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Raffi Hovasapian

Raffi Hovasapian

Peptide Synthesis (Merrifield Process)

Slide Duration:

Table of Contents

I. Preliminaries on Aqueous Chemistry
Aqueous Solutions & Concentration

39m 57s

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

38m 53s

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

29m 1s

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

39m 11s

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

41m 33s

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

44m 19s

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

18m 45s

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

38m 19s

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

27m 14s

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

48m 28s

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

45m 18s

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

42m 47s

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

1h 2m 33s

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)

49m 12s

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

54m 31s

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

50m 52s

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

51m 36s

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

1h 3m 36s

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

1h 7m 16s

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

41m 38s

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

44m 2s

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

56m 40s

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

20m 37s

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

51m 37s

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

51m 23s

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

54m 49s

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

1h 17m 46s

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
1:01:39
Haworth Projection
1:07:34
Pyranose & Furanose Overview
1:07:38
Haworth Projection: Pyranoses
1:09:30
Haworth Projection: Furanose
1:14:56
Hexose Derivatives & Reducing Sugars

37m 6s

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

43m 32s

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

39m 25s

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

44m 15s

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

44m 23s

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

40m 22s

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

54m 55s

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

38m 51s

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

38m 20s

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

48m 36s

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

45m 51s

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

37m 6s

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

44m 32s

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

30m 8s

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

49m 46s

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

56m 34s

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

42m 12s

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

43m 32s

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

1h 1m 47s

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

59m 17s

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

39m 47s

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

41m 34s

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

34m 18s

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

42m 52s

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

36m 10s

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

49m 20s

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

44m 11s

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

48m 11s

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

45m 58s

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

33m 18s

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

40m 59s

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

39m 18s

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

36m 21s

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

47m 58s

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

41m 11s

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

36m 27s

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
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Lecture Comments (7)

1 answer

Last reply by: Professor Hovasapian
Fri Feb 26, 2016 1:50 AM

Post by Jinhai Zhang on February 16, 2016

Dear Prof:
when you mention in the lecture that Ala is the N-teminus and Ser is the C-terminus you based upon the condition from you are given, Ala-Gly-Ser(N-->C), so if the condition becomes Ser-Ala-Gly now, and we can infer that Ser is the N-terminus and Gly is the C-terminus, is that right? Thanks!

1 answer

Last reply by: Professor Hovasapian
Sun Feb 8, 2015 3:02 PM

Post by Richard Meador on February 6, 2015

How is the bead constructed?  I read where the beads are typically 50 um in diameter.   In the lesson, you show one serine monomer attached to the bead but I assume there are thousands of serine monomers attached to each bead so that after synthesis there are thousands of polypeptides per bead?? Do the recommended textbooks cover this in detail? Thanks for your help.

1 answer

Last reply by: Professor Hovasapian
Fri Aug 29, 2014 7:28 PM

Post by David Gonzalez on August 26, 2014

What are some of the applications of the finalized protein once it has been synthesized? Can it be integrated into an organism? If so, is there a lesson on that? Thank you professor.

0 answers

Post by Matthew Humes on October 1, 2013

Is there any way to download all of the images/slides at once? I can't seem to find a way other then saving them all individually one at a time...is there an easier way?

Peptide Synthesis (Merrifield Process)

Lecture Slides are screen-captured images of important points in the lecture. Students can download and print out these lecture slide images to do practice problems as well as take notes while watching the lecture.

  • Intro 0:00
  • Peptide Synthesis (Merrifield Process) 0:31
    • Introduction to Synthesizing Peptides
    • Merrifield Peptide Synthesis: General Scheme
    • So What Do We Do?
    • Synthesis of Protein in the Body Vs. The Merrifield Process
  • Example: Synthesis of Ala-Gly-Ser 9:21
    • Synthesis of Ala-Gly-Ser: Reactions Overview
    • Synthesis of Ala-Gly-Ser: Reaction 1
    • Synthesis of Ala-Gly-Ser: Reaction 2
    • Synthesis of Ala-Gly-Ser: Reaction 3
    • Synthesis of Ala-Gly-Ser: Reaction 4 & 4a
    • Synthesis of Ala-Gly-Ser: Reaction 5
    • Synthesis of Ala-Gly-Ser: Reaction 6
    • Synthesis of Ala-Gly-Ser: Reaction 7 & 7a
    • Synthesis of Ala-Gly-Ser: Reaction 8
    • Synthesis of Ala-Gly-Ser: Reaction 9 & 10
  • Chromatography: Eluent, Stationary Phase, and Eluate 45:55

Transcription: Peptide Synthesis (Merrifield Process)

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

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