#08 Biochemistry Hemoglobin Lecture for Kevin Ahern's BB 450/550

Uploaded by oharow on 12.10.2011

Captioning provided by Disability Access Services
at Oregon State University.
Ahern: We are moving rapidly through stuff
and I finally sat down and looked at when the exam is
and the exam is this Friday,
so I guess...
[classroom chatter]
Ahern: It's not?
The exam is a week from Monday.
I'm guessing you guys wouldn't be ready for it this Friday?
There will be fewer things on it.
Think about that.
A smaller nice, little tidy exam.
Student: And the next will destroy us.
Ahern: And the next will kill you.
Or you will kill me, I don't know.
The second exam is not cumulative.
The final is cumulative.
I just have a couple fairly minor things
I want to say about characterizing proteins
and I do that in recognition of the fact
this is not a biophysics class.
The things that are there are really more biophysical in nature
than they are biochemical in nature.
I want you to be aware of what we can do
with certain techniques.
One of the techniques,
last time I talked about MALDI-TOF and MALDI-TOF
is a fantastic method as I say that I realize
I didn't post that figure for it.
I will get that figure posted showing you the set up
for a MALDI-TOF instrument
and you can hopefully see it better than my words can describe.
I will get that posted for you.
What I want to talk about today
are a couple other very powerful techniques
that come to us from biophysics.
I'm not going to go in depth
but you should know the basics of them
and you should be able to understand why they're useful for us.
The two techniques I want to talk about
are X-Ray crystallography and nuclear magnetic resonance.
You've probably had some exposure
at least with nuclear magnetic resonance,
I'm guessing in your organic chemistry class.
Has anyone had exposure to X-Ray crystallography before?
Little bit, okay.
I'll start with X-Ray crystallography.
X-Ray crystallography,
both these techniques, by the way,
are extraordinarily useful for helping us to understand
relative positions of nuclei in space.
I always tell the story when I give a tour of our facilities
in ALS in Biochemstry and biophysics to students
that using the tools of X-Ray crystallography
and nuclear magnetic resonance,
biophysicists can determine the position
in three dimensional space of every atom
that might exist in an enzyme that has 10,000 or 50,000 atoms.
That's a really remarkable thing
because knowledge about structure
leads to understanding about function.
We've heard structure function before.
When we think about how drugs get designed,
the design of drugs is happening increasingly
as a result of the molecular knowledge of the structure
of the proteins that they're targeting.
If I know the position or I know the structure
of the active site of the enzyme,
the place where the reaction is catalyzed,
I know the dimensions of the molecule
I need to design to plug up that enzyme.
That knowledge of structure is really valuable for us
to have for whatever purpose.
And there are purposes aside from designing drugs as well.
X-Ray crystallography arises from the fact
that X-rays get diffracted
which means they get bent when they encounter electron clouds.
That diffraction process is depicted here.
To do X-Ray crystallography,
one has to have first of all a crystal
and a crystal is a as you guys see crystals
you don't think at a molecular level what a crystal is,
but a crystal is a perfectly packed homogenous molecule
that has a regular repeat to it.
That is all the molecules in there are the same composition
and they're organized in a regular fashion.
That regular repeat is what gives rise to the crystal itself.
The process of making a crystal for a lot of X-ray crystal analyses
is actually the thing that takes the longest.
A lot of different, there's no one formula for making a crystal.
Different proteins crystallize in different to way.
Suffice to say that making the crystal,
which is already shown right here,
can be a very and time consuming step
and a frustrating step in the process.
Once one has a crystal,
one can take they crystal
and put in the path of an X-ray beam.
That X-ray beam will have its rays diffracted again
according to the electron clouds that it encounters.
The importance of the regularity of the molecules
in the crystal are very important
because those really add up
and give us the diffraction patterns that we see.
Now interpreting the diffraction patterns
obviously isn't a thing we're going to do here,
but sufficed to say that diffraction gives,
this is what a diffraction pattern
of a given crystal might look like and we say wow,
there are some spots.
These are the spots correspond places
where X-rays got diffracted to.
So a biophysicist can take information
from a diffraction pattern and work backwards,
ultimately to determine where the individual electron clouds
where it caused the diffraction pattern to exist.
So X-Ray crystallography is extraordinarily powerful
because it does give us three dimensional information about
the position and orientation of electron clouds
and a working map might look something like what you see right here.
Here are patterns of electron clouds and then within there
we decide what are the individual atoms that correspond to there,
you see different carbons and hydrogues and oxygens
and so fourth scattered through there.
And interpreting these patterns can again be a very time
consuming process it's very computationally intensive.
The result is at the end of this
that one has that structural information that's very useful.
Again, that's just a very cursory
description of X-Ray crystallography.
X-Ray crystallography has its advantages
and it has its disadvantages.
The advantage of X-Ray crystallography is if you get crystals,
then you can determine these positions very nicely.
Sometimes you can't always get crystals and that's one limitation.
And the other is crystals may or may not correspond to the natural,
whatever that means, shape of the solution.
So we think about the enzymes we have in our cells,
most of them are dissolved the in water of the cytoplasm.
So when I'm making a crystal,
I'm basically taking it out of the solution
so one thought is well is this reflecting
the actual structure it has when it's in the solution?
So to partly address some of those concerns,
this additional technology of nuclear magnetic resonance
is very useful because nuclear magnetic resonance analysis
allows one to determine molecular structures in aqueous solution.
They work in different ways and nuclear magnetic resonance
relies on the fact that certain nuclei have spins
that are characteristic of them and those spins
can be altered in the presence of an electromagnetic field.
So understanding the energies it takes to alter those spins
of given nuclei for example protons.
Protons are very commonly used in analysis
to understand the changes in those spin gives us
some knowledge about the structure.
I'll just show you a very brief example here.
Here's a nucleus that has a characteristic spin.
There are two possible spins that can exist.
One has a slightly higher energy than the other
and the difference between that energy
is what is excited using the electromagnetic radiation.
It turns out the different nuclei have different spins
corresponding to the electronic environment
in which they find themselves.
This depicts the nuclear magnetic resonance signal
of a very simple molecule.
This is ethanol.
We can see that ethanol has three different kinds of protons in it.
It has methyl protons that are farthest off in the end.
It has methylene protons in the middle here
and it has hydroxyl on the end.
These give rise to characteristic signals.
These signals have known positions,
we know where hydroxyl protons arise,
we know where methyl protons arise, etc.
And so we can examine the spectrum that comes from this.
The spectrum is called the chemical shift which I won't go in to.
This molecule has some hydroxyl,
it has methylene, it has methyl protons.
As you can imagine for a molecule like a protein,
it's not not nearly as this.
We might get very complicated spectra
and in fact we do get complicated spectra.
So this is a little bit more challenging to interpret.
We're not going to do that obviously here.
But sufficed to say that analysis of nuclear magnetic spectra
does allow, ultimately a scientist,
to allow which signal corresponds
to which groups inside the protein.
Now understanding the different kinds in this case of protons
that exist in a protein is useful information
but one of the things we're interested
in as biochemists is how do proteins fold.
Because remember that folding really gives the protein
its characteristic shapes so we would like to get more information
because the knowledge of different protons we have
in the protein really isn't sufficient enough to tell us structure.
One of the techniques done with that is an enhancement
as it were of nuclear magnetic analysis.
It's called the nuclear overhouser effect
and it arises from the fact that,
this clip doesn't work,
it arises from the fact that nuclei,
when they interact with each other,
also have effects.
In this case, here are two protons that,
as a result of folding,
have been brought into close proximity.
And if they're brought into close enough proximity
they actually do affect the signal of the other one.
This requires a very sophisticated analysis called 2D
and that's obviously a lot more complicated
than 2D gel electrophoresis I'm not going to go into that.
But sufficed to say that with this type of an analysis,
one generates some even more interesting spectra
but this information that we see here now
tells us not only what kinds of protons that we have
but how close those protons are to each other.
That's very, very useful when one goes to trying to determine
the overall structure of a protein molecule.
So that can be very, very useful.
Commonly these two techniques are used in combination
with each other to help elucidate molecular structure.
There is a biophysics course in 5 minutes.
How's that?
Questions or comments about that?
Let's get away from biophysics and talk about,
this is the lecture I'm going to give today
is most of the most popular lectures
I give throughout the entire term.
It's the lecture on hemoglobin.
And hemoglobin is, I hope to convince you
by the end of the lecture on Friday,
one of the most magical molecules in our body.
It is absolutely incredible the abilities
that are built in to the structure of this protein.
I start talking not about hemoglobin
but about a related protein called myoglobin
and I introduced myoglobin to you before
as a protein related to hemoglobin.
It's found in our muscle cells primarily
and there it serves the function of storing oxygen.
It's a very good way to store oxygen.
Hemoglobin is very good at delivering,
that is picking up and dropping off oxygen.
The difference you recall structurally I hope
between the two proteins and that myoglobin
has a single protein subunit.
And hemoglobin has four protein subunits.
So hemoglobin has quaternary structure, myoglobin does not.
And this quaternary structure that myoglobin has is,
that hemoglobin has is what gives rise
to all of the properties that the molecule has.
Well, you've seen myoglobin before,
it's mostly alpha helical structures,
it looks something like this.
There's the amino terminus and there's the carboxyl terminus.
And here you see alpha helix bend,
alpha helix bend, alpha helix bend,
a lot of alpha helices here.
We see the amino acids, we see 146 amino acids.
Myoglobin was I believe the first protein
whose structure of this nature was actually determined
and so that has some biochemical significance.
Not of any concern to us at the moment.
But the other concern for us
is it has an oxygen binding group in it called a heme.
So the heme, and yes, myoglobin has a heme
just as hemoglobin has a heme.
The heme is located right here.
Yes, sir?
Student: So wait, is this myoglobin or this hemoglobin?
Ahern: Actually, I'm sorry, this is the beta chain.
I have a link to it that says myoglobin.
They're very, very similar so this is the beta chain of hemoglobin.
We can think of this as myoglobin
because as I said the structure is very similar between the two.
Thanks for noticing that.
Anyway, both myoglobin and hemoglobin have a heme.
Hemoglobin you recall has four chains
to call beta and to call alpha.
This is one of the betas right here.
Now the heme turns out to be really important for several reasons.
The number one being of course that it's the place
where the oxygen is bound by this protein subunit.
There's the heme.
Heme is a flat ring.
It is something we refer to in these proteins as a prosthetic group.
Sounds like a very mouthful name.
Prosthetic group is simply a molecule
bound to a protein that helps the protein do what it does.
It's a non-amino acid.
So it's a non-amino acid bound to a protein
that helps the protein to function.
The heme group of hemoglobin and myoglobin,
the two are essentially identical and they're very,
very similar in structure to chlorophyll.
The electron gathering component of chlorophyll
that we find in plants.
The difference in plants that instead of having iron in middle,
we have a magnesium in the middle.
Student: You said this is planar?
It has like 20 carbons and stuff in the middle.
How is it possible?
Ahern: How is it possible?
Well, if we're talking about an exact plane,
there's nothing that's exactly flat.
Generally, it's a flat structure.
You can see when I talk to you about this
that the places actually pucker.
So it's planar, but I wouldn't say it's a perfect plane, no.
Alright, that puckering that we will see is very, very important.
It's actually seen right here in this figure.
What I'm getting ready to tell you about here
occurs in both myoglobin and hemoglobin
but the impact is felt in hemoglobin because of its four subunits.
What you see happening on the screen happens in both proteins.
Let me describe to you what's going on here.
If you we look at the deoxyhemoglobin on the left,
that's the way it normally sits.
Shannon says, "well that's not exactly planar."
And I say well okay, look.
It is slightly puckered.
We can imagine it being a little concaved downwards
like my hand is.
When the oxygen binds and we see oxygen bound over here,
there is a very, very tiny change.
So instead of being slightly puckered,
it flattens a bit.
Why does that happen?
It happens because the oxygen pulls up the iron atom.
The iron atom physically gets lifted.
This change is minuscule.
We're talking fractions of angstroms.
Very, very tiny change.
Yes, sir?
Ahern: His question is,
"Is this just the heme group?"
It turns out this movement affects a lot of things.
It's a very good question.
For the moment, we're thinking only about the iron atom.
The heme group itself is not moving.
It's the iron atom that's moving.
So the iron atom moves up a very tiny fraction of an angstrom.
And if you look at the structure,
you'll notice that the iron atom is not floating freely there.
It is in fact attached to an amino acid beneath it.
This amino acid that it's attached to is a histidine.
Now, if I pull up on iron and iron's attached to histidine,
you can do the math and figure that the histidine
is probably moving a fraction of an angstrom as well
and you'd be exactly right.
And you'd say histidine is attached
to another amino acid in the protein,
is it moving also?
Yep, so the foot bone,
the toe bone's connected to the foot bone,
and the foot bone,
this isn't going to be a song by the way.
The foot bone's connected to the ankle bone,
and the ankle bone is connected to the shin bone
and by pulling on the toe,
I'm ultimately going to affect the hip.
Even if it's by a very tiny amount.
And this very tiny amount,
I can't emphasize enough the importance of this very tiny change
because I'm going to hopefully convince you by the end
that the result of this very tiny amount of movement
allows us to be animals.
Without this movement, animal life is essentially not possible.
This is a scary thought.
Why is it that this makes animal life possible?
We'll talk about that.
Hemoglobin of course doesn't exist as a single subunit,
it exists as 4 subunits.
All 4 subunits have a,
each subunit of the 4 has a heme group of its own.
So when this guy binds an oxygen,
and by the way, because it's a schematic,
they're not showing the connection,
but in each case it's connected to a histidine.
When this guy binds to an oxygen,
let's see I've got this hemoglobin that's got no oxygen
This guy binds an oxygen,
it's going to cause the iron atom to move a slight distance,
it's going to cause that histidine to move a slight distance.
It's going to cause that entire chain to very slightly shift.
That very slight shift changes the overall shape of this subunit.
And guess what?
That shift affects how it interacts with its adjecent subunits.
And the adjacent subunit now
becomes more favorable for binding oxygen.
So this one, when we have hemoglobin that's empty of oxygen,
it's not real keen on binding oxygen,
but once one of them binds oxygen,
these changes get communicated between the subunits
and each additional oxygen is increasingly favored for binding.
This phenomena I've just described to you is called
The bonding of one molecule to a protein
affecting the binding of others.
In this case,
it's positive cooperativity.
It's favoring more binding.
Now this is really important.
We have to, we are animals,
we are moving creatures,
we have to have an adequate oxygen supply.
Plants don't have this issue.
Plants don't have to get up and run around and jump
and go chase things or run away from things.
Their oxygen needs are more constant.
Ours are rapidly changing.
We need oxygen, we need it now.
When our hemoglobin gets to our lungs,
it doesn't have an awful lot of time to be there.
We want it to load up on oxygen as much as it can
and take that oxygen out to the tissues where its needed.
Cooperativity as we will see,
plays a very big role in the loading up of hemoglobin.
So we're loading up hemoglobin.
If we can't load up hemoglobin,
we don't have enough oxygen,
we can't go run, we can't go escape,
we can't do things that animals do.
Very, very important.
The other thing I want you to look at in this structure,
it's actually easier to see right here
is that when we look at hemoglobin from above as we are in this case,
we see that hemoglobin is shaped sort of like a doughnut
and there's a little hole in the middle.
That little hole turns out to be extremely important.
Extremely important.
So I'm going to talk back and talk about that hole
but before I do that I want to tell you a little bit
about the needs of oxygen in the cell
and how hemoglobin helps to supply those.
Questions on this before I move forward?
Everybody understands what cooperativity is?
Yes, sir?
Student: Does the inverse occur when it unbinds the oxygen?
Ahern: Good question.
Does the inverse occur when it unbinds oxygen?
The answer is to some extent, yes it does.
Loss of one will favor the loss of additional ones.
So it would be a negative cooperative.
So you start to see where this is heading, right?
If we look at the oxygen binding of myoglobin, this is a plot.
Again, whenever I show you a plot,
I always want you to know what the axes tell us
because without the axes,
the plot has no meaning.
This is the fractional saturation meaning
what fraction of all the myoglobin molecules
in the solution have an oxygen bound to them?
Myoglobin can only bind one oxygen per protein
because there's only one subunit and each heme only binds one oxygen.
Either it's bound or it's not bound.
What percentage of those guys are bound with oxygen?
What we see that it takes very little oxygen.
This is very low, the pressure of oxygen on the X-axis.
Very little oxygen for us to get 50% saturated.
What does that mean?
It means that myoglobin when it has the chance
is grabbing a hold of oxygen.
It's very good at storing oxygen.
It grabs it, it holds onto it very well.
Well that's nice,
but it's not ideal for delivering oxygen
because if myoglobin didn't give up its oxygen
til the oxygen concentration got very low,
it could travel all the way through the body,
get all the way to the lungs and it hasn't given up its oxygen.
"No, it's mine.
I'm the big kid I get the quarter."
"I'm not going to give this up for you."
Myoglobin only gives up its oxygen
when the oxygen concentration gets very low.
How many people have UPS on their computer?
Anybody know what a UPS is?
It's an uninterruptible power supply.
It's there to give you power when the power goes off
so you have a chance to shut down and save your work.
Myoglobin is the UPS for your muscles.
When you're working very hard,
it's very easy for you to use oxygen
faster than your blood can deliver it.
Well oxygen is important.
It's not essential,
but it's important so that more oxygen your muscles have,
the better off they are because muscles need it for contracting,
we gotta run away from something,
we gotta beat something up, we gotta do whatever,
hopefully we're not doing too much of that.
If hemoglobin can't supply all the oxygen that's needed,
we want something there to back it up and this is backing it up.
When the oxygen concentration starts getting very low,
myoglobin says, "Oh, here's some oxygen."
That's the only time myoglobin gives up oxygen.
When the concentration gets very, very low.
But it helps us when we need that.
Let's compare that with hemoglobin.
Ahern: That's the oxygen concentration.
Oh, here?
How much does it take to get half of it saturated,
half of it bound to oxygen.
Is P 1/2 just refers to 50% of it.
So very, very low number there.
If we compare this with hemoglobin,
hemoglobin has a different looking curve.
So the curve that corresponds to myoglobin
is what we call hyperbolic.
It's a hyperbolic curve.
It's a hyperbolic function that will fit that curve.
The curve that hemoglobin gives is called sigmoidal
because it has sort of a S shape.
It's sigmoidal.
Look at this.
At low oxygen concentrations,
there's not very much oxygen bound.
When hemoglobin travels through our body,
it goes from places of high oxygen concentration,
our lungs, high oxygen concentrations,
essentially 100% of it gets bound with oxygen.
As it travels through the body,
the oxygen concentration starts dropping
because the cells are using oxygen and hemoglobin is the only source,
oxygen concentration drops
and hemoglobin starts letting go of its oxygen.
It's a perfect system for delivery.
We see it cooperatively binding oxygen to begin with
and we see cooperatively letting go of oxygen.
Binding in a negative sense
when the oxygen concentration starts to fall.
Hemoglobin, because of cooperativity can satisfy an immense,
or a diverse set of oxygen concentrations
as they occur in our bodies.
This is essential for an animal.
I just cannot, I keep coming back to that,
but I can't emphasize that enough.
It binds like myoglobin at the very highest concentrations.
It will get 100% bound.
But as oxygen concentration falls,
it's down over here.
And that's pretty cool.
Then by the time it's dumped its oxygen,
it goes back to the lungs and it gets more oxygen.
So let's think about that a little bit.
No, I'm not going to ask you to draw this particular figure
although you should be familiar with any of these figures.
We see the differences in oxygen concentration,
this figure is nice in that it shows us the concentration
of oxygen roughly that occurs in tissues
and the concentration of oxygen that roughly occurs in lungs.
We see that again, way up here in the concentration of the lungs,
myoglobin and hemoglobin are essentially the same.
And then way out over here,
when this thing gets out to the tissues,
only what is 38%, no, I'm sorry, what is this,
about 30% or so of the hemoglobin is actually still bound to oxygen.
So that flexibility of hemoglobin for oxygen is very,
very valuable for us and allows us to things like sitting here,
or getting up and running if we have to go running.
And both of those work.
Okay, if I exercise, blah blah blah, if I rest,
I have different needs, if I have lungs, same sort of thing.
This shows us the quaternary changes
that happen as a result of oxygen binding to hemoglobin.
Quaternary changes mean the four subunits
are actually changing on oxygen binding.
Notice that doughnut hole that I had before.
The doughnut hole has largely closed up.
It turns out that these two different states of hemoglobin
have names that we give.
And we're going to hear more about these names later.
They're called the R state and the T state.
I need to define them for you.
The one on the left is called the T state.
The T stands for tight.
I like to think about it as people I know who are uptight.
People you know who are uptight,
you can just sense it around them and they can't take anything more.
They're very rigid.
Give me some oxygen,
"No, I don't want any oxygen!"
Okay, tight structures.
Ahern: What's that?
Student: [inaudible] the right?
Or the left?
Ahern: The left?
Yeah, that's tight, yeah.
They're not very flexible.
They have poor binding of oxygen.
So when hemoglobin has no oxygen, it's in the T state.
It doesn't want to bind more oxygen.
On the other hand,
once it's bound and it's gotten full of oxygen,
its structure changes to what we call the R state.
R is the relaxed state.
The relaxed state, "yeah, come on, we'll take this, we'll have it,
I can take a lot of stuff, man."
[class laughing]
I grew up in the 60s, guys,
you gotta give me credit for the language at least.
So the relaxed state is a high affinity binding for oxygen.
Once we put one oxygen on there,
we start flipping it into the R state
and we've got affinity to find more oxygen.
We flip it into the T state,
it doesn't want to bind oxygen.
That's kinda good.
If you think about it.
Let's imagine that I'm a hemoglobin
that's floating around and I've just gone to let's say
the muscles where they're exercising pretty heavily
and I dump all my oxygen.
On the way back to the heart or to the lungs,
I pass through the kidney.
Do I want the hemoglobin taking the oxygen away from the kidney?
That wouldn't be a good career move, right?
So I only want it to flip when there's a high oxygen concentration.
That's what's going to happen when it gets back to the lungs.
So the T state and R state
really serve the body's needs very, very usefully.
There are a couple of ways of describing
how this phenomena occurs.
The way I've described to you is called a sequential mains.
That is the binding of the first one affects the second one,
affects the third one, affects the fourth one.
And that's not shown on the screen,
this is a different model.
If I were to show what happens,
we've got T state above and R state below.
In the sequential model, one of these guys turns circular,
it favors the next one turning circular,
the next one turning circular, etc.
That model's called a sequential model of binding.
Changes in the structure of one changes the next one,
which changes the next one, etc.
It's sequential.
This model you see on the screen is the opposite of the sequential,
it's called the concerted model.
These are models.
Models are way of explaining things.
This model says that we don't see one followed by the other,
followed by the other, followed by the other.
Instead, what we see is we're either in one state
or we're in the other state
and binding of things locks them into that state.
Now hemoglobin is not a good model for this.
We'll see in next week's lecture
how an enzyme is a much better model for this.
This model and I'll say more about this model next week
so I'm not going to go into much here,
but this model says that the changes happen all at once
and they're independent of the binding of anything.
We see this as back and fourth.
But they get locked into one vs. the other based on what they bind.
We'll come back to that next time.
For right now, think sequential.
Binding the first changes the second,
changes the third, changes the fourth.
Changing the T to R state changes significantly
the binding affinity of hemoglobin for oxygen,
which I showed you before
and if we look at what hemoglobin would look like in the T state,
this is what it would look like.
If it were only in the R state,
this is what it would look like and in fact,
hemoglobin goes through a transition from T to R
and that's what we're seeing,
why this curve has a couple of shapes in it.
We're seeing a change from T to R.
That change is what we've already described as cooperativity
and that cooperativity is favoring bonding in this case is oxygen.
If we're getting more oxygen or favoring the release of oxygen
if we're getting into lower concentrations of oxygen.
This is what a sequential looks like.
Nothing bound, first one changes this one,
which causes this one to chances,
which bind has caused this one to change,
which caused this one to change its find.
It doesn't matter if it binds to an alpha subunit
or a beta subunit. It doesn't matter.
They're essentially the same as far as this molecule exists.
Yes sir?
Student: Shouldn't the [inaudible] under K4 be switched
where there's a higher affinity to drive further to the right?
Ahern: All of these depend on oxygen concentrations,
so you're exactly right.
In the concentration of the lungs,
even though you've got a lower going to the right,
there's enough oxygen concentration to drive that.
Student: You're showing a sequential increase in K1
and [inaudible] the last one is shorter.
It's counter-intuitive.
Ahern: No, because it doesn't want to bind that first one.
I agree that this is important for the releasing of oxygen.
This, from what I've told you sounds a little odd
in terms of putting that last one on there.
But the reason this is the case
is because that first one doesn't want to bind.
And that's because this guy here is in the T state.
That should answer your question.
Student: Once you have the three on there,
it seems like it should be more of a push
in the equilibrium to push [inaudible].
Ahern: Right, so his point is that this equilibrium
is favored actually in the leftward direction
and that would be true if it weren't
in the high oxygen concentration in the lungs.
The lungs are loaded with oxygen and that drives it to this state.
We want this guy to dump off oxygen once it gets out of the lungs.
That's why the arrow is back to the left.
Where am I at here?
That's the basics of hemoglobin
but there's so much more that's built into this molecule.
The first one I'm going to show you right here
is a really interesting and cool molecule called 2,3-BPG.
We'll talk more about this molecule later in this term
when I talk about glycolysis
but this molecule turns out to be a fascinating molecule.
You don't need to know the structure
but you definitely need to know
at least this part of the name: 2,3-BPG.
It's real name is 2,3-bisphosphoglycerate.
I need to tell you why this molecule is important.
If my microphone works that is.
Why this molecule is important.
This molecule is a molecule
that is released by rapidly respiring cells.
If I'm a muscle cell and I'm doing my business I'm making 2,3-BPG,
we'll see later it's actually a byproduct,
but that doesn't matter for our purposes right now.
Actively respiring cells release 2,3-BPG.
So my muscle cells may have a lot of 2,3-BPG,
my nose cells may not have so many.
With me?
Unless I'm sneezing,
I've got that cold everybody else does,
I might have more 2,3-BPG.
It turns out that 2,3-BPG affects hemoglobin.
If we look at hemoglobin in the presence of 2,3-BPG and red,
we see that it binds less,
and they're not exaggerating the S so much here,
they've sort of drawn this to make their point.
The point is that in the presence of 2,3-BPG,
hemoglobin holds onto less oxygen.
So 2,3-BPG it turns out causes the hemoglobin to release oxygen.
How does it do it?
It's very simple.
2,3-BPG has a shape that fits exactly in that doughnut.
It fits exactly in that doughnut
and when it fits into the doughnut,
it favors the conversion of hemoglobin
from the R state to the T state.
T state has low affinity for oxygen,
guess what hemoglobin's going to do when 2,3-BPG binds to it,
it's going to start giving up more oxygen
and that's exactly what this curve is telling us.
This turns out from a bodily perspective
to be very useful because when I've got
actively respiring tissue and I've got a lot of 2,3-BPG,
what's 2,3-BPG going to do?
It's going to bind hemoglobin and hemoglobin's going to say,
"Okay, flip into the T state,
I'm going to let go of the oxygen."
And as concequence of letting go of oxygen
the tissues that need the oxygen get it.
That's great.
But wait, there's more.
But wait.
If hemoglobin has bound to 2,3-BPG,
it's going to be in the T state and when it gets back to the lungs,
it's still got 2,3-BPG,
it doesn't want to bind to more oxygen, I've got trouble.
Well fortunately, remember these are not covalent bindings,
fortunately 2,3-BPG fits in that pocket.
But it goes in, comes out, goes in, comes out.
Like any binding that occurs,
binding and letting go happens all the time.
On the way back to the lungs, 2,3-BPG,
when it gets off of the hemoglobin
can get grabbed by cells and be metabolized.
So as hemoglobin is making its way back to cells in most people,
the cells are grabbing it, burning it up,
and hemoglobin gets back to the lungs and
it has no 2,3-BPG in it.
If it had 2,3-BPG in it, you wouldn't bind as much oxygen.
Now all of you pre-meds,
everybody looks up at this point,
smokers are full of 2,3-BPG.
Smokers are full of 2,3-BPG.
The reason that, one of the reasons that smokers huff and puff
going up stairs is that 2,3-BPG
doesn't get all the way broken down.
The hemoglobin gets back to the lungs, uh oh.
My oxygen carrying capacity is lower,
that's why smokers huff and puff going up stairs.
They've got too much 2,3-BPG.
The next question is why do they have more?
And that we will save and talk about when we talk about glycolysis.
Suffice to say,
they have much more 2,3-BPG in their blood than do non-smokers.
Yes sir?
Student: So is that a large contributory factor to say COPD?
Ahern: COPD being?
Student: Chronic obstructive pulmonary disease.
Ahern: Is it a contributor to COPD?
Not to my knowledge.
There are other things that give rise to that.
I'm not a medical person so I can't tell you that.
But sufficed to say that the primary physical observation
that you could make with respect to hemoglobin,
respect to BPG in smokers is that they huff and puff.
They puff and then they really huff and puff.
If you smoke, quit doing it.
Now you know at the molecular basis
why smokers are having a hard time going up stairs.
Their hemoglobin is stuck in the T state
and they can't get it out of that T state very well.
There we go.
There's your doughnut and there's the binding.
We'll need to worry about the various other stuff that's here.
Now, hemoglobin is some pretty cool stuff.
There's a problem, though.
We are not chickens.
And some would say that's probably good.
But chickens lay eggs and the fetus
that develops inside the egg has its own resources
and doesn't have to rely on mom except to the point
where the egg is laid.
Mom, however, is the source of nutrients in mammals
for food, for water, and for oxygen.
Now there's a problem.
The problem is what if mom's hemoglobin
is competing with the baby's hemoglobin?
They both have oxygen,
why should the baby's hemoglobin have to fight with mom for that?
What turns out that fetuses have a modified hemoglobin.
They have a different hemoglobin than adults do.
They have something called fetal hemoglobin.
Adults have alpha-2 beta-2,
meaning we have two alpha subunits,
two beta subunits and that makes up four subunits which you saw.
A fetus on the other hand has two alpha sub units
and twp slightly different gamma sub units.
So you've got alpha-2, gamma-2.
Those two gamma subunits give the hemoglobin of a fetus a very,
I shouldn't say very,
but a slightly different property than mom's.
Do they have cooperativity?
But they have a greater affinity for oxygen than mom's hemoglobin.
They can literally take oxygen away from mom.
Talk about a little parasite.
[class laughing]
A little parasite sitting there sucking my oxygen, right?
How do they do it?
The gamma subunits in addition to having a slightly
different structure cause the hemoglobin
that they're in to not have a doughnut.
That little doughnut hole where the 2,3-BPG fits
doesn't fit 2,3-BPG anymore.
The fetal hemoglobin essentially stays in the R state all the time.
Essentially stays in the R state all the time.
Now, you say that's great,
so obviously it can take oxygen away from mom
and yes, it can, and yes, it does.
But we just saw how the T state helped to release oxygen, right?
Is the fetus starved for oxygen?
What do you think?
Okay, there's a no, there's a no.
Nobody thinks yes?
No, it's not.
Why is it not starved for oxygen?
Student: Higher net oxygenation level?
Ahern: Higher net oxygenation level.
It does have a higher net oxygenation level,
but it also has trouble releasing oxygen so the answer is no.
The answer is simpler than you think.
Student: I mean it just sits there, right?
Ahern: Okay, and that's the answer.
It just sits there.
It doesn't have widely varying oxygen needs.
Mom goes and climbs the stairs,
she needs more oxygen than when she's sitting around in a chair.
All the fetus does is kick.
[Class laughing]
So it doesn't have widely varying oxygen needs.
It needs a relatively constant supply of oxygen
and because it does have a high oxygen
carrying capacity as you noted,
there's enough release so that it can satisfy those needs.
If it had very diverse and very challenging needs,
then you betcha there would be an issue.
Other questions?
I'm going through this kinda quickly.
Student: So if you're like a mom
and maybe you have a very low blood concentration,
like iron concentration,
would it be smart to take supplements for that?
Ahern: If you were a mom and you had anemia,
is that what you're saying?
Student: Yeah.
Ahern: Would it be smart to take, I don't know,
iron supplements or something like that?
Yeah, for people that are anemic,
that can be a consideration with mom but again,
I'm not a physician but I'd imagine that yes,
they would use that.
Student: So when the fetus is born, it becomes...[inaudible]
Ahern: Yeah, yeah.
Ahern: Yeah, when the fetus is born,
it has got fetal hemoglobin.
So that change over happens in the first year or two
where the gamma sub units stop being made
and the beta subunits start being made.
And so the fetus transitions to adult hemoglobin
fairly early in its life.
But you're exactly right, yeah.
Student: Does it change based on how active the baby [inaudible]?
Ahern: Does it change based on how active the baby is?
I honestly don't know the answer to that question.
I don't know.
As far as I know, it's simply a developmental thing.
Your question is whether it responds to environment
and I don't know the answer to that.
Well we've gone through a lot I thought
we should finish with a song today.
What do you guys think?
We haven't done a song in awhile.
I have a cold so I think it will be worse than usual
so I want you to sing really loud today.
And by the way, I have an idea I will do with my classes.
If you guys sing loud,
I assure you you'll have an extra credit question on the exam.
But if I can't hear you...
Everybody ready?
Okay, everybody sing.
"Biochemistry, biochemistry,
"I wish that I were wiser.
"I feel I'm in way over my head.
"I need a new advisor.
"My courses really shook me.
"Such metabolic misery, biochemistry, biochemistry.
"I wish that I were wiser.
"Biochemistry, biochemistry, reactions make me shiver.
"They're in my heart and in my lungs."
"They're even in my liver.
"I promise I will not complain.
"If I could store them in my brain.
"Biochemistry, biochemistry, I wish that I were wiser.
"Biochemistry, biochemistry, I truly am in a panic.
"The mechanisms murder me.
"I should've learned organic.
"For all I have to memorize, I outta win a Nobel prize.
"Biochemistry, biochemistry, I wish that I were wiser."
Alright, guys.
See you Friday.