Biochemistry Protein Function III: More on Hemoglobin

Hello and welcome back to Educator.com, 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 in...no, 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