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JOHN ESSIGMANN: Let's go now to storyboard 36,
starting with panel A, Regulation of Metabolism.
So far in 5.07, we have looked at metabolic pathways
from the perspective of the cell in organelles within the cell.
Using physiological scenarios, I've
tried to show you how pathways within the cell
respond to a change in the environment, for example,
by running away from a stressor, such as a dog
or dealing with the problems of starvation or diabetes.
We have looked a little bit at the ways
that individual steps and pathways are regulated.
We have not, however, looked at the ways
that individual pathways and individual organs
coordinate their respective activities in order
to accommodate the needs of the entire organism.
Coordinated pathway networking is the focus of this lecture.
Let's turn to panel B. I'm going to start out
by talking a little bit about the seven pathways
that we have studied in detail in 5.07
and look at how they're regulated.
As I've said before, typically pathways
are regulated at their rate-determining steps, that
is, the step at which you'll find an enzyme that
has a large free-energy change associated
with the conversion of substrates to products.
In the first pathway we studied, glycolysis,
there are three irreversible steps, specifically
the glucokinase/hexokinase step; secondly,
the phosphofructokinase-1 step; and thirdly, the last step,
pyruvate kinase.
These are all sites of pathway regulation.
Our second pathway, the tricarboxylic acid cycle,
or Krebs cycle, was regulated at every step at which NADH
is produced, that is, citrate dehydrogenase,
alpha-ketoglutarate dehydrogenase,
and malate dehydrogenase.
While pyruvate dehydrogenase is not formally
part of the TCA cycle, I'll point out here
that it is also regulated by NADH.
In all four cases, NADH feedback inhibits
the enzymes that produce it.
Under conditions of excessive TCA cycle activity,
you'll find that NADH levels will drop,
and accordingly NAD+ levels will increase.
The reduction in NADH will result
in activation of the pathway.
In other words, the disappearance of NADH
results in the uninhibition of the pathway.
Our third pathway, gluconeogenesis,
is regulated at the pyruvate carboxylase step
at the fructose 1,6-bisphosphatase step
and at the glycogen synthase/glycogen phosphorylase
steps.
We're going to be looking at this pathway in some detail
in a little while.
Our fourth pathway, fatty acid catabolism,
is regulated at the CAT, or Carnitine Acyltransferase step,
which is inhibited by malonyl coenzyme A. Malonyl CoA
is one of the key precursors for fatty acid biosynthesis.
It makes sense that the concentration
of malonyl coenzyme A, if high, would
inhibit the uptake of fatty acids
into the mitochondrial matrix.
Keep in mind that uptake into the matrix is the job of CAT1.
Shutting down CAT1 by the high concentration of malonyl CoA
prevents entry of fatty acids into the mitochondrial matrix,
where they otherwise would be subjected to beta-oxidation.
By turning off CAT1, malonyl coenzyme A
prevents the futile cycle of simultaneous fatty acid
degradation and biosynthesis.
The fifth pathway is fatty acid biosynthesis.
Acetyl-CoA Carboxylase, or ACC, is the enzyme
that makes malonyl coenzyme A out of its precursor,
acetyl coenzyme A. This enzyme, ACC, is activated by insulin.
As we've seen before, insulin detects the fed state
in the organism.
Hence, following a meal, insulin levels rise,
and that's the signal that tells the cells of the body
to take up nutrients.
That's just one example.
After a meal, glucose levels will rise in the blood.
Insulin will be released and drive the glucose
into the cell.
The cell will then activate pathways
by which the glucose is converted to acetyl-CoA.
Then the acetyl-CoA will be converted into fatty acids,
and then ultimately, into triacylglycerides
for energy storage.
Our sixth regulated pathway is the pentose phosphate pathway.
Glucose 6-phosphate dehydrogenase
is the first step in the pathway.
And as we often see, an early or committed step
is where regulation happens.
One of the products of the pentose phosphate
pathway is NADPH.
If the cell has used a lot of NADPH,
for example for biosynthesis, its levels will drop.
Coordinately, there will be an increase in NADP+.
In the case of glucose 6-phosphate dehydrogenase,
we find that NADP+, which is the product of excessive reductive
biosynthesis, as well as other activities,
such as combating oxidative stress,
activates the dehydrogenase in order to enable the synthesis
of additional NADPH in order to sustain biosynthesis.
The last pathway, the seventh, is electron transport
and oxidative phosphorylation.
This is our most robust pathway for the production of ATP.
When there's a lot of demand for ATP,
ADP levels will rise in the cell.
And it's ultimately the level of ADP
that regulates electron transport
in oxidative phosphorylation.
More about that in a few minutes.
Let's now look at panel C. Organs communicate
with one another by the nervous system, of course,
but also by hormones, small molecules that
are secreted by one organ that will
have an effect at one or more organs
distal to the first organ.
Changes in our environment are detected
by the brain with input from our sensory organs.
Our internal organs, such as the liver, pancreas, kidneys,
adrenal glands, and muscles will detect signals,
either independently of the brain or after an instruction
set is received from the brain.
The resulting signal network will allow the entire organism
to be able to adapt to the new environment,
be it one of stress, for example, being chased by a dog,
or one of, for example, hunger.
We'll talk later about the adrenal glands, which
are going to respond to signals that
come in from sensory organs that tell our muscles to start
running and to tell the liver to start to provide
the muscles with the glucose they need in order
to sustain running.
Later we'll be looking in some detail
at the adrenal-produced hormone epinephrine, also called
adrenaline.
Adrenaline will have a profound impact, both in the muscles
and in the liver, to allow these organs
to do their respective jobs.
The pancreas is a very important organ
in that it provides exocrine functions that
help with digestion and endocrine
functions that enable us to regulate or balance
fuel metabolism.
The alpha cells of the pancreas produce glucagon.
The alpha cells sense hunger.
They secrete glucagon into the blood,
which travels to organs that represent our fuel depots.
And fuel from those depots, for example, fatty acids
and glucose, will then be supplied
to other tissues of the body that are in need of nutrition.
The beta cells of the pancreas sense
what we call the fed state, and they produce insulin.
Insulin instructs the various organs
of the body to take up fuel in the wake of us
having eaten a meal.
Now let's look at panel D. There are
three general paradigms by which pathways are regulated.
The three hormones that I've just mentioned typically
will cause the activation of a kinase that will phosphorylate
a target protein, resulting in either increased or decreased
activity of that protein.
We call this hormonal or, more properly, covalent control,
because there will be a covalent modification of a protein that
will be responsible for pathway regulation.
The second major paradigm by which pathways are controlled
is allosteric regulation.
Earlier in 5.07, JoAnne Stubbe showed us
how hemoglobin, the molecule that
carries oxygen in the blood, is controlled
by bisphosphoglycerate and protons, which
act as allosteric effectors.
Similarly, other small molecule effectors
will control the major pathways of metabolism.
We're going to be looking at phosphofructokinase-1
as our prime example.
The third strategy of regulation is
what I call "acceptor control."
This is the way that electron transport and oxidative
phosphorylation are regulated.
To the right side of panel D is an abbreviated metabolic
pathway that I'm going to use to describe each of these three
paradigms.
At the upper left is a box that contains
glycogen synthase and glycogen phosphorylase.
Earlier in 5.07, I told you how phosphorylation
of a specific C-ring residue on glycogen
phosphorylase results in a dramatic increase
in the activity of that enzyme.
A little later, we'll see that covalent modification, again,
by serine phosphorylation of glycogen synthase, results
in a decrease in activity of that enzyme.
Hence, covalent modification of these two proteins
enables, in one case, activation of the enzyme
and, in the other case, inhibition of the enzyme.
This reciprocal control prevents futile cycling.
Now let's look at the box in the center
of this abbreviated metabolic pathway.
The glycolytic enzyme, PFK-1, Phosphofructokinase-1,
converts fructose 6-phosphate to fructose 1,6-bisphosphate.
We're going to see that a small molecule derivative of fructose
6-phosphate, specifically fructose 2,6-bisphosphate,
will act as a powerful allosteric stimulator of PFK-1.
I'll also point out that AMP, Adenosine Monophosphate,
will also stimulate this enzyme.
I'll also point out here that PFK-1 is a tetramer.
Oftentimes, as JoAnne taught us, proteins that are multimers
are the ones that are subject to allosteric regulation.
Our third paradigm of regulation is accepter control,
shown in the box at the bottom of panel D.
During periods of intense physical activity,
ATP is consumed and converted to ADP,
and NADH is converted to NAD+.
Rising levels of ADP in the mitochondrial matrix
activate the proton-translocating ATP
synathase.
As more protons flow through the synthase, more ATP is made.
They call this accepter control, because the regulatory molecule
is ADP, which is the, quote unquote, "acceptor
of phosphate" in the synthesis of ATP.
Because we covered accepter control
in some detail in the lecture on electron transport
and oxidative phosphorylation, I'm
going to leave that topic for now
and focus on covalent control and allosteric control.
Let's turn now to storyboard 37, starting
with panel A. With regard to the paradigm of covalent control,
I'm going to focus on the pair of enzymes, glycogen
synthase and glycogen phosphorylase,
which reciprocally regulate glycogen synthesis
and glycogenolysis.
Let's consider a scenario in which
there's some kind of stressor.
The muscles have to be activated in order
to be able to deal with the stress,
and the liver has to be able to provide the resources
to the muscles that will allow the muscles to continue
intense physical activity.
Now let's look at panel B. In our scenario,
you've just seen something frightening.
Your brain then sends an electrical signal
to the adrenal glands, which then
send a chemical signal, specifically adrenaline,
to the liver and to the muscles.
At the top part of this response scenario,
both the liver and muscles are going
to be responding in a very similar way.
However, at the bottom part of the response scenario,
the liver and muscles are going to be activating
very different pathways.
Specifically, we're going to see that muscles will strongly
activate glycolytic pathways in order
to enable the production of ATP to keep the muscles running.
The liver, by contrast, is going to be activating pathways that
are more like those of gluconeogenesis, that is,
spilling out fuel from the liver to provide
the muscles with the energy-rich resources
that it needs to keep us running away from our stressor.
Adrenaline, or epinephrine, travels from the adrenal
glands through the blood.
It takes only a second or two for this to happen.
It's received by the liver and the muscles
at the beta-adrenergic receptor.
The blood concentration of epinephrine is very low,
something of the order of 10 to the minus 10
molar at this point.
There's going to be a very substantial signal
amplification as we move ahead.
Keep in mind for later that the initial triggering
signal is in the 10 to the minus 10 molar range.
The arrival of epinephrine at the receptor
results in structural changes in the transmembrane domain
of the beta-adrenergic receptor.
The signal is received by a heterotrimeric G
protein, shown as the circle with alpha, beta, and gamma
subunits.
This G protein, in its inactive state,
has a bound molecule of GDP, that is, guanosine diphosphate.
Upon receipt of the signal from the beta-adrenergic receptor,
the heterotrimeric G protein ejects the beta and gamma
subunits, and the GDP molecule, which is non-covalently bound,
is released.
The GDP is replaced by a GTP, guanosine triphosphate.
This replacement results in the formation
of the alpha subunit with a non-covalently bound GTP.
This is the active form of the enzyme.
It translocates along the inner surface of the membrane
until it encounters AC, or adenyl cyclase.
The GTP-bound G protein activates adenyl cyclase.
The activated AC dynamically starts
converting ATP, which is very abundant in the cell,
into cyclic AMP.
We call cyclic AMP the second messenger.
In our scenario, the first messenger
is epinephrine, which interacted with
the beta-adrenergic receptor.
The second messenger is cyclic AMP,
which is produced by the activation of adenyl cyclase,
which is asymmetrically associated
with the inner surface of the cell's membrane.
Let's turn now to panel C. In the upper left of panel C,
you'll see a box.
This box depicts the chemical mechanism
leading to the production of cyclic AMP.
Let's now look at the main part of the panel.
In a very short period of time, a matter
of seconds, the concentration of cyclic AMP within the cell
goes from about 1 micromolar up to about 5 micromolar.
So the presence of epinephrine at the 10 to the minus 10
molar concentration outside the cell
results in a perturbation of cyclic AMP concentrations
inside the cell, bringing cyclic AMP concentration to about 10
to the minus 6 molar.
This is a very substantial increase
in signal, which an engineer would call gain.
The increasing cyclic AMP concentration
is going to activate a kinase cascade.
And the initial kinase that's going to be activated
is called Protein Kinase A, or PKA,
which is represented symbolically as a C
with a circle around it in my drawing.
In its inactive state, protein kinase A
is in a complex involving two molecules of itself
and two molecules of a regulatory protein, which
I've indicated as an R in the middle of a circle.
Cyclic AMP forms a tight complex with R.
This basically causes R to dissociate
from the active subunit C, that is,
the catalytic portion of protein kinase
A. The catalytic portion of protein kinase A
is now free to act as a kinase to phosphorylate
the next kinase in the cascade, which
is called SPK, or Synthase Phosphorylase Kinase.
A serine residue on SPK is phosphorylated,
and that event converts this kinase into its active form.
I'll also point out that one of the subunits,
the delta subunit of SPK, is the calcium-binding protein
calmodulin, which gives a second level of regulation
to this enzyme.
Specifically, this enzyme is regulated
not only by the concentration of cyclic AMP,
but also by the levels of calcium within the cell.
The activated SPK, or synthase phosphorylase kinase,
is now going to phosphorylate two other proteins, glycogen
phosphorylase and glycogen synthase.
As I mentioned earlier, phosphorylation of glycogen
phosphorylase results in the enzyme becoming more active.
That is, it starts breaking down glycogen by phosphorolysis
to produce glucose 1-phosphate, which
will then be available for further metabolic processing.
In the muscle, glucose 1-phosphate
will be converted by phosphoglucomutase
into glucose 6-phosphate, which will then be a substrate
to initiate glycolysis, with the ultimate generation of lots
of ATP for the muscle.
In the liver, phosphoglucomutase will
convert glucose 1-phosphate into glucose 6-phosphate,
but in this case, gluconeogenesis
will be activated.
Glucose 6-phosphatase will remove the phosphate
from glucose 6-phosphate, converting it to glucose.
The glucose will then be secreted from the liver,
go out into the blood, and then go
to the muscle to help the muscle carry out glycolysis.
Now let's look at the other target of SPK,
synthase phosphorylase kinase, specifically,
glycogen synthase.
In this case, glycogen synthase phosphorylation results
in a less active protein, and hence, glycogen synthesis
will cease.
This makes sense, because a cell under stress
would not want to be making glycogen. It wants to be using
energy and not storing fuel.
Let's step back and look at the big picture at this point.
The interaction of a hormone resulted
in the covalent modification of proteins.
One of those phosphorylated proteins,
glycogen phosphorylase, triggered glycogen breakdown.
Phosphorylation of a second protein, glycogen synthase,
turned it off.
Turning off glycogen synthase helped
us avoid the otherwise futile simultaneous synthesis
and breakdown of glycogen.
So overall, this is an example of a hormone causing
covalent modification of proteins in such a way
that it altered the activity of those proteins
in a manner that made sense given
the physiological challenges to the organism.
Next, let's turn to storyboard 38,
panel A. Our next regulatory paradigm
will be the use of allosteric effectors
to regulate a pathway.
As before, I'm going to use the physiological scenario
of stress.
But in this case, I'm going to show
how stress will cause the production
of small-molecule allosteric effectors
that will activate glycolysis in the muscle
and activate gluconeogenesis in the liver.
Our focal point is going to be PFK-1, phosphofructokinase-1,
of glycolysis.
Of central importance is going to be a small molecule,
fructose 2,6-bisphosphate, which we'll see is going to be a very
powerful allosteric stimulant of PFK-1.
Fructose 2,6-bisphosphate is made by the enzyme PFK-2,
phosphofructokinase-2.
Let's start out by looking at the very
short biochemical pathway shown in panel A. We see glucose
converted by hexokinase or glucokinase
to glucose 6-phosphate and then some equilibrium
steps up until we get fructose 6-phosphate.
Ordinarily, we think of fructose 6-phosphate
as continuing in the glycolytic pathway,
but let's not think of it that way right now.
The chemical structure of fructose 6-phosphate
is shown slightly to the right.
Note that fructose 6-phosphate has
a phosphate on the 6-hydroxyl group,
and that the 1-hydroxyl functionality
has only a hydrogen.
This is the alpha anomer of fructose,
as evidenced by the fact that the hydroxyl group on carbon-2,
the anomeric position, is down, or under the furanose ring.
With a little bit of electron pushing,
you can change the stereochemistry
at the 2-carbon such that the hydroxyl group would be up,
or in the beta configuration.
In the beta configuration, the hydroxymethyl group
of fructose 6-phosphate, which includes its 1-carbon,
would be down.
The beta and alpha-anomers of fructose 6-phosphate
are in equilibrium.
That is, they both exist at the same time.
The kinase, phosphofructokinase-2,
catches the beta anomer of fructose 6-phosphate
and phosphorylates it, producing the molecule at the lower
right of this panel.
That's the actual structure of fructose 2,6-bisphosphate.
This is the molecule that's going
to serve as the allosteric effector
of the glycolysis pathway.
The PFK-2, or phosphofructokinase-2 enzyme,
is shown to the left.
It's a complex enzyme having lots
of different functionalities.
The hatched circle within the larger circle at the top
represents the phosphofructokinase domain,
which converts fructose 6-phosphate to fructose
2,6-bisphosphate.
That is the structure at the lower right.
The hatched domain at the bottom is the fructose bisphosphatase
domain.
This is an unusual enzyme.
In the top domain, it acts as a kinase, in the bottom domain,
acts as a phosphatase.
PFK-2 has two hydroxylated amino acids,
one at about 3 o'clock as drawn and the other at about 6
o'clock.
When the hydroxyl group at the bottom
is present as a pure, unmodified hydroxyl group,
the protein acts as a kinase.
When the 6 o'clock domain is phosphorylated,
the enzyme acts as a phosphatase.
Protein kinase A, or PKA, from the previous storyboard,
where we talked about hormonal regulation,
is the kinase that phosphorylates
the hydroxyl group at 6 o'clock on the protein.
As we'll see later, the hydroxyl group at 3 o'clock
is targeted by a kinase called AMP kinase in the muscle.
Before we go on, I want you to look carefully at the structure
of fructose 2,6-bisphosphate at the bottom right of panel A.
Compare that structure with the structure of fructose
1,6-bisphosphate at the right in panel B.
If you squint and look at these two molecules,
you'll see that they look remarkably alike.
In both cases, the phosphate group
is above the plane of the furanose sugar.
However, in the case of fructose 2,6-bisphosphate,
the phosphate is on the 2-carbon,
whereas with fructose 1,6-bisphosphate,
the phosphate is on the 1-carbon.
The way these molecules look will become important
in a few minutes.
Now let's take a look at the pathway
as shown in panel B. Panel B reflects
the liver in its normal, that is, non-stress state.
So this is just the liver doing everyday liver business.
Glucokinase of the liver is converting glucose
to glucose 6-phosphate.
The alpha anomer of fructose 6-phosphate is produced.
Most of the fructose 6-phosphate is converted to fructose
1,6-bisphosphate.
In the liver cell at this time, there's
probably some low level of glycolytic activity going on.
However, because of natural anomerization,
some of the alpha-fructose 6-phosphate
is converted to its beta-anomer.
Phosphofructokinase-2, or PFK-2, is shown to the lower left,
and it's active.
Note that neither of its hydroxyl groups, at this point,
are phosphorylated.
And its kinase site will be converting the beta-fructose
6-phosphate anomer to its phosphorylated form, that is,
beta-fructose 2,6-bisphosphate, which has the structure shown
at the bottom right of the previous panel.
Fructose 2,6-bisphosphate is a positive allosteric effector
of phosphofructokinase-1.
So it facilitates the glycolytic pathway
that is enabling net flux from left to right in the pathway
as shown.
In addition to stimulating glycolysis,
fructose 2,6-bisphosphate strongly inhibits
gluconeogenesis.
It does this by binding at the active site of fructose
1,6-bisphosphatase, the gluconeogenic enzyme,
clogging up the active site, and thus disabling
the gluconeogenic pathway.
The reason that it's such a good inhibitor of gluconeogenesis
stems from an examination, once again,
of the structures of fructose 2,6-bisphosphate in the bottom
right of the previous panel and fructose 1,6-bisphosphate shown
to the right on this panel.
Keep in mind that fructose 1,6-bisphosphate,
while it's the product in glycolysis,
is the substrate for the gluconeogenic reaction.
If the active site of the fructose 1,6-bisphosphatase is
clogged by fructose 2,6-bisphosphate,
then it's not going to be able to process the fructose
1,6-bisphosphate in the gluconeogenic direction.
Hence, gluconeogenesis is strongly inhibited.
All of the above shows us that fructose 2,6-bisphosphate is
really important.
It stimulates the forward direction
in the liver that is causing glycolysis to have
a net flux from left to right.
And in addition, it strongly inhibits the back reaction,
that is, the reaction in the gluconeogenic direction.
Turn at this point to panel C. Now let's ramp up the scenario
and think about a situation in which you're
being chased by a dog.
You want to be able to generate glucose in the liver
and then export that glucose to the muscles
so that you can run away from your foe.
This is a situation in which you would like to have your liver
stop doing glycolysis.
You want the liver to strongly turn to gluconeogenesis
in order to manufacture glucose for the muscle.
At this point, refer back to the storyboard
that dealt with covalent control, where protein kinase
A was activated.
Recall the part of the lecture when
I talked about covalent control, and we
were talking about the glycogen phosphorylase
step in the liver.
In the liver, we generated first glucose 1-phosphate
and then glucose 6-phosphate.
And then the phosphate was lopped off the glucose
6-phosphate to produce glucose that went out into the blood.
In the present case, we're looking
at a step that's further down in the gluconeogenesis pathway.
Specifically, we want to activate the fructose
1,6-bisphosphatase in order to push even more carbohydrate
toward the production of glucose.
So in panel C, we see stress inducing the cyclic AMP cascade
that results in activation of protein kinase A. And protein
kinase A is going to phosphorylate
the southernmost, that is, 6 o'clock domain, on the PFK-2,
or phosphofructokinase-2 protein.
As I indicated earlier, phosphorylation
of the 6 o'clock domain results in conversion
of phosphofructokinase-2 from a kinase into a phosphatase.
The phosphatase produced from PFK-2 will act upon the pool
of fructose 2,6-bisphosphate and convert that pool to fructose
6-phosphate.
That is, the 2-phosphate will be lopped off of the fructose
2,6-bisphosphate molecule.
By doing this biochemistry in the liver,
we've done two things.
First, we destroyed the fructose 2,6-bisphosphate pool.
Hence, this molecule is no longer
available to act as an allosteric stimulator of PFK-1.
Secondly, removing fructose 2,6-bisphosphate from the pool
has resulted in removal of the active-site inhibitor
of fructose 1,6-bisphosphatase.
This enzyme, therefore, becomes active
and is able to push the net flux of carbon
from right to left in the direction of gluconeogenesis,
producing glucose that will then spill out into the blood.
Please turn now to panel D. We just
saw that the liver, post-stress, produces glucose.
Now we're going to take a look at the muscle, which,
of course, is going to be using that glucose.
In response to the stress state, the muscle
is going to activate an enzyme called
the AMP-dependent protein kinase,
or AMP kinase, also known as AMPK.
AMPK activates metabolic pathways that generate ATP.
The 3 o'clock phosphorylated PFK-2 proves to be
an exceptionally competent enzyme for converting fructose
6-phosphate into fructose 2,6-bisphosphate.
This increases further the concentration of fructose
2,6-bisphosphate in the muscle cytoplasmic pool.
And keep in mind that fructose 2,6-bisphosphate itself is
a powerful allosteric stimulator of glycolysis by its effect
on PFK-1.
So in the muscle cell, we're able to achieve extraordinarily
high concentrations of fructose 2,6-bisphosphate,
thus favoring glycolysis.
Let's turn now to storyboard 39.
Let's start with panel A. The way
that AMP kinase stimulates glycolysis in the muscle
post-stress is illustrated in this abbreviated metabolic
pathway.
Starting at the lower left, the stressor
has created the cyclic AMP cascade
that activates AMP kinase.
AMP kinase phosphorylates the 3 o'clock domain on PFK-2,
converting it to its phosphorylated derivative that
shows powerful kinase activity to convert fructose 6-phosphate
into fructose 2,6-bisphosphate.
This enhanced pool of fructose 2,6-bisphosphate will now act
as a powerful allosteric stimulator of PFK-1 to favor
glycolysis.
I want to point out that muscles do not do gluconeogenesis.
Only the liver and renal cortex do this pathway.
So muscles do not have fructose 1,6-bisphosphatase,
the gluconeogenic enzyme.
Accordingly, in muscles we don't have
to worry about gluconeogenesis being turned on to some extent
and thus dampening the glycolytic effect
in the muscle that's needed in order for the muscle
to generate the ATP needed to outrun your foe.
On this panel, we also see that there
are several other effectors that influence
the activity of PFK-1.
One is citrate.
Another is ATP.
Another one is AMP, or Adenosine Monophosphate.
I want to say a couple of words about AMP in this regard.
Let's look at panel B. AMP is not as powerful an allosteric
effector on PFK-1 as is fructose 2,6-bisphosphate.
But we know a lot about its activity
as an allosteric effector because it's
been studied quite thoroughly.
PFK-1 is a tetramer, and its activity
is regulated by cooperativity.
This graph shows the activity of PFK-1
as a function of the concentration of one
of its substrates, fructose 6-phosphate.
When there's no ATP present, which
is not a realistic situation, you
see the regulatory pattern following that
of a rectangular hyperbola.
That's scenario A.
Scenario B shows what would happen
to the activity of the enzyme when
ATP is present at a concentration of about
1 millimolar, which is quite realistic,
and what you see is suppressed activity.
When the heterotropic allosteric effector AMP is added,
you see the curve shift to the left.
That is, you get enhanced activity at a given substrate
concentration.
My guess is that fructose 2,6-bisphosphate would push
the curve even further to the left.
Let's now look at panel C. I think it's useful to use AMP
to help us construct a model for what fructose 2,6-bisphosphate
might be doing as an allosteric stimulator of the PFK-1 enzyme.
The equilibrium diagram that I've drawn
shows the relationship between the relaxed, that
is, more active state of the PFK-1 tetramer,
and the tense, that is, more inactive state of the enzyme.
When ATP is abundant within a cell,
it binds more strongly to the tense state,
and hence, ATP inactivates PFK-1.
This makes sense, because if ATP is abundant,
you do not want to be doing glycolysis
to make even more ATP.
However, when the ATP pool of a cell
is challenged by doing heavy work or biosynthesis,
the AMP is going to bind more tightly
to the R state of PFK-1.
And that's going to help convert the enzyme
into its more active state.
That's a little snapshot of how a small molecule, in this case,
AMP or perhaps fructose 2,6-bisphosphate can
dramatically regulate the activity of an enzyme
at a specific step in metabolism.