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Kevin Ahern: How's everybody doing today?
Student: Wunderbar.
Kevin Ahern: How was the exam?
Student: Good.
Student: It was fine.
Kevin Ahern: Did I hear, "Good," "Good," "Good"?
Is that what I heard?
Wow!
All right.
Student: We'll see if numbers bear that out.
Kevin Ahern: We'll see if numbers bear that out.
Comments or questions or... ?
Student: Good extra credit question.
Kevin Ahern: You liked the extra credit question?
[laughs]
I thought you might like the extra credit question.
No comments on the exam?
Student: When are we getting it back?
Kevin Ahern: The exam, as I said last time,
will not be available until Monday.
I apologize, but the TAs are just too busy with exams
this week, themselves, to do that.
I fully intend to make it available Monday morning first thing,
and I have told the TAs that I expect that's how they're going
to spend their Thanksgiving break,
so not everybody has it as bad as they do, I guess.
So you will have it back on Monday.
I will put a note out when it's available, as I always do,
but it will be available in the BB office, as before.
Student: So extra points if it comes back
with cranberry sauce on it?
Kevin Ahern: Extra points gets what?
Student: If it comes back with cranberry sauce all over it.
Kevin Ahern: I may see cranberry sauce on
the turn-ins, I don't know.
You guys might see them as you pick them up.
Actually, that reminds me.
I always invite my classes over to my house,
so if you guys are in town on Thursday and you would like to
come over for turkey, seriously, give me a holler.
I'd be happy to have you over.
So if you're not going home and you have no other plans,
give me a holler and I'll try not
to food poison you with turkey or something.
[laughter]
I'm serious.
I'm serious.
I usually get one or two people who take me up on it.
We have usually a big get-together of students
and faculty at our house, so that'd be kind of fun.
Student: Thank you.
Kevin Ahern: Well, I guess without any comments for exams,
we'll move forward.
We are nearing the end, believe it or not.
After today, there are only four lectures left in the term.
Kevin Ahern: Where did our term go, right?
It's hard to believe.
Yeah!
After today there's... are you upset?
Student: Yes!
Kevin Ahern: We need some counseling on
the front row here, I think.
[laughter]
Try not to take it too hard, but I can promise you
we'll have more of it next term.
I guess we're going to have class in here next term, as well.
We used to have it in Gilfillan but they decided to give us
this classroom again, so we'll have to bear with it.
I like this classroom, though, now.
What's that?
Kevin Ahern: 451, yes.
I've had a couple of questions...
451 does not have a recitation.
"Where's the recitation?"
“Where's the recitation?"
There's no recitation in 451 because we don't work
through problems like we do in 450,
so there's no calculations, as such, in 451.
The first half of 451 is that of metabolism,
the kind of things that we're doing right now,
and the last half is on molecular biology—DNA synthesis,
RNA synthesis, protein synthesis, gene expression
and one brief thing on sensory, the senses,
and one on the immune system.
So that's what's there.
The last thing I'll say is, I will tell you that for
the final exam I allow you to have a note card.
I'm kind of picky on that note card.
You have to get the note card from me
and you have to turn it in with your final exam.
That's the two rules about the note card.
Even if you don't want a note card,
you have to turn in a note card that you get from me.
So not having a note card is going to cost you points.
Having a note card besides the one that I give you
is going to cost you points, as well.
So make sure you get a note card from me.
I'll make those available next week.
It's a fairly large note card, and, yes,
you can use both sides and so on and so forth,
the primary rule being that everything on the note card
must be in your own handwriting.
You cannot paste figures on there.
You cannot print on there.
You have to use it as it is.
I had to institute that rule, I've told that story
to a few of you, I think, but I had to institute the rule
a few years ago when I had a young man who had
the really brilliant idea that if he printed the note card
in red and then printed over it in green,
that if he used red-green glasses he would
double the capacity of his card.
And it worked!
[laughter]
It worked!
Student: Wow.
Student: Oh, my gosh.
Kevin Ahern: Yeah, It was a clever idea.
I said, well, maybe that's just taking it a little too far.
I honestly think there's some benefit from writing
things with your own hand, anyway.
So that's the rule.
It has to be in your own handwriting.
No printed cards.
No copy and paste.
No figures pasted on there.
The card has to be as it is.
I'll say more about that when I hand the cards out on Monday.
The final exam in here is on Monday at 9:30 a.m.
It's the first day and it's one of the first finals.
I've never had that happen with this class before,
so we'll actually have that final done with fairly early.
So that's good news, maybe bad news.
I don't know.
Last time I talked about... shh!
People are doing a lot of talking.
When you're talking, it's hard for people around you to hear.
Last time I talked about my own, Kevin Ahern's,
pet theory about why Americans are getting obese,
and I hope that I made a reasonable case for you
about why that's the case.
The next thing I'm going to talk about is something
that's going to reinforce something else I've been saying
during the term and probably you never thought
about it that way, either.
I've been telling you all along that glucose is a poison,
and, in general, sugars are poisons for cells.
In general, sugars are poisons for cells.
This next example I'm going to give you,
you're going to see how it actually,
this poison can manifest itself.
Before I tell you about that, though,
I have to tell you a little bit about how galactose
is normally metabolized.
Galactose is a sugar.
It's a monosaccharide.
It's very closely related to glucose.
It's actually an epimer of glucose, and that epimer
of glucose we get in our body by drinking dairy products.
Galactose, as I maintained previously,
is half of the disaccharide known as lactose,
the other half being glucose.
Our body has to deal with galactose because, like glucose,
galactose is a poison and if we don't deal with it,
we've got problems.
Well, in our body, we deal with glucose being
a poison by making glycogen.
Question?
Student: You said lactose was an epimer of glucose.
Kevin Ahern: I'm sorry. If I said "lactose," I meant "galactose"
is an epimer of glucose.
Galactose is an epimer of glucose.
Galactose we have to deal with,
but we don't make polymers of galactose.
That's one of the things we don't do.
So let's look to see, first,
how galactose is normally metabolized in our bodies.
Galactose is first converted from the free sugar
form to galactose 1-phosphate by this enzyme
known as galactokinase.
Galactokinase catalyzes the reaction here.
It's rather similar to the reaction
that you saw with hexokinase, the difference being that
it's working with a galactose and it's putting the phosphate
on position 1 instead of putting it on position 6,
but pretty much everything else is the same.
So this is the first step that we have
in detoxifying galactose.
It would be nice to be able to use galactose for energy
and so forth, and it turns out that I mentioned last time
that glycolysis is a very useful pathway because
it allows us to metabolize many sugars.
One of the ways that that happens is that these sugars
can get converted into fructose or, more commonly,
into glucose, and that's what we see happening on this figure.
Now, I'm going to step you through this figure
and try to hopefully ease some of the confusion or concerns
that students have about what's happening in this process.
It's not nearly as complicated as it looks.
How many times have I told you that, this term?
You're never going to believe me again, right?
Here's our product of the last reaction, galactose 1-phosphate.
Galactose 1-phosphate plus UDP-glucose—so that's
just glucose linked to a UDP and we'll see that molecule
is important in glycogen metabolism later
but if I take these two and I combine them with
this enzyme—whose name I'm not even going to mention,
just simply because it's not really important
for our purposes—what happens?
Well I see that the glucose gets released,
as glucose 1-phosphate, and the galactose becomes
linked to the UDP.
So instead of having UDP-glucose,
I have UDP-galactose and I have some glucose 1-phosphate.
Everybody see what's happened there, so far?
We've just swapped the galactose 1-phosphate for
a glucose 1-phosphate and we're left with UDP-galactose.
Now, glucose 1-phosphate, as we will talk about next week,
is readily convert into glucose 6-phosphate.
There's an enzyme that's known as phosphoglucomutase
that will convert glucose 1-phosphate into glucose
6-phosphate, and, of course,
you know glucose 6-phosphate can get burned in glycolysis.
So already we see how this pathway is contributing
things that can be used in glycolysis.
But, we're not done yet because we have to convert
UDP-galactose into something useful.
All we've done, so far, is just put the galactose onto a UDP.
That comes up with the next reaction here,
UDP-galactose 4-epimerase.
Well, that last enzyme name should tell you
something about what's going on here.
An epimerase is going to make an epimer,
and galactose is an epimer of glucose and guess what it does?
It converts UDP-galactose into UDP-glucose.
So, now, we're right back where we started.
So, in essence, every time this wheel turns
we're seeing a little wheel turning here, every time
this wheel turns, a glucose 1-phosphate is kicked out.
In essence, we're bringing in galactose and
we're kicking out glucose... galactose 1-phosphate,
kicking out glucose 1-phosphate.
All right?
So this pathway that you see on the screen is allowing us
to metabolize galactose in glycolysis.
Now, there's always confusion as to what's happening here,
so I'll stop and take questions on that.
Yes, Connie?
Student: Could you go over how glucose 1-phosphate
is used in glycolysis again?
Kevin Ahern: Glucose 1-phosphate is not used in glycolysis,
but glucose 1-phosphate can be converted readily
into glucose 6-phosphate that is used in glycolysis.
Kevin Ahern: We'll talk about the enzyme
that does that next week.
It's involved in glycogen metabolism, actually.
But this is only one step away from glycolysis, basically.
Yes, sir?
Student: Is there a beta form of glucose 1-phosphate?
Kevin Ahern: Is there a beta form of glucose 1-phosphate?
There probably is, but the product here is an alpha.
Enzymes are always specific for what they will make.
So, in this case, you're only going to get
the alpha out of this guy.
That's it?
Was I that clear or were guys that much asleep?
Yes, sir?
Student: Just [unintelligible]
would it be glucose 1-phosphate [unintelligible]
would it be a mutase, to change that from 1 to 6?
Kevin Ahern: Yeah.
The enzyme that will convert that from 1 to 6 is a mutase,
and again, I'll talk about that next week.
Kevin Ahern: It's a good question,
but it is actually a mutase that does that.
When I say "mutase," what comes to mind?
What's the intermediate?
Student: [unintelligible]
Kevin Ahern: Glucose 1,6-bisphosphate, right?
[laughing]
Not 2,3-bisphosphoglycerate.
Not every intermediate is 2,3-BPG.
Shannon?
Student: So I think, I don't know if you said this before,
but what's the difference between a mutase and isomerase?
Kevin Ahern: What's the difference between a mutase and isomerase?
Isomerase simply does the rearrangement
by moving one piece to another piece.
A mutase has an intermediate where we add and then we subtract.
In the case of 2,3-BPG, we started with 3-PG,
then we had 2,3-BPG, and then we took off
the phosphate to make 2-PG.
In this case, you guys are getting ahead of me,
but since you've asked the question, I'll answer it,
in this case, we have glucose 1-phosphate.
The mutase puts a phosphate on,
so we have glucose 1,6-bisphosphate,
and then it takes off position 1 and we're left
with glucose 6-phosphate.
So the mutase, that name "mutase" will always
tell you it's putting both of them on
in the process of doing what it does.
Okay?
So, this is a very useful pathway.
It's a very important pathway for us because it allows us
to metabolize galactose if we are, like most of us are,
fairly, relatively enriched, our diets
are relatively enriched in dairy products.
We're getting plenty of lactose and if we don't
have a way of metabolizing that galactose,
we've got a problem, and that comes up next.
So if we have, for example, a genetic problem
where we're lacking either of these enzymes,
galactose 1-phosphate uridyl transferase,
whose name I don't expect you to know,
or UDP-galactose 4-epimerase, if we're lacking either one
of these enzymes, we can't do this cycle.
Well, if we can't do this cycle, what's going to happen?
Galactose 1-phosphate is going to accumulate,
and when galactose 1-phosphate accumulates,
then, as this accumulates, so, too, is free galactose
going to accumulate.
Now, free galactose is the problem, as I said.
It's a poison.
When it accumulates, one of the problems
that we experience is a product of this reaction.
Our body recognizes, or our cells recognize,
that galactose is a poison so it does something
to convert it into something that's less poisonous.
What it does is it reduces the aldehyde
to an alcohol to make galactitol.
If you recall, aldehydes are fairly reactive,
so this is a way of making this molecule much less reactive.
It's a protective mechanism.
Unfortunately, certain places in our body,
this guy is a problem.
It turns out that in the lens of our eye
this will form crystals.
Galactitol will form crystals in the lens of our eye,
and can lead to the formation of cataracts.
It's not the only cause of cataracts,
but it's a potential cause of cataracts.
So if you're not metabolizing galactose properly,
you make this compound here that becomes a crystal
in the lens of your eye.
So, again, reinforcing the poison nature of galactose.
Yes?
Student: Are people who are lactose intolerant...
Kevin Ahern: Yes.
Student: ... lacking these enzymes?
Kevin Ahern: Are people who are lactose
intolerant lacking these enzymes?
It turns out, no.
So you're anticipating my next thing,
which I'll talk about in just a second,
but lactose intolerance involves something else
and other problems.
Student: So do people with diabetes get cataracts?
Kevin Ahern: Do people with diabetes get cataracts?
People with diabetes get a lot of things,
but I'm not aware of them getting cataracts
at any higher level than anybody else, no.
I always tell this story at this point, actually,
because I watched young people do some things
that are really stupid.
As my capacity as advisor, or as instructor,
is I see people do things that are stupid.
There's one that people do that they don't realize
that it's stupid, and I'll tell you about it because,
when I was your age, I did the same thing.
How many of you have ever popped something in the microwave
and gone and just watched that little
puppy cook in there, right?
Have you done this?
You want to do something really fun, take an egg.
Have you done this?
You take the egg and you put it in the microwave and,
at some point, it goes, kapoof!
Yeah, right?
Now, I'll tell you a statistic that will surprise you.
One of the greatest incidences of cataracts happens
among people who work in the fast food industry.
Did you know that?
You know why?
The thinking is that, because of all the microwaves
that are used in food prep, that they're getting exposed
to microwaves and that's causing some crystals
of some sort to form in their eye and leading to cataracts.
You should reduce your exposure to microwaves.
Just because it's got a screen on there doesn't mean
that there aren't microwaves that are coming out there.
And, yes, cell phones use microwaves, too.
Whatever you do with microwaves, you should be reducing
your exposure to them, not increasing them.
So don't go press the eyeball up against the microwave oven,
watching that thing happen.
When I use the microwave at home, I usually stay at least
six to eight feet away from it at all times.
Seriously.
Yeah.
"Whoa, look at Kevin, he's over there!"
Right?
But, seriously, why should you increase your exposure to it?
Just like I wouldn't go and expose myself to X-rays any more
than I need to, I wouldn't expose myself to microwaves
any more than I need to, either.
All right, last thing on lactose.
The question about lactose intolerance always comes
up at this point, and you guys are on top of this, as well.
Lactose intolerance arises from a different problem.
Lactose intolerance arises because we lack the enzyme...
I shouldn't say we “lack."
Our body changes over time the amount of the enzyme
called "lactase" that it synthesizes.
When we're young, when we're an infant,
we're drinking milk, we need a lot of lactase because
that's the primary source of sugars and so forth
that we're getting in our diet.
It's coming through the milk.
Over evolutionary time, what's happened is,
we look at animals drinking milk or we look at humans
drinking milk, over evolutionary time, the only times
that really we needed to make that enzyme
is during that infant stage.
As humankind has developed organized farming
and we've had availability of dairy products, and so on,
and so forth, we have tended to eat those
dairy products longer in our lives.
While our body does continue to make some lactase
as we get older, it varies considerably from one culture
to another, from one ethnic group to another,
in terms of how much lactase is made.
If you don't make sufficient lactase,
what will happen is you're left with lactose.
You don't break it down into glucose and galactose,
and, as a consequence, lactose is metabolized by bacteria
in your stomach in a different way than these guys
are which get dumped into your bloodstream.
So the bacteria get a hold of this guy and they go crazy.
Of course, one of the byproducts
of metabolic action is carbon dioxide.
Well, gas, some severe problems relating to discomfort
and so forth, happen as a result of a deficiency of lactase.
It's happening because, again, as we're getting older,
we're making less of that enzyme.
There are commercial versions of lactase
that are available for people who are what are called
"lactose intolerant," that they can actually just
simply swallow when they have dairy products
and help to relieve that problem.
Other questions, comments?
Yes?
Student: So what happens to people who are, like,
deathly lactose intolerant, like, they can't stand
the smell of cheese, it just makes them feel bad?
Kevin Ahern: What happens to people who are,
what she says, "deathly" lactose intolerant?
I've never heard of such a thing, so I don't know about that.
A lot of people have either mental notions
of problems or there's other things that may relate.
So people who have issues with gluten intolerance
and so forth sometimes are diagnosed mistakenly
with lactose intolerance, and that may be a problem.
But I don't know of any deathly afflicted
with lactose intolerance.
Do you?
Student: Yes.
Kevin Ahern: You do?
Again, it could be an allergy or something else.
It's not directly lactose intolerance.
Lactose intolerance mostly comes about as a result
of the discomfort, the most common thing that's happening
is the discomfort that's there.
Student: I guess as long as that is related
in some form to, like, celiac disease?
Kevin Ahern: Celiac disease is related to, actually,
gluten intolerance, and there are some connections,
although they are distinct diseases,
but there are some connections between those two, yeah.
But, again, lactose intolerance,
different area, different area.
I had a student who was originally diagnosed
with lactose intolerance and then she just got the paranoia.
"I'm absolutely not going to have anything,"
“I can't be around milk," and so on and so forth
ane discovered that it wasn't lactose that she was intolerant to.
It actually was she was exquisitely sensitive to gluten
and was getting that in a variety of ways.
She could actually get gluten through milk,
that the cow was eating.
So she was very sensitive.
So when they got diagnosed they realized
what it was that was causing her problem.
Now we've talked about the metabolism of the sugars.
We've talked about how glucose gets broken down.
I want to spend a little bit of time talking about
regulation of the pathway and then,
when we come back on Wednesday
and I know you all are going to be here on Wednesday
when we come back on...
[scattered laughing]
Kevin Ahern: You're not all going to be here on Wednesday?
I know I'll be here on Wednesday.
When we come back on Wednesday I will then talk about
regulation again, in view of gluconeogenesis.
So we're going to get sort of a cursory look at
the regulation of glycolysis and then on Wednesday
we'll see how that ties to the regulation
of the synthesis of glucose,
as well, and the two are actually coordinated.
Let's start talking, first, about a really,
really interesting enzyme
[coughing]
excuse me, that, as I said before,
is the most important regulatory enzyme
that we see in glycolysis.
This is the enzyme phosphofructokinase,
or as you probably memorized it, PFK.
PFK is molecule that has a, I'm sorry,
is an enzyme that has a very interesting structure.
You see it is actually existing here as a tetramer,
and that enzyme has a very unusual behavior.
Let's take a look at this figure.
What we see is we're plotting, on the y-axis,
the velocity of the reaction that it catalyzes.
Now, you may not remember the reaction, so I will tell you.
The reaction that PFK catalyzes is as follows.
Fructose 6-phosphate plus ATP makes fructose 1,
6-bisphosphate plus ADP.
So it's using the energy of ATP to put the phosphate
onto fructose 6-phosphate to make fructose
1,6-bisphosphate.
Okay, simple enough, right?
If we do this reaction and we're plotting V versus S,
where the substrate that we're using is fructose 6-phosphate,
we see something very odd happening.
Now this is one of the few enzymes I know
that behaves in this way.
You might say, "What's so unusual?"
We look at this and we say,
"Okay, so it's got sigmoidal nature."
There's sort of an S shape there,
and in the presence of ATP, it's sigmoidal.
So ATP is an allosteric effector, right?
Well, ATP is a substrate, kind of like we saw with ATCase.
Remember, aspartate affected the enzyme, right?
Very similar.
However, if we do the same reaction with a small amount
of ATP, look how much the velocity goes up.
Now, remember, ATP is a substrate.
How do we increase the velocity
by decreasing the amount of a substrate?
We haven't seen that happen before.
So the enzyme is getting turned on by having
a less amount of one of its substrates.
That's very odd.
How would that manifest itself?
Any thoughts?
Yes, sir.
Student: Possibly the concentration is less important
than the effect of feedback inhibition
from high ATP concentration?
Kevin Ahern: He says possibly the effective concentration
is less important than the feedback inhibition
resulting from ATP concentration.
Well, yes, that's true, but that doesn't tell us
how that can happen.
You're right.
Yes?
Student: Is it possibly due to the Gibbs equation?
Kevin Ahern: Like what?
Student: Does it have to do with the Gibbs free energy?
Kevin Ahern: Does it have to do with the Gibbs free energy?
No.
Remember that enzymes never change
the overall Gibbs free energy.
So that's not it.
Yes?
Student: How many sides does the enzyme have?
Kevin Ahern: How many sides does the enzyme have?
The enzyme has, it's a tetramer,
so there are four different subunits that are there.
Student: Then the other subunits would
be available if low ATP was there.
Kevin Ahern: Well, you're getting there.
You're getting there.
She says the other subunits would be available
if low ATP was there.
Student: Are there two sites to bind, for ATP?
Kevin Ahern: Ahhhhh!!
Over here!
He'*** it on the nose.
It turns out the enzyme has two places to bind ATP.
One is an allosteric site and one is a catalytic site.
Which one do you suppose has the higher Km?
Which one is the enzyme going to have greater affinity for?
Student: Catalytic?
Student: Allosteric?
Kevin Ahern: You two want to duke it out?
[laughing]
Let's think about this.
When we have low amounts of ATP, the enzyme is more active.
What does that tell us?
Something.
So ATP is inhibiting the enzyme, right?
We all agree on that, right?
That's what we see here.
ATP is inhibiting the enzyme in some way.
Student: It's allosterically regulated, right?
Kevin Ahern: It's allosterically regulated.
So I'm asking you, does the allosteric site have a higher
affinity or does the catalytic site have a higher affinity?
Student: The catalytic site.
Kevin Ahern: The catalytic site's got
to have a higher affinity.
Right.
Because only when the ATP concentration is high does it start
banging into the allosteric site and turning it off.
That's a really cool enzyme, a very cool enzyme.
Student: [inaudible]
Kevin Ahern: Yeah.
It's responding to two signals.
This turns out to be really important because,
why do we want the enzyme turned off if there's high ATP?
Well, think about it.
Do we want to be burning gasoline,
do we want to be burning our furnace when it's summer?
No.
When we have plenty of energy,
do we want to be burning our glucose?
No, we don't, and we have plenty of energy
when we have high ATP.
High ATP should be turning this enzyme off,
and that's basically what we're seeing.
It's turning the enzyme, it's turning
the volume of that enzyme down.
That's really important.
On the other hand, what happens if we don't have much ATP?
Well, you betcha we want this enzyme going,
because we want to burn glucose so that we can get
pyruvate and we can get ATP and we can get the citric acid
cycle and we can get all these things going.
Low ATP is turning that enzyme on.
Really cool!
It's a very cool thing.
Now, phosphofructokinase turns out to be affected
by several things, as we shall see.
PFK gets affected by several allosteric effectors.
One of them is ATP.
Another very important one is this molecule called F2,6BP,
which stands for "fructose 2,6-bisphosphate."
One of the things you're going to see as we talk about
the regulation of the metabolic pathways relating
to sugars is, many of the names are going to sound similar.
"Fructose 2,6-bisphosphate" sounds an awful lot like
"fructose 1,6-bisphosphate" and because the numbers
ain't equal, you know that ain't the same thing.
Right?
You're going to have to spend some time getting straight
numbers and names, and you're going to see enzyme
names are going to overlap with these, as well.
For the moment, we're going to focus simply on this molecule,
fructose 2,6-bisphosphate.
Look what happens with this molecule.
Relative velocity, and we're doing the same plot
that we did before.
Here's the enzyme with no fructose 2,6-bisphosphate.
Here's the enzyme in the presence of 0.1 micromolar.
That's a very, very tiny amount.
Here's the enzyme in the presence of 1 micromolar.
Look at that!
***!
It's on!
Fructose 2,6-bisphosphate is present in very vanishingly
tiny quantities in our cell,
such vanishingly tiny quantities,
it wasn't even discovered until like the 1980s.
But in very tiny amounts, it turns this enzyme on superbly.
Yes, sir?
Student: Should the x-axis also be labeled
"F2,6BP" instead of "F6P"?
Student: No.
Kevin Ahern: This is a substrate.
No.
Student: Okay.
Kevin Ahern: This is an allosteric effector.
Student: Gotcha.
Kevin Ahern: Now, a very, very sensitive switch
to turn that enzyme on.
If we do the same plot and, instead of measuring the substrate
in fructose 6-phosphate, we measure it with ATP
we know ATP's got some weird relation with this enzyme.
We see that this activates the enzyme,
even in higher concentrations of ATP.
In other words, fructose 2,6-bisphosphate is a more
important regulator than ATP, itself, is.
Cells use fructose 2,6-bisphosphate as a way
of controlling this enzyme at very, very,
very sensitive levels, and we will see next week,
when I talk about glycogen metabolism,
how this ties into all of this.
It's a master picture that we think about
with respect to sugar metabolism that
is a really interesting and elaborate control.
But fructose 2,6-bisphosphate is probably
the most important regulator of this enzyme.
Now, the synthesis of fructose 2,6-bisphosphate
is a little complicated, and I'm going to save
that until I talk about gluconeogenesis,
but suffice it to say, cells have interesting ways
of making this molecule and it doesn't take
very much to get glycolysis going.
For those of you who wonder about structure,
there's the structure of fructose 2,6-bisphosphate.
We look at this and we remind ourselves
because I had to, myself, this morning, when I looked at this,
that we always want to number our carbons.
That will help us keep track of things.
Carbon 1... 2... I'm sorry.
Carbon 1... 2... 3... 4... 5... 6.
That's carbon-2 with the phosphate on it, right there.
Now, I've described situations to you where we have
muscle and where we have liver,
and we have to think about the different conditions
that the body has to respond to, relative to those situations
that muscle and liver find themselves in.
This schematically shows us
the overall pathway of glycolysis.
I told you that there are three enzymes
that play important regulatory roles.
The first is hexokinase and, as I said,
its regulation is a little odd.
Its product actually helps to turn it off.
That's not totally surprising.
It's known as "substrate-level regulation."
But that product affects this enzyme, hexokinase.
So hexokinase is the first regulated enzyme.
PFK is the second regulated enzyme, and we see,
for example, that ATP can turn it off.
This is a little confusing.
AMP can actually turn it on.
That makes sense.
AMP is an indicator of low energy inside of cells.
Low energy, you want to turn this enzyme on.
We also see that ATP, of course, as I mentioned earlier,
turns this enzyme off.
Fructose 2,6-bisphosphate turns the enzyme on.
Pyruvate kinase is the third regulated enzyme,
and I'm going to show you something about
that enzyme in a minute.
But I want to sort of remind you of something
that I talked about earlier, but I haven't had a chance
to finish the story on it.
Pyruvate kinase is regulated,
there goes our bouncing ball,
pyruvate kinase is regulated in two ways.
One, by phosphorylation.
Phosphorylation tends to turn it down or turn it off.
The other is by allosteric regulation,
and the allosteric regulation involved in controlling
pyruvate kinase is fructose 1,6-bisphosphate.
Fructose 1,6-bisphosphate will activate that enzyme.
It activates that enzyme.
And it activates that enzyme in a mechanism I refer to as
"feed forward activation."
Feed forward, that's the opposite of feedback inhibition.
Feedback inhibition said that the last molecule
turned off the first enzyme in the pathway.
Feed forward activation says a molecule in
the pathway turns on an enzyme further ahead.
Let's think about how this actually works in our body,
because it's kind of cool.
Let's imagine that we are sitting here and,
all of a sudden, the fire alarm goes off.
I've actually had this happen in this class.
The fire alarm goes off and we have to all go racing outside.
When that happens, the first thing that we have
to do is we have to get out.
We had to run a long ways, not in this building,
but we had to run a long ways to get out.
We need energy to get out, right?
What's going to happen?
Well, if I'm sitting here resting,
what's happening with my pyruvate kinase reaction?
Very little.
It's not doing much.
I'm not burning energy.
I'm not going through glycolysis much.
Things are just kind of sitting there
doing their thing, right?
However, when I get up and I start running,
my body starts, through epinephrine,
dumping glucose into my bloodstream.
My muscles take up that glucose
and now their glucose concentration is high.
What's going to happen with glycolysis?
It's going to start.
Great!
Glycolysis starts.
Glucose goes to glucose 6-phosphate.
Glucose 6-phosphate goes to fructose 6-phosphate.
Fructose 6-phosphate goes to fructose 1,6-bisphosphate.
And then we hit a wall.
Anybody remember what the wall is?
There is one enzyme that had a high positive Delta G.
Aldolase.
Aldolase is a wall.
We've got a high positive Delta G zero prime
at the aldolase and, all of a sudden, what starts accumulating?
The substrate for aldolase.
Look what it is, fructose 1,6-bisphosphate.
As fructose 1,6-bisphosphate accumulates,
trying to get over that energy hump,
what do you suppose it's going to do?
It's going to start binding to pyruvate kinase
and what's pyruvate kinase going to do?
It's going to take whatever phosphoenolpyruvate
that's there and convert it into pyruvate.
Now, because there's none of this, there was less of this,
this reaction goes forward.
This reaction goes forward.
This reaction goes forward.
And, finally, we've decreased the product
of the aldolase reaction, which is glyceraldehyde 3-phosphate
[stuttering]
and DHAP and G3P.
[laughing]
We've decreased the products of the aldolase reaction.
We've increased the reactants, we've decreased the products,
and that's how we get over the energy hump!
Feed forward activation is important for helping
us to get over that aldolase barrier.
So this occurs in the phenomenon I think I've talked about
previously, called "pushing and pulling a reaction."
We push a reaction when we increase substrate.
We pull a reaction when we decrease product.
The feed forward activation is decreasing product.
That's what it's ultimately doing.
As a result of that, this now goes all the way down
to pyruvate and we start making those ATPs we need to run away.
Student: You said decreasing the product is the pull part?
Kevin Ahern: Is what?
Student: The pull part?
Kevin Ahern: The pull part is decreasing the product.
That's right.
So, if we wanted to get our automobile out of the street,
if we had us pushing it, that's one thing.
But if there's only one or two of us pushing it,
that's not so good.
But if we had somebody on the other side with a truck
and a cable pulling the car and us pushing at the same time,
it's much more likely to move.
Same thing occurs with reactions.
Questions about that?
Let's go and take one quick look at
I think, yeah, let's not mess with that,
talk about pyruvate kinase.
Pyruvate kinase is the last enzyme in the glycolysis pathway.
As I said, it's catalyzing the big ***...the big ***.
So we want to have this enzyme under control because,
if we don't, as soon as we have PEP it's going to be
going straight to pyruvate.
One of the ways that we regulate this enzyme
is with phosphorylation.
Phosphorylation occurs as a result of action
of our friend protein kinase A.
Protein kinase A will catalyze the addition
of a phosphate to pyruvate kinase and cause it to become
into the less active state.
Pyruvate kinase is also regulated allosterically.
It's regulated by alanine, for one, alanine.
Alanine's an amino acid.
Why is alanine important?
Well, alanine is actually a good measure of pyruvate,
because pyruvate is readily converted into alanine.
When we have high alanine, we have high pyruvate,
we don't want this enzyme working, we will actually
allosterically turn this enzyme off with alanine.
ATP will also turn this enzyme off.
We have too much ATP, what's going to happen?
Well, we don't want to keep dumping more
and more of this stuff.
We're going to stop making pyruvate
and we're going to clog up the enzyme.
The allosteric activator, of course, as I said earlier,
is fructose 1,6-bisphosphate.
So we can turn the enzyme off, we can turn the enzyme on,
allosterically and by phosphorylation, and, of course,
dephosphorylation, to turn it on.
Now, this theme of allosteric regulation superimposed
on covalent modification is one that we will see
a lot of in glycogen metabolism.
So this is a little complicated, but the important
thing is understanding what all the pieces are,
not how they all play together.
What if I have a phosphorylated enzyme
and I have fructose 1,6-bisphosphate?
Well, it's kind of hard to mentally balance all of that.
I'm not going to go through and do that.
But I do think that you should know
the effects that phosphorylation has.
You should know the effect that fructose 1,
6-bisphosphate has, and you should know the effects
that ATP have on this enzyme.
Student: So phosphorylation,
the alanine and the ATP are offs?
Kevin Ahern: ATP and alanine are offs.
Student: And phosphorylation?
Kevin Ahern: And phosphorylation, too.
Right.
That's a lot of stuff with regulation there.
I want to say a little bit about GLUTs.
We talked a little bit about GLUTs earlier.
You may not remember, but when I talked about
the insulin signaling pathway,
I told you that the way that the body deals
with high blood glucose is by synthesizing insulin.
insulin went through that mulit step pathway
that involved the Hulk, that you recall.
The end result of the insulin signaling pathway, at least,
one of the end results, was that the movement of GLUT proteins
from the cytoplasm to the cell membrane was affected.
It favored the movement of those GLUTs and the "GLUTs"
stands for "glucose transport proteins."
There are many different GLUTs,
as you can see on the screen,
and they're located in various places in our body.
These GLUTs turn out to play some very important roles,
from a human health perspective,
and they may have some very, very important considerations
with respect to treating cancer.
So with that introduction, I want to tell you
about a very interesting phenomenon.
When we think of the development of a tumor,
as many of you have noted in here,
there's a multi-step process it takes to get to being a tumor,
and that multi-step process is probably different
for every different tumor that forms.
There's no one way of making a tumor.
There are many ways of making a tumor,
but there's no one single step that gets us to a tumor.
What people have noted about tumors is the following,
that tumors do tend to grow more rapidly than do other cells.
Tumor cells tend to grow more rapidly.
They are not organized.
They grow as a clump.
So what happens is, the needs and demands of a tumor cell
are greater than those of a non-tumor cell.
What happens in the process of that is really
interesting and cool.
It turns out that tumors,
because they are just growing in a place,
they don't have good access to blood supply, normally.
One of the things that tumor cells probably have to do
in order to survive, is they have to stimulate
the growth of blood vessels to supply them with blood.
There's a protein known as angiogenin that stimulates
the growth of blood vessels, and many tumors will,
in fact, activate or synthesize or accumulate angiogenin
and favor the growth of blood vessels to supply them with blood.
Yes, sir?
Student: Is the angiogenin a direct result of VEGF?
Kevin Ahern: I do not know.
Don't know.
Student: Because that's one thing we went over in cell biology,
the vascular endothelial growth factor.
Kevin Ahern: I don't know the answer to that question.
I can look it you up for you, though.
Now, angiogenin is favoring the growth
of blood vessels supplying tumors with blood.
Blood contains glucose, blood contains oxygen,
all the things that tumor cells, or any cell,
would like to have.
One strategy for treating tumors is to inhibit the growth
of the blood vessels and there are some promising drugs
that appear to do very well at basically
starving tumors to death.
That's kind of cool.
What is interesting is, well, if the tumor, as it's growing,
it takes a while to get these things synthesized,
it takes a while to get these blood vessels there.
As it's growing, if this tumor cell is growing faster
than its surrounding tissues,
its energy needs are greater.
There's not a blood supply.
There's not an oxygen supply.
These tumor cells are going to be what we call "hypoxic."
They're going to be low in oxygen.
I've already told you oxygen is necessary
for rapidly metabolizing cells.
We think of this tumor cell as a rapidly metabolizing cell.
It's taking up oxygen.
It's taking up oxygen faster than it's getting it
from the environment in which it finds itself.
[student sneezes]
Gesundheit!
Now, hypoxia is a normal phenomenon.
Our body goes through hypoxia all the time.
Our body has a response to hypoxia.
When we are hypoxic,
we make a protein called "hypoxia-induction factor."
It's a protein.
It's a protein that is a transcription factor.
A transcription factor activates
transcription of certain genes.
I want you to look at the genes that
hypoxia-inducing factor—"hypoxia-induction factor"
is what I call it—makes.
Look at this!
It's making GLUTs.
It's making hexokinase.
It's making phosphofructokinase. That's PFK.
It's making aldolase.
It's making...
look at all these enzymes of glycolysis!
This makes a lot of sense.
Let's think about what we've learned about glycolysis.
I told you that when we had plenty of oxygen
we produced a heck of a lot more ATP, right?
When we don't have plenty of oxygen,
we've got to burn more glucose to get the same amount of ATP.
These hypoxic cells are recognizing that.
They're making more glycolysis enzymes
so they can take in more sugar
so they can keep that cell alive.
What you know about glycolysis says,
"This makes perfect sense!"
And it also means maybe I've got a strategy
for how I might stop a tumor from growing.
If I can find ways to starve it to death,
by perhaps stopping this transcriptional activity,
affecting any of the enzymes preferentially in tumor cells,
I got a very cool way to knock out cancer.
Almost at the end.
Any questions about what I've just told you?
Question, yeah?
Student: What's the difference between a benign
and a malignant tumor?
Kevin Ahern: A malignant tumor is growing uncontrollably.
It will metastasize and kill you.
A benign tumor is growing controllably.
Other questions?
Was there another hand back there?
Okay, so I'll see all of you on Wednesday, I know.
Captioning provided by Disability Access Services
at Oregon State University.
[END]