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Lecture Comments (2)

1 answer

Last reply by: Professor Hovasapian
Thu Feb 27, 2014 7:35 PM

Post by Yanet Ortiz on February 26, 2014

Great lecture!!! very clear explanation!!! thank you

Protein Function III: More on Hemoglobin

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
  • Protein Function III: More on Hemoglobin 0:11
    • Two Models for Cooperative Binding: MWC & Sequential Model
    • MWC Model
    • Hemoglobin Subunits
    • Sequential Model
    • Hemoglobin Transports H⁺ & CO₂
    • Binding Sites of H⁺ and CO₂
    • CO₂ is Converted to Bicarbonate
    • Production of H⁺ & CO₂ in Tissues
    • H⁺ & CO₂ Binding are Inversely Related to O₂ Binding
    • The H⁺ Bohr Effect: His¹⁴⁶ Residue on the β Subunits
    • Heterotropic Allosteric Regulation of O₂ Binding by 2,3-Biphosphoglycerate (2,3 BPG)
    • Binding Curve for 2,3 BPG

Transcription: Protein Function III: More on Hemoglobin

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

Today, we are going to continue our discussion of hemoglobin and discuss cooperative binding a little bit more.0004

OK, let’s get started.0010

OK, now, as far as the cooperative binding of hemoglobin is concerned, there are 2 models that have been proposed for how this actually happens.0014

Let’s see what we have here.0023

There are 2 models for cooperative binding.0029

And again, cooperative binding just refers to the fact that when one particular ligand attaches to one part of this hemoglobin molecule, it causes certain changes to take place in the conformation - things like that - that makes the binding of the second, third and subsequent ligands either a little easier or a little harder.0039

That is all cooperative binding means; it means it is not just a simple process of bind, unbind, bind, unbind.0059

There is more interaction going on.0066

Two models for cooperative binding, they are called the MWC model, and the MWC just refers to the last names of the men who proposed this and the sequential model, OK, MWC model and sequential model.0070

Now, for the MWC, the basic assumptions of the model is that the subunits of the protein - and again, in our case, the protein that we are interested in is the hemoglobin, so we have the 4 subunits, the 2 alphas and the 2 betas, I will go ahead and write hemoglobin in this case - they function identically - OK - that they exist in 2 conformations; and that all subunits, they transition from one state to the other state simultaneously.0091

Excuse me.0160

OK, in other words, in the case of hemoglobin, everything is either going to be in the T-state, the tensed state, the deoxyhemoglobin state or everything is going to be in the R-state, the relaxed state, the oxyhemoglobin.0181

Remember, the T-state is the one that does not want to bind H2, does not want to bind O2 - I am sorry - not H2.0198

The R-state is the one that does tend to bind O2.0205

Let’s go ahead, and let me go to blue for this one.0214

That is for hemoglobin, Hb, all the subunits are either T or R- one or the other.0220

OK, pictorially this is what it looks like; you are going to get something like this.0239

It is going to take a couple of minutes to draw out this picture here; we have 4 subunits in our hemoglobin molecule.0244

We have 1, 2, 3, 4; I will just do it that way, and then, let’s see.0251

I hope I have enough room here, I should; let me make my...I am sorry.0260

I am going to make my circles a little bit smaller; I would like all of these to be on one page.0262

I am going to go 1, 2, 3, 4, and I will do my equilibrium arrows afterward actually, 2, 3, 4, 1, 2, 3, 4.0266

That is 2, 1, 2, 3, 4; that is 3.0280

1, 2, 3, 4, that is 4, and 1, 2, 3, 4, that is 5.0282

That is going to be our 4 subunits of hemoglobin, 1, 2, 3, 4, the 2 alphas, the 2 betas.0288

This is going to be the T-state; we will circle, we will represent the T-state.0293

Now, I will go ahead and draw the R-state.0298

The R-state is going to be represented as squares; let me go ahead and draw them in, and then, we will talk about the movements between these states.0302

Here we go; this is our T-state over here on the left, and we have our R-state.0315

And again, for the MWC model, they are either all in the T-state or all in the R-state.0319

What you have is a bunch of equilibriums that exist between all of these forms.0325

Let me go ahead and draw in all of my equilibrium arrows, and I will talk about this in just a minute.0331

Just let me make sure to get all these drawn in; I hope I have not forgotten anything here.0345

OK, now, I am going to go ahead and put 1 ligand which is the oxygen.0350

I will go ahead and do this in red; that has 1 ligand attached, and of course, when it makes the transition, this one has 2 ligands attached, and then, in the case of oxygen, the ligand we are talking about is diatomic oxygen, oxygen gas, O2.0354

Let’s go ahead and, now, do a 3 L, L, L, L, L, L, and, of course, our fourth final, of course, has 4.0372

Each of the subunits has an O2 attached; the MWC model, it looks something like this.0385

You start out with, let’s say, 4 empty subunits.0391

It is in equilibrium with the R-state of the 4 empty subunits.0396

From here, either one of these can bind a ligand.0401

If this one binds one ligand, 1 O2 molecule, OK, now, this one might bind another one.0404

It might go this way, the transition from the T-state to the R-state completely.0411

Again, the MWC model says that the transitions take place for all of them simultaneously, all T-state or all R-state.0416

Maybe, it makes the transition over here, and then, maybe, now, in the R-state, it binds another ligand.0425

Well, we know that in the R-state, as more ligands bind, there is more of a transition and the O2 tends to bind easier.0431

Maybe, it does not make the transition back here, or maybe, a few of the enzymes actually do make it back here.0440

They bind the third, and maybe it comes back here; at some point, we go from all empty to all full, and all of these equilibriums exist.0446

It can go this way, this way, this way, this way, this way, this way, this way; they can go this way, this way.0454

That is what all of these equilibrium arrows mean; these are pathways that it can actually follow to get from no binding to every single subunit is completely bound by its ligand, which is oxygen in this case.0460

This is a pictorial model of how the MWC mechanism works.0473

OK, now, in sequential model, I will just say…actually, let me go back to blue here.0479

I like the blue very much.0490

OK, in sequential binding, each individual subunit can be in either state, can be in either T - well, I will just say in either state - and the conformational change in 1 subunit can induce a change in the other or others, can induce a change in another subunit or others.0494

It can be one or more; a change in one subunit can cause, maybe, another subunit to change, or it might cause a second subunit to change, or it might cause all 3 subunits to change, but they are free.0557

Each one is individual; they are not necessarily...they are cooperative in the sense that they affect each other, but they do not all have to be T or all have to be R- either one can be either.0567

OK, induce a change in the other causing the ligand or ligand - again, pronunciation - to bind more tightly.0578

And we know, again, T-state to R-state, as it makes the transition from the T-state to the R-state, the O2 is bound more tightly.0598

OK, now, let’s do a pictorial version of this; the sequential binding, the pictorial version of this has actually a lot of those states.0607

In the previous picture that we saw, there were not too many; this one has quite a few because each individual subunit is free to be in either state.0615

I am going to draw a portion of it that involves change in 2 of the subunits.0624

It actually has more when you have changes in all 4 subunits, but I am only going to do a portion of it; but you should be able to understand what it is that is going on.0630

You should be able to hopefully, you will actually finish off the picture.0639

Let me go ahead and do this, you know what, this time, I think I will do black.0644

Excuse me; I have got 1, 2, 3, 4, 1, 2, 3, 4, 1, 2, 3, 4.0651

OK, this time, I have got 1, 2, 3, and this one is a square.0662

I have got 1, 2, 3; this top right is a square, and I have got 1, 2, 3.0670

This top right is a square.0675

OK, this time, I have circle, circle, square, square or T-state, R-state.0679

The Ts are the circles; the squares are the R: circle, circle, square, square and circle, circle, square, square0685

Alright, now, let’s go ahead and draw in our equilibrium arrows; these are going to be a little bit more complex.0697

It is not going to be just horizontal and vertical; there is actually going to be equilibriums that exist diagonally.0702

Let me go ahead and put the...let’s be nice and systematic; let’s go horizontal first, get those out of the way.0708

Now, let’s go ahead and do the vertical, there - actually I prefer that - and then, this way and that way and this way and that way, this way and that way, this way and that way.0714

And now, we have an equilibrium that exists here also, so equilibriums all over the place.0728

OK, now, let’s go ahead and fill in with the ligand, our oxygen.0736

We can have it in this state where there are all the T-state.0742

We can have the transition of 3 of them can be in the T-state; 1 of them can be in the R-state.0748

Two of them can be in the T-state; 2 of them can be in the R-state, and what I mean by you finishing off, that this is being just a portion of the whole picture, the next column over will be 3 squares and 1 circle, and then the next column will be 4 squares.0750

The same thing down here, it is just a little broader.0764

I am just doing a portion of it here; these can be all empty.0767

They can be all empty; they can be all full.0772

One could be empty, the others full; one could be full, the others empty.0774

Let’s go ahead and draw in some ligands here: L, L, L, now, for 2 ligands bound.0779

OK, and again, it goes a little bit further this way and a little bit further this way.0792

OK, all this means is that at any given time, any subunit can be in any state at any given time with ligand or without.0798

That is what is key here; OK, any subunit can be in any state, and by "any", we mean the 2 states, the T or the R.0808

In the case of hemoglobin, another enzyme another protein might have 5 states.0823

It could be in any of those 5- any state at any time with ligand or without.0829

All kinds of possibilities are available here.0843

You can start here; maybe, you can bind the ligand to the T-state.0848

Maybe, this particular subunit starts to make the transition, itself, from the T-state to the R-state, and now, it has moved over here.0853

Now, maybe, this one decides to bind another ligand, and it binds it over here; and then, maybe this subunit, it bounded in the T-state, but now, it has gone ahead and transitioned to the R-state, or maybe, this one, this particular subunit, when this one binds, it causes a conformational change in this subunit over here on the bottom left.0861

It causes it to transition into the R-state, and then, the R-state actually binds another ligand.0882

We can go here or here; that is what these arrows mean.0888

They are pathways that this particular protein can take to get from one place to another.0892

It does not have to always be in one way; if I wanted to go from here, the top left where everything is in the T-state, and everything is empty, all the way down here where I have 2 of them in the T-state, 2 in the R-state and the ligands happen to be in the R-state subunits, I can go 1, 2, 3, 4.0898

That is 1 path; I can go 1, 2, 3, 4.0918

That is another path; I can go 1, 2, 3, 4, 5, 6, 7, 8, 9.0920

It is an infinite number of pathways.0926

What this model represents is pictorially, it gives you an idea of the complexity of the interactions of cooperative binding.0928

It is not a simple thing; we can describe it, but the idea is to understand that this is very, very complex and very, very subtle.0937

This gives you an idea of just how complex and how subtle.0947

Pathways, double arrow, if the arrows were in one direction, it would actually minimize the pathways, but it can transition from any state to any state with ligand or without.0954

OK, this is the sequential model for binding.0961

Now, you should now, just because 2 models have been proposed, it does not necessarily mean that they are mutually exclusively.0965

We are not saying that it is either sequential or MWC.0971

The same protein can and will often demonstrate both.0975

Maybe, sometimes the binding will happen in a sequential pattern.0980

Maybe, the binding will happen in an MWC pattern, in a simultaneously pattern.0984

Again, when we propose models in science, we are not necessarily being exclusive of the other models especially, when it comes to something as complex as biochemistry.0988

Lots of things are going on, and we still do not have a full grasp on what and why.0997

That is what makes biochemistry so rich, so full, so exciting, such a fertile field for any of you that are interested in research- so many wonderful things to discover.1003

OK, that takes care of the cooperative binding of hemoglobin.1014

Now, we are going to go on, and I am going to talk about a couple of the other things, other ligands that hemoglobin actually binds.1020

As it turns out, hemoglobin does not just bind oxygen.1026

It binds hydrogen ion; it binds CO2, and it binds something called biphosphoglycerate, which is a molecule that regulates the extent to which it binds O2.1030

Now, let’s go ahead and discuss that.1040

Now, let me go back to blue; I like blue.1044

Hemoglobin, in addition to the oxygen, it also transports hydrogen ion and CO2.1050

Hemoglobin not only carries O2 to the tissues - I should say from the lung to the tissues - but also carries the hydrogen ion and CO2 from the tissues to the lung.1070

I mean, in terms of physiological efficiency, in terms of just efficiency alone, I mean, it makes sense.1114

Why are you going to have this molecule that is going to go, take the oxygen from the lung, deliver it to all of the tissues and then, you are going to have another molecule that has to bring those waste products, the CO2, and the hydrogen ion that are created by the metabolic processes the body needs to get rid of them?1119

Why have another enzyme, another protein floating around, that is just going to have to bring them back?1138

You can have the same molecule, hemoglobin, do double duty.1142

It will take the oxygen; it will bring back the H+ and the CO2.1146

It will take the oxygen, bring back the CO2; that is what is so amazing about this hemoglobin molecule.1150

I mean, it really is kind of, sort of, staggers the imagination when you think of this collection of a couple of a hundred of amino acids and look at it what it does and it does it so beautifully and so perfectly and for so very long- absolutely magnificent.1155

OK, let’s see here.1170

Now, very, very important, the H+ - excuse me - and CO2, they do not bind to the same site that O2 binds.1176

O2 binds to the iron in heme; the H+ and the CO2 do not bind to the iron in heme.1192

Actually I will not say Fe2+; I am going to say iron.1209

I am going to say 2+ 3+ because again, once it know what, I am just going to say "does not bind to the Fe in heme"- there we go.1218

OK, the H+ binds to one of many amino acids that make up the protein- makes sense.1231

We have plenty of nitrogens and oxygens that can actually be protonated by this H+, so it can bind anywhere.1253

Now, CO2 binds to the NH2 group, to the amino group at the amino terminal - oops, excuse me - end of each subunit forming something called carbaminohemoglobin.1261

The name does not matter actually.1310

Again, a lot of these names, they are actually not that important; they are just extra fancy stuff that just really gets in the way.1317

As long as you understand that CO2 is actually binding to an amino group, that is at the amino terminal of each subunit, that is what matters.1323

Here is the reaction that takes place; let me go ahead and go back know what, I think I will do this in red.1331

We have our CO2 molecule plus I will do H2N.1336

We have a C; this I going to be the amino terminal here, then, we have C, and of course, the polypeptide chain of the protein goes on.1344

This is the amino terminal end.1356

This reacts, CO2 reacts with this, and what you end up with is the following.1360

You end up with C - I will go ahead and make it a little bent here - N, H, C, H, N, C.1366

Yes, here we go; let's just make sure we keep track of all these atoms- there we go.1379

We end up with this thing right here.1386

OK, the amino terminal end of the subunit, OK, this amino group, it acts as a nucleophile.1390

It attacks here; it pushes these electrons up, and it binds CO2.1399

That is where the CO2 binds; now, let's go back to black.1404

OK, the CO2 produced in metabolic processes is converted to bicarbonate to make it soluble in aqueous solution.1412

OK, in some of the CO2 that is produced in metabolic processes, about 20% actually, is going to be bound to hemoglobin.1449

Hemoglobin is what is actually going to take it back to the lungs to release it; that is one of the things that you exhale.1457

Not all of this CO2 that is created in the metabolic processes is actually bound up by hemoglobin, only a part of it is, again, about 20%, maybe, 25%.1464

The other CO2 produced in metabolic processes, CO2 is not soluble in aqueous solution.1474

In other words, if I bubble CO2 into water, the bubbles come up because it is not going to dissolve in the water.1480

The only time that CO2 is actually soluble in water is when I put it under high pressure, which is what you get in soda.1488

That is why soda has the fizz.1495

What we have done is we have actually pushed CO2 into the water, and in that process, by putting all that pressure on it, we can make sure it stays in the water.1499

When it is in the water, it reacts with the water to form carbonic acid.1508

Well, the CO2 that is produced in metabolic processes, the part that is not actually attached to hemoglobin directly, is converted to bicarbonate in order to keep it soluble in aqueous solution in the blood, which is not a high pressure system - excuse me - under conditions of normal pressure.1512

When we do not have the pressure to actually keep the CO2 dissolved, we have to turn it to something else to make sure it does dissolved.1548

We have to turn it into an ionic species - bicarbonate, normal pressure - and here is what it looks like.1552

Let's see here; let me see.1563

Let me go to the next page here; what we have is CO2 is going to react with the H2O - OK - and it is going to produce HCO3- + H+.1570

OK, this reaction is catalyzed by an enzyme called carbonic anhydrase- very, very important enzyme.1588

We are not going to talk about it here, but you definitely want to know this carbonic anhydrase.1595

OK, here is your bicarbonate right here, OK, hydrogen bicarbonate, carbonate.1600

Notice that 1 of the things that is actually produced in this is H+.1608

High CO2 concentration actually ends up producing a high H+ concentration, high acid concentration, which means a lower pH.1614

That is what CO2 does; there is more CO2 in the body.1623

If you cannot exhale it fast enough, it is actually going to start producing H+, and it is going to start to lower the pH of the blood.1626

OK, high CO2 concentration lowers the pH because high CO2 concentration...what is not attached to hemoglobin will actually react with water to produce bicarbonate and hydrogen ion.1634

Alright, now, as we said - excuse me - hemoglobin accounts for about 40% of the H+ and about 20% of the CO2 produced in tissues during metabolic processes.1648

What happens to the rest, well, the rest is actually absorbed in this bicarbonate and carbonate buffer system.1688

This is part of the buffer system of the blood, the carbonate, bicarbonate buffer system, that controls the amount of CO2 and controls the amount of H+ in the blood to make sure that it remains at a reasonably stable pH, somewhere in the range of about 7.2.1695

OK, now, H+ and CO2 binding are inversely related to O2 binding, and you know what inverse means.1712

The more that hydrogen ion and CO2 bind, they cause conformational changes, which makes it less likely for O2 to bind and vice versa.1739

As more O2 binds, it makes it less likely for hemoglobin to bind CO2 and hydrogen ion.1748

This makes sense because of what hemoglobin does, delivers the O2, drops its pay load, delivers the O2 to the tissues.1755

Now that the O2 is no longer bound, now, it is going to pick the H+ and the CO2 and take it back to the lungs- it makes sense.1763

Now, under conditions of low pH - and again, low pH means high hydrogen ion concentration - and high CO2 concentration, the affinity - excuse me - of hemoglobin for O2 is lowered.1772

It releases its O2 to the tissues.1819

Again, the hemoglobin travels to the tissues.1831

The tissues tend to have a very, very high CO2 and a high acid concentration.1836

Under those conditions, the binding of O2, the affinity of hemoglobin for O2 drops.1840

When the affinity drops, it releases its O2, which is exactly what we want to happen.1850

It is not just pressure, the low 4 kPa pressure at the tissues versus the 12 or 13 kPa of the lungs.1855

It is not just the pressure that actually affects the extent to which hemoglobin will bind O2 or release O2.1863

It is also the presence of CO2 and hydrogen ion, and as we will see a little bit later, the presence of this regulating molecule called 2,3-biphosphoglycerate.1871

OK, now, in the lungs, as CO2 is released and the pH comes back up to normal - in other words, under conditions of normal pH - hemoglobin’s affinity for O2 rises.1883

Now, it wants O2, which is perfect because the lungs are flooded with oxygen gas.1913

It binds the O2.1918

Let’s try to make this a little bit more legible, shall we?1925

Hemoglobin's affinity for O2, it rises, and it binds oxygen, which exactly what we want it to do.1936

OK, now, let’s go back to blue here.1955

The effect of H+ and CO2 on O2 binding and release - well, there is a name for it - by hemoglobin is called the Bohr effect.1964

It is named after Christian Bohr, who was Niels Bohr father.1994

It is not a Niels Bohr effect; it is a Christian Bohr effect- the Bohr effect.1998

The extent to which the hydrogen ion and CO2 actually affect the binding and release of oxygen by hemoglobin, we give it the name the Bohr effect.2002

OK, now, let’s go a little bit deeper into this, not too, much but a little bit of detail is good.2012

A major contributor to the hydrogen ion Bohr effect is the histidine 146 residue on the beta subunits.2022

This beta subunit, it is a polypeptide chain.2063

The 146 amino acid on that chain is a histidine.2067

OK, now, under conditions of low pH...and again, low pH means high hydrogen ion concentration.2073

It is really, really easy to confuse that; I actually confuse it all the time.2090

When we talk about...which is why I personally prefer to talk about high hydrogen concentration, low hydrogen ion concentration because then, I am actually speaking about something that is high or low directly.2094

When we talk about low pH, because pH is a negative logarithmic scale, a low pH means high hydrogen ion concentration- very, very acidic.2106

High pH is low hydrogen ion concentration- very, very basic.2117

Again, in the literature and just in general, you are going to find that they use pH to describe concentration of H+ as opposed to directly saying high or low hydrogen ion concentration.2123

So, be very, very careful with that, stop for a second just to make sure you understand- low pH, high hydrogen ion concentration.2136

I will go ahead and put it in here: high hydrogen ion concentration.2143

And I know my brackets are not exactly pretty, but there it is.2149

OK, under conditions of high hydrogen ion concentration, low pH, this histidine 146 actually becomes protonated, which makes sense, and it forms an ion pair, in other words, an ionic bond, an ion-ion bond, ion pair with aspartate 94.2152

The aspartate 94, you have got a negative charge on it; the protonated histidine has a positive charge on it.2189

Well, negative and positive, they tend to attract each other; they form an ion pair.2196

OK, this ion pair, this positive negative attraction - OK - it actually stabilizes the T-state of hemoglobin, and the T-state is the deoxyhemoglobin state.2201

It is the state that does not bind O2 very easily; in fact, it wants to give up its O2.2227

Sure enough, the more the hydrogen ion concentration, the more acidic this histidine 146 is protonated.2233

It is going to form some ionic pair with this aspartate 94 that is going to, sort of, lock the hemoglobin in to this T-state.2242

It is not going to bind to O2; that is where this comes from.2251

It stabilizes the T-state, and the T-state is the deoxy state- deoxyhemoglobin.2256

OK, now, the bound CO2 that we saw earlier, that is bound to the amino terminal - OK - also forms ion pairs.2267

Again, when we bound the CO2, you notice that one of the double bonds actually moved up to the oxygen to form a negative bond on one of the oxygens.2292

The oxygen is carrying a formal charge of negative one.2301

That is going to form an ion pair with other things that happen to be positively charged, which also tends to stabilize the T-state.2304

The bound CO2 also forms ion pairs that stabilize T-state- both of them do, the deoxy.2313

OK, the conclusion, really, what we want you to take away from all of this is the fact that cooperative binding is profoundly complex.2329

Cooperative binding in hemoglobin - well, cooperative binding in anything that actually binds ligands - is a very complex function, in not only O2 but also H+ and CO2- there you go.2346

OK, now we are going to start.2378

Now, let's talk about the regulation binding, the hemoglobin’s binding of O2 via this molecule called 2,3-biohosphoglycerate- just BPG as what we call it.2383

Let’s start on that, and I think I am actually going to keep it in blue because I like the blue.2393

Regulation of O2 binding by 2,3-biphosphoglycerate, and by now, you are accustomed to this nice long words in biochemistry, otherwise known as just 2,3-BPG or just BPG.2400

Regulation of O2 binding by 2,3-biphosphoglycerate is heterotropic allosteric regulation.2436

OK, here we go; regulation of O2 binding by 2,3-BPG is heterotropic allosteric regulation2459

Let's recall what this means; remember we said we have this enzyme - not enzyme, this protein - that binds some ligand, in this case, O2.2467

Well, there is another site on this protein that another ligand combined, and this binding of this ligand can actually affect the extent to which the main ligand, the O2, binds.2478

Heterotropic means that it is a different molecule altogether.2492

Allosteric means it binds to another site on the protein and regulation is just regulation- that is all.2496

This is just a fancy word for the fact that there is another molecule that is going to control the extent to which O2 binds- that is all.2503

I mean in some sense, the H+ and the CO2 that we have talked about are also allosteric heterotropic modulators of O2 binding, but we tend to call 2,3-BPG, we tend to refer to that more so because it is a larger molecule; but for all practical purposes, they are all doing the same thing.2511

They are not the same molecule as O2, and they control how O2 binds.2532

OK, let's draw the structure for 2-BPG and see what is going on here.2538

Let’s see; let’s write 2-BPG.2543

We have got...let me do this...that is OK.2549

I will go ahead and do it in black; I have got C, C and C.2552

I will go ahead and do this as COO-; I will go ahead and just write this as PO32-, and I will go ahead and put a PO32- here.2559

That is fine; I will go ahead and put the hydrogens in- that is it.2573

2,3-biphosphoglycerate, this is the no. 1 - I will go back to blue - no. 2 carbon, no. 3 carbon, 2,3-biphosphoglycerate, this carboxylic acid a molecule right there.2577

OK, now I am going to go ahead and stay with blue here.2596

Now, BPG, the biphosphoglycerate is always present.2601

When we talk about biphosphoglycerate as regulating the amount of O2 that is binding, we are not talking about that it is not bound to hemoglobin, and then, all of a sudden, when BPG does bind to hemoglobin, things change.2605

The BPG is always bound to hemoglobin.2618

What we are discussing here is that when you go to higher altitudes, it is the increase in the BPG concentration on the blood.2621

Therefore, more BPG starts to bind to hemoglobin.2628

BPG is always present.2633

In fact, the normal binding curves that we saw in previous lessons for hemoglobin, that actually includes the bound BPG that is always there with the hemoglobin.2639

OK, in fact, the standard binding curves we have discussed, or I should say we have seen so far, involve hemoglobin with bound BPG.2654

I repeat again, we are concerned not with the binding of BPG.2681

We are concerned with the binding of more BPG.2685

OK, now, let me see the binding of BPG.2691

BPG and hemoglobin’s affinity for oxygen, again, are inversely related.2700

What that means is, as the BPG concentration rises, that means that the affinity for O2 by hemoglobin, drops.2718

That means, hemoglobin does not want O2 as badly as it did before.2730

Hemoglobin's affinity for O2 decreases.2737

As the BPG - let me make this arrow a little bit better - concentration increases, well, when a concentration increases, more BPG is going to bind to hemoglobin.2745

That causes hemoglobin to have less affinity for O2; it is going to want to get rid of its O2, or another way of looking at it, it does not want to bind O2 as strongly.2757

It just depends on your perspective.2765

OK, let’s take a look and see what is going on; let’s say a few more words actually, and then, we will go ahead and draw what this looks like.2769

We will draw a couple of binding curves, standard binding curve and then, the binding curve with the extra BPG that is bound to the hemoglobin.2774

Now, at sea level - that is fine, I will go ahead and stay with the blue - the BPG concentration is somewhere in the neighborhood of 5mM.2784

Now, at about 4500 feet, as you go up to about 4500 feet or 5000 feet, now, what happens is the concentration of BPG in the blood starts to increase - OK - and it actually is about 8mM.2804

The concentration of BPG goes up, so more BPG is going to bind to the hemoglobin.2827

Now, here is what is important; here is how this works.2831

BPG does not - repeat - does not affect the binding of O2 very much in the lungs but effects the release of O2 in the tissues very, very much.2837

I will write very, very much.2880

OK, let me talk a little bit about what is actually happening here.2887

Now, recall that at sea level, that hemoglobin delivers about 40% of the oxygen that is bound to it, delivers about 40% of its bound O2.2891

OK, 100% saturated hemoglobin is fully saturated with O2.2920

It only delivers in the blood; only about 40 % of the oxygen that is available is delivered to the tissues.2926

It keeps about 60%.2930

Now, well, let me go ahead and write it out and then, discuss it.2934

At high altitudes, the availability of oxygen is less.2942

In other words, the partial pressure of oxygen is less.2953

There is less oxygen; that is all that means.2956

The pO2 is reduced; OK, there is less oxygen.2958

The pO2 is reduced, so less O2 binds to hemoglobin.2966

Therefore, less O2 is delivered to the tissues.2979

Now, this is before anything happens; OK, this is before any BPG happens.2992

If I am at sea level, 40% of my oxygen that is bound to the hemoglobin is being delivered to my tissues.2996

If I, all of a sudden, take a plane, take a train, whatever, and all of a sudden, I am at 4500 feet sea level, well, there is less oxygen up there.3002

Before anything happens in the body, all of a sudden there is less oxygen binding to the O2.3010

However, the body will still only deliver up to a certain amount, but because there is less O2 having bound, there is less O2 delivered; and here is what it looks like pictorially.3016

Let's say that this is a 100% here and here; OK, now, let’s say this is a 100% of O2.3030

Well, we said that it binds; we said that it releases about 40% of it.3036

About that much is released; this right here, this remaining 60%, that stays bound.3041

Well, if you go to higher elevation, before anything happens, OK, before BPG has a chance to do what it does, now, instead of binding a 100%, now, it is binding 90%.3047

There is less oxygen to bind; all of the hemoglobin is now no longer saturated.3058

However, when it gets to the tissues, it still releases that amount, that same amount.3063

Now, instead of 100%, now, because less is bound, it is still going to deliver the same amount from the 0 mark.3069

Now, instead of delivering this much, it delivers this much.3080

The bottom end, the delivery end, that stays the same, but because there is less oxygen bound to hemoglobin, there is less oxygen delivered because hemoglobin will always keep about 60% of its oxygen.3086

From 60 to a hundred is 40, but now, if only 90% is saturated, 60 to 30 is 30%.3103

That is how our bodies experience the diminished oxygenation.3106

It is because it delivers the same level, but there is less to deliver.3111

What BPG does is at higher elevations, once the BPG concentration rises, the BPG binds to the hemoglobin.3118

Now, it does not affect too much, as far as the binding of hemoglobin in the lungs.3127

Let's say from a 100%, it binds 90%, but now, instead of delivering from 60-90, now, what happens, is it actually ends up delivering more.3133

Now, the 40% or the 60% drops down to 50 to 40 to 30.3145

Although, it does bind less because of the increased BPG concentration, it ends up delivering more, so here, here, here, here.3152

OK, once I have gone to a higher elevation, yes, it is true that less has bound, but now, I am dropping this down here.3165

So, more is delivered, and it brings it back up to the 40% level, which was the original amount that the body needs in order to sustain its natural function.3175

Do not worry about this; I am going to actually write it out and then, I am going to show you what this looks like on a graph on a binding curve.3185

I will be discussing this again in just a moment; let me just go ahead and write a few things down here, though.3192

Now, OK, what did I write?3198

At high altitudes, the PO uses less oxygen to bind, therefore, less O2 is delivered to the tissues.3203

Yes, now, BPG concentration, it rises.3208

Once the BPG concentration starts to rise, the body is adjusting for the higher elevation by raising the concentration of BPG.3221

Higher concentration of BPG, it starts to bind to hemoglobin.3228

BPG starts to bind to the hemoglobin.3233

OK, as more binds, it causes hemoglobin to lose affinity for its oxygen/O2 but not so much at the pressure of the lungs, rather, very much at the pressure of the tissues.3242

OK, again, I have a certain amount of hemoglobin, 100% saturated, OK, 0, 100- that is like that.3301

It is going to deliver about 40% of its payload; it is going to keep 60%.3307

Well, now, I go to higher elevation; the BPG, it is true that it ends up dropping from about...the BPG causes the O2 to bind a little bit less in the lungs.3312

Now, I am about 90% saturated, but because it causes hemoglobin to lose affinity for oxygen, it actually causes more of it to release.3321

Even though I have dropped this to 90%, the 60, the 90 is 30, but now, it is no longer 60.3334

Now, it releases up to 50, 40 and even more.3341

If it releases now 50% instead of 60%, now, the difference between 50 and 90 is back up to 40%.3347

As far as the body is concerned, it is still getting the oxygen that it needs.3354

We have just gone to this much to this much; we have dropped the lower end.3360

It binds a little bit less, but it releases a whole lot more once it gets to the tissues.3365

I hope that make sense; OK, now, let’s go ahead and draw this out, and I think it will make sense.3371

Let me go to the next page here, and let me go ahead and draw the axis in black.3381

I have got this; I have got that.3388

OK, now, this is going to be 1; we are doing a binding curve here.3393

These are percentages, and this is going to be 0.3401

This is going to be 0; this is pressure.3406

OK, this is going to be partial pressure of O2, and we are going to go ahead and use kPa; and this right here, is the percentage of O2 actually bound.3412

I am going to go ahead and mark this as 4.3423

This is 8; this is 12.3425

This is 4kPa, 8kPa, 12kPa.3428

Now, you remember, this 4kPa, that is about the pressure at the tissues, and just about 12 or 13kPa, that is the pressure in the lungs at sea level.3432

At 4500 feet, the pressure in the lungs is about 7kPa; let’s go ahead and mark those points off.3446

I am going to mark those points in red; I am going to put a little X here, and I am going to put a little X here.3451

And now, let me go ahead and draw some lines.3458

Actually, let me go ahead and draw that; yes that is fine.3462

I will go ahead and draw the lines.3464

That is there; that is there, and that is there, and I will go ahead and label everything in just a minute.3469

Now, let me go ahead and draw my lines here.3481

This is going to be the normal binding curve at sea level, and remember, it is going to be, sort of, sigmoid.3486

Let me mark a couple of points, though, so just I know where I am.3492

Let me go ahead and go over this way.3499

OK, it is going to be something like that.3503

It is going to look something like that, the normal sigmoid binding curve.3510

OK, now, let me go ahead and do my labels here.3515

This point, this is the - I am going to do this in black - partial pressure of O2 in the tissues.3519

OK, partial pressure is 4kPa.3530

OK, now here, this is going to be the partial pressure of O2 at 4500 feet elevation.3534

Here, this is going to be the partial pressure of O2 at sea level- normal.3541

OK, that is what is going on.3550

Now, let me go ahead and draw the other cure before I discuss distances.3554

I am going to do this curve, I think in...that is fine.3558

I will go ahead and do it in blue also; that is not a problem.3562

Let’s make sure this is blue; now, this is going to be here, and it is going to be here- something like that.3567

OK, this graph, that is the graph with no BPG or normal BPG.3582

OK, this graph right here, is the binding curve for hemoglobin under conditions of elevated BPG concentration - OK - elevated BPG, or I will put extra bound BPG.3595

OK, now, let’s take a look at what happens; I am going to go ahead and do this in red.3623

Under normal conditions - OK, we are going to follow this top curve right here, OK - it is going to go ahead and - you know what, I am going to do it in black, I am sorry, OK - bind at about 12 or 13kPa.3627

It is going to bind to oxygen in the lungs; now, at the tissues, it is going to be right there.3644

The distance between here and here, this distance, that is the 40%.3651

OK, under normal conditions, that is what happens.3658

Now, let’s go ahead and go all of a sudden to 4500 feet, before any extra BPG has bound.3661

So, we are still going to follow the top curve.3667

Now, under conditions of 4500 feet, now, the pressure in the lungs is here.3672

It is going to bind that much oxygen, but still it is going to release that much.3676

Now, there is your 30%.3683

That is what your body experiences; from normal sea level, all of a sudden, if you jump up to a high altitude before the body has had the chance to adjust the BPG concentration, all of a sudden you are delivering less oxygen from 40% to 30%.3687

OK, now, after the body has had a chance to adjust, a couple of hours, a couple of days, BPG concentration in the blood goes up.3700

The BPG binds to the hemoglobin; now, the hemoglobin is going to behave differently.3710

It has less affinity for oxygen, so the binding curve is going to be different.3716

Now, we are going to follow this curve right here.3720

OK, now, at 4500 feet, it is going to bind that much.3724

It is going to bind less but not too much less, but now, at the pressure of the lungs, but now, as it makes its way to the tissues, now, let’s go to the tissues, come down this curve.3731

Now, look where we are; the binding in the lungs has diminished a little bit, but the release of the O2 in the tissues has increased a lot.3743

Now, this distance right here, that distance is back up to the 40%, in fact, more that 40% sometimes.3753

That is what is going on.3763

This is absolutely extraordinary; there is less oxygen available at higher elevations, and the body knows this.3768

It can only do what it can with what is available to it.3775

If less oxygen is available, the body adjusts at the lower end that says: OK, if I am going to keep 60% of my payload, now, instead I will keep 50% of my payload.3781

A certain amount has bound, but now, I am going to release more of it because, again, it wants to be at the 40%.3796

By releasing more, it brings it back down to the 40% despite the fact that there is less oxygen at higher elevation.3804

Again, it does not make too much of an adjustment at the high end; it makes the adjustment at the low end to bring the delivery back up to normal.3812

And again, that is all the tissues are concerned about; it is the tissues that want the oxygen.3820

That is where you notice it; you do not notice it in the lungs.3824

You notice it because less oxygen is being delivered to the tissues; well, if BPG ends up causing more oxygen to be delivered to the tissues, your body does not notice.3828

It is just going to function normally again as if it were at sea level; that is what is important.3837

OK, now, let’s go ahead and circle these points because these are the important ones- over here.3843

Here, the binding of O2 in the lungs at this higher elevation is only slightly diminished or decreased.3851

Here, the release of O2 in the tissues is greatly increased creating more of a gap to bring us back up to that 40%.3871

That is what is happening; OK, this top curve is normal hemoglobin under standard sea level conditions.3902

This curve is after BPG has had a chance to have its effect.3909

Now, the binding of oxygen has changed and by picking 2 points, we can actually see the numbers, from here to here, 30%, from here to here 40% under normal conditions.3915

Now, under BPG conditions - once the BPG has had a chance to do its magic - now, it binds less oxygen, but it releases so much more oxygen bringing the number back up to the 40% that the body is accustomed to.3930

That is all that is going on; now, I will finish off by saying, BPG binding, it does effect O2 affinity but not equally at both ends, in other words, the lung end and the tissue end or the lung end and the tissue end.3945

It affects it tremendously at the tissues end I will say.3993

The delivery of O2 is brought back to about 40%.4014

I hope that made sense.4030

Thank you for joining us here at; we will see you next time, bye-bye.4032