these things are going to be very, very important.0871

In a minute you will see an example of one that is not neutral, so positive, negative, neutral, charges- very, very important.0876

This one is called - oops - phosphatidylcholine because we have this right here.0883

This group is called choline.0898

Ethanolamine has 3 hydrogens; choline has 3 methyl groups attached to the nitrogen.0903

And again, nitrogen has 4 things attached to it, so it is carrying a formal charge of +1.0908

OK, let’s take a look at another one.0914

We have got C, and we have got C, and C, O, C.0920

This is R1, O, C; and this is R2.0929

We have our O; we have our phosphate group.0935

The oxygen is carrying a -1 charge; we have our O there.0938

And now, we have C and C; we have N there.0943

We have C here; I will go ahead and put, yes, I will go ahead and just leave it like this.0948

And then, this is going to be NH3+, and there is a negative.0956

There is a negative, and hopefully you will recognize this particular molecule right here, this particular residue.0963

This is serine; here is the N, C, C.0971

OK, this is an amino acid; and here is the C, and this is an OH, so it is an alcohol, right?0976

Serine has, its R-group has CH2 and then OH.0983

This is phosphatidylserine- that is it.0986

Now, what is the net charge on here?0994

Well, we have -1, +1, -1, so the net charge here is -1.0996

Looks different and because it is a different, it is going to have a different biochemistry- net charge, -1.1002

OK, just to reiterate that, the charges on the head groups.1009

When we say head groups, we are talking about the polar group.1024

This right here, this whole thing, is referred to as the head group; and that, right there, is going to be the tail.1027

Again, this R1 and R2 - I probably should have drawn them out - these are just long hydrocarbon chains- that is it.1038

That is all they are, the fatty acid chains, the 14, 16, 18, 20, 22, 24, 26, length, saturated, unsaturated.1043

That is what these are, so that is the tail group; this is the head group.1053

We often refer to it that way; the charges on the head groups are important and do have consequences at the layer - that is fine, I will just go ahead and write it this way - at the head group, water interface.1057

The charge is going to affect how that particular group reacts or interfaces with whether it is in the cytosol or outside of the cell.1079

The aqueous environment, it is going to have an effect on the biochemistry there.1092

OK, let me see if I should do a little, yes, let’s go ahead and give a schematic version of this.1099

Just schematically, basically, you are going to have your glycerol.1106

OK, you are going to have glycerol, and then you are going to have some fatty acid attached in ester linkage, another fatty acid attached in ester linkage.1119

You are going to have a PO3-, and then you are going to have some alcohol.1151

This is for the schematic representation of a phosphoglyceride, glycerophospholipid- that is it, glycerol, 2 fatty acids, phosphate, diester linkage here, a phosphodiester.1156

In other words, it is an ester linkage with a phosphate instead of a carbon, OC, OC.1171

This is called a phosphodiester because this is phosphodiester linkage.1180

That is your 1 ester; this is another ester.1187

That is you are phosphate group and it is attached to some alcohol; that is the general schematic for that.1190

OK, let’s see what else we have got here.1195

OK, slight variation.1203

Some glycerophospholipids have 1 of the 2 fatty acids connected to glycerol with ether linkages instead of ester linkages.1209

OK, let’s do some examples here, and I will go ahead and keep these in black.1251

Let’s see, so let’s go C, C, C; and this time, I am going to go O, C, double bond C and some R1.1260

I will go ahead and - yes that is fine - I will go ahead and leave it like that, and then this one will be O.1274

This is our ester linkage, so I will go ahead and leave this as R2.1281

And over here we have our O; we have our phosphate group.1286

So, this is the head group, and I will go and leave it as, let me see O, C, C, C; and I will go ahead and just put (CH3)3 there and this, oops, that is not a C, that is a nitrogen, because it is choline.1291

That is that; that is that.1312

OK, when you have something like this, same basic structure - you have the phosphor, you have the alcohol group - one of the fatty acids is the normal fatty acid connected in ester linkage, but one of the fatty acids, instead of an ester right here, instead of an oxygen double bonded to a carbon, what you have is these 2.1315

The first carbon and the second carbon are attached in alkene linkage.1337

OK, this right here, this is our alkene connection.1342

I do not know why I am having such a hard time writing today.1355

This is our alkene, and, of course, this right here - we will do it in blue - this is our ether linkage.1359

OK, it is not the ester linkage, the C, double bond O, single bond O, C; it is C, O, C single bond.1365

This right here, this is your ether linkage, and the general name for this class of molecule, it is still a glycerophospholipid.1373

It just happens to be called a, again, a plasmalogen or a plasmalogen, depending on where you want to put the stress; and these are characterized by the alkene linkage at no. 1, no. 2 carbon.1390

OK, let’s go ahead and move on to our second class, which is going to be the galactolipids.1404

Let me go ahead and start drawing a little bit of a line here.1413

Let me do this one in red.1417

Our second class, these are going to be our galactolipids.1421

OK, these predominate in plant cells; where the glycerophospholipids tend to predominate in animal cells, these predominate in plant cells.1430

OK, and they are characterize by having 1 or 2 galactose monomers; and you remember galactose, it is the c4-epimer of glucose.1446

Monomers is connected; it is connected to the no. 3 carbon of glycerol - yes, that is right - glycerol while the other 2 have their 2 fatty acids, while the other 2 Cs are attached to fatty acids, as usual.1462

OK, let’s go ahead and do a structure for a monogalactolipid and a digalactolipid, so 1 or 2 monomers attached.1507

Let’s go ahead and do a monogalactolipid; this time, let’s do it this way.1519

We will go C, C, and we will do C here.1525

Here we have our normal fatty acid linkage; we will call it R1.1530

This one is going to be R2, and now, this is carbon.1535

So, we have oxygen - wait, let me do this a little bit differently, carbon, no, that is alright, I will go ahead and do it this way - oxygen, OK, there, there, there, there.1545

And, of course, we have our galactose, so this down.1564

This is up; this is up and this is up.1568

This is our galactose monomer, and let me go ahead and draw those in, just for the heck of it.1573

OK, here we go; we have our glycerol backbone- 1 fatty acid, 2 fatty acids.1585

And here, instead of phosphate, the sugar is just directly attached, so this is your anomeric carbon right here, right?1593

And notice, you have - let me do this in blue - this is the beta-1.1602

The beta configuration is actually going up.1609

This is our galactose monomer, and a couple of words, R1 and R2 are generally the same.1612

They can be different, but they are generally the same; and they tend to be linoleic acid, which is better represented as 18:2 delta 9, 12- there is that.1626

This is often called an MGDG stands for monogalactodiacylglycerol- that is it.1646

Diacyl, 2, monogalacto has one sugar- that is it.1665

Now, a couple of things you want to note to be in a net neutral.1670

The charge on this is neutral; nothing is ionized here.1678

Nothing is carrying a charge, so it is net neutral.1683

There is no charge on the head group, and, of course, the beta configuration at the anomeric carbon of the galactose.1686

OK, now, let’s go ahead and do a digalactolipid.1693

I will go ahead and draw that down here; let me do this in blue.1699

Actually, let me do it in red again.1703

I have got C; I have got C.1705

I have got C; let me make sure I have enough room here.1708

Yes, I should have enough room; this is O, C.1711

It is really interesting; when you actually draw these structures out by hand, it is kind of nice to look at illustrations and see them passively because when you are drawing them out by hand, especially these ester linkages, you will often forget an oxygen or forget a carbon or something like that.1717

It is really kind of, it is interesting; but it a good idea to be able to reproduce them by hand.1731

We have C; also, hopefully, I am not forgetting any of my atoms here because that would be really bad.1738

OK, let’s go here.1745

And we said we have O, and we have this, that O.1751

That is one of them, and then we have our CH2; and I will go ahead and put that there.1762

This is going to be O; I will go ahead and do it this way, that, that, that, that, that, that, that.1771

I think that is right, so we have a down.1781

This is going to be up; this is going to be up.1785

That is one galactose monomer, and notice, 1, this is the anomeric carbon.1789

Notice we have a - let me do this in black - this is going to be beta-1 configuration.1794

Over here, 1, 2, 3, 4, 5 connected to the 6 carbon.1800

OK, this is going to be a, this is an alpha, so we are on alpha-(1,6) glycosidic bond, between 1 galactose monomer and another galactose monomer.1808

And, let me go ahead and put the rest of the substituents in here.1817

This down; this is up.1821

This is up, and, of course, we have our CH2OH, which is up; and I hope that you will confirm this for me, but I think I have got everything here.1824

R1, this is R2, not R1, R2.1835

Again, they tend to be the same, but that is OK; we will go ahead and leave it as R2.1840

This is called a DGDG- digalactodiacylglycerol.1842

Again, just names, not all together that important.1851

Notice net neutral beta-1 configuration connected to glycerol, alpha-(1,6) configuration connected from 1 galactose monomer to the other galactose monomer, galactose, galactose, c4-epimer.1855

So, this is up instead of down like it would be for glucose- that is it, just a little variation, that is all.1870

OK, now, what I am going to do is, I am just going to go ahead and jump to some illustrations.1879

Instead of drawn out by hand, I would like you to actually see what it is that they are going to look like in your book or in any other media source that you happen to consult.1885

Let’s take a look at some of these things, so we have a better sense of what is going on.1895

OK, first of all, let’s take a look at a nice, general...we are going to work our way down.1900

We are going to look a little bit deeper in magnification.1910

We have our general lipid bilayer; this is a nice cell membrane.1913

We see some integral proteins; we see the lipid bilayer.1917

This is our top layer here; this is our bottom layer there.1924

We have outside of the cell; we have inside of the cell.1929

We would call this the outer leaflet; We would call this the inner leaflet, and you can see this little blue thing, this is our head group, and you can see the tails a little bit.1931

So, when we magnify this, now, we are going to come over to this level, so nice cross-sectional view.1942

We have our head groups; these are the head groups.1949

These are the tails; notice, there are 2 tails attached.1954

These are our glycerophospholipids; in general, you have the 2 fatty acid tails attached.1958

They are going to arrange themselves like this, again, because polar with polar, nonpolar with nonpolar.1965

These are nonpolar tails attaching with nonpolar tails- outer leaflet, inner leaflet.1970

This is something called a micelle; often times, they will arrange themselves.1978

One layer of these can arrange itself in a circle and create this little container, if you will, where the inside is actually nonpolar; the outside is polar.1981

The soaps and detergents that you use, they often do it like this; they trap the oil inside here, but they interact with water outside here because these things are polar.1996

OK, now, let’s magnify a little bit more.2006

Now, we have an idea of what it is that they look like.2010

This is what you might call the, well, separation; this is what they are showing.2012

What you have here, in this particular case, we have our glycerol right there.2022

OK, this is our glycerol; we have one of the fatty acids attached.2030

We have another fatty acid attached; here is our phosphate group.2033

And then, of course, in this particular case, we have C, C, N; we have C, C, C, so this is choline.2038

This is our polar head group.2045

OK, that is that, and these down here, that is that.2048

In this particular case, same thing, except we have ethanolamine instead of choline, but we have our phosphate; and here, we have our serine.2058

Again, that is the tail; that is the tail, and this is our phosphatidylserine, phosphatidylethanolamine, phosphatidylcholine.2070

This is the general structure; this is what you see.2078

This is one of these, is one of these- there you go.2082

Again, you have seen this stuff before, should be reasonably familiar.2088

Now, we are just getting to the actual structure of these lipids.2092

OK, this is a nice blown up view of a cross section of a cell.2097

In the outer leaflet, the inner leaflet - let me go ahead and go to blue - in the outside of the cell, the inside of the cell, this is going to be, let see, sphingomyelin, cholesterol, phosphatidylcholine.2106

A couple of these, we actually have not talked about yet; they are going to be the other classes that we are going to talk about, the sphingolipids and the sterols, but this one, we have talked about.2122

Here, you have your glycerol; you have your phosphate.2132

You have your choline.2138

OK, this is the outer leaflet of the membrane, inner leaflet of the membrane; and then, of course, you have your tail group, and, of course, you have your tail group.2141

And these are just some other lipids, some other fats that are actually - well, we should not call them fats, let’s just call them lipids - some other lipids that are in the outer leaflet, which we will talk about in just a little bit.2153

Here we have our phosphatidylserine; here we have another cholesterol.2164

We have phosphatidylethanolamine, and we have phosphatidylinositol, which again, we will talk about in subsequent lessons- that is it.2169

It is just an arrangement of these lipids, the various lipid classes arranging themselves and making up the cell membrane- that is all.2177

OK, a couple of structures here, so here, we have a monogalactolipid.2191

This one is a monogalactolipid; this is the MGDG, OK, or the monogalactodiacylglycerol.2197

This is the digalactodiacylglycerol; this is our galactose monomer, down, up, down, down.2210

Now, notice in this particular case, they actually show you the stereochemistry in a different way.2220

Instead of looking at it in Howarth projection, they are looking at it directly like this.2224

Again, just another way of looking at the molecular structure of something.2230

You are going to often see structures like this; that is the thing with biochemistry- different projections, different views give us different bits of information.2235

I just wanted you to see something a little bit than a Haworth projection, back, forward, forward, forward.2245

Here, we have our 2 galactose monomers; they are connected in what we said was alpha-(1,6) because this is the 1, 2, 3, 4, 5, 6 carbon, right there.2252

This is going to be a bet