What do you think about ketogenic diets?
Whether you're on one or thinking about
it, you love it, you hate it, or you're curious,
you should know how it works. And this
video is about how ketogenesis works.
A ketogenic diet has neurological benefits.
Why do we have to eat such an enormous
amount of food?
Complex science.
Clear explanations.
Class is starting now.
Hi I'm Dr. Chris Masterjohn of
chrismasterjohnphd.com. And you're
watching Masterclass with Masterjohn.
We are now in our 32nd in a series of lessons on
the system of energy metabolism.
And today we are talking about ketogenesis.
Ketogenesis is an important component
of how we react to glucose deprivation.
In the last three lessons we talked
about gluconeogenesis in the
context of how if we don't have enough
glucose we need to make it.
But remember the main thing that we
make glucose from is protein. And if we
are in a state of glucose deprivation,
we want to take down our glucose
requirement because if we
just turn all the protein we have into
all the glucose we would normally use on
a mixed diet, we're going to burn through
a lot of protein really fast. And if you
imagine that you're on a diet that's
mostly fat, which is often used as a
ketogenic diet today, or in the context of
why these systems evolved to handle
glucose deprivation in the first place,
like fasting; you're going to have to tap
into your skeletal muscle and burn
through it really fast if you can't push
down your glucose requirement.
So while gluconeogenesis is how we make the
glucose that we don't have,
ketogenesis is an important part of how
we conserve the glucose that we do have,
or that we do get from protein, so we
don't use it unnecessarily when we can
get alternative fuels.
And the main thing that ketogenesis is
doing is allowing tissues that aren't good at using fatty
acids themselves to use ketone bodies,
which are water-soluble much smaller
pieces of fatty acids made mostly by the
liver; and use those for energy in place
of using glucose or using fatty acids
themselves. So clarifying the purpose as
we just have, let's dig right into how
the process of ketogenesis works.
As we first covered in lesson 4,
in order for acetyl CoA to enter the
citric acid cycle it needs to condense
with oxaloacetate to form citrate,
leaving coenzyme A free in that
reaction to pick up the next acetyl
group. That means that in order for
acetyl CoA to enter, the supply of
oxaloacetate, whether oxaloacetate is
there, is important. The biochemical event
that initiates ketogenesis is the
accumulation of acetyl CoA when there
is no oxaloacetate to drive it into the
citric acid cycle to form citrate. This
doesn't mean there's ever no
oxaloacetate that oxaloacetate is at
zero supply. What this means is that if
acetyl CoA is coming into the citric
acid cycle and then oxaloacetate gets
used up because there's only a certain
amount available, all the excess acetyl CoA
that's leftover that can't enter
the citric acid cycle will then undergo
ketogenesis. But as we covered in lesson
16 on anaplerosis, normally pyruvate
serves as the source of oxaloacetate
whenever it's limiting.
Remember the main fate of pyruvate is to
undergo pyruvate dehydrogenase to form
acetyl CoA.; if the acetyl CoA has
oxaloacetate to enter the citric acid
cycle it does, if it doesn't it inhibits
the pyruvate dehydrogenase complex
temporarily and stimulates pyruvate
carboxylase. The inhibition of pyruvate
dehydrogenase conserves pyruvate to
enter through pyruvate carboxylase
instead and acetyl CoA increasing the
activity of this enzyme, which would
otherwise be one of the few things in
biochemistry that ever is at zero
activity in the absence of acetyl CoA
binding to it. Acetyl CoA amping up
pyruvate carboxylase
allows pyruvate to form the oxaloacetate
that's needed to allow acetyl CoA to
enter again. Then that acetyl CoA stops
accumulating, pyruvate carboxylase goes
back to zero, and the main fate of
pyruvate gets resumed to form acetyl CoA.
This means that for ketogenesis to
happen you have to have depletion of
oxaloacetate that is not supplied in
this way immediately thereafter. That
means in order to get ketogenesis
you need depletion of pyruvate.
Although the main source of pyruvate on a mixed
diet is carbohydrate, because pyruvate is
the end product of glycolysis and
therefore is derived from glucose, we
also talked in lesson 16 on anaplerosis
that the amino acid alanine can form
pyruvate. And in fact we also talked in
lesson 8 about how aspartate, another
amino acid, can directly form
oxaloacetate. And how glutamate, another
amino acid, can form alpha-ketoglutarate.
If alanine forms pyruvate, the pyruvate
supplies oxaloacetate that allows
acetyl CoA to enter the citric acid cycle
and that prevents acetyl CoA from
accumulating and entering into ketogenesis.
If glutamate supplies alpha-ketoglutarate
that alpha-ketoglutarate
becomes oxaloacetate that does exactly
the same thing again, preventing
ketogenesis.
Not shown on this screen,
there are many other amino acids that we
will talk about when we get to protein
metabolism that can either form pyruvate
or oxaloacetate or one of the precursors
of oxaloacetate in a manner analogous to
what's shown on the screen for alanine,
aspartate, and glutamate, the amino acids
that we've already talked about in
previous lessons. What that means is that
anything that makes oxaloacetate
suppresses ketogenesis. And therefore
protein, through a great variety of amino
acids, as well as glucose, can equally
suppress ketogenesis at a biochemical
level. Now it's important to distinguish
between the biochemical suppression of
ketogenesis and the physiological
suppression of ketogenesis. Biochemistry
is the study of the reactions and the
enzymes and the pathways that they come
together to make. Physiology is the study
of how biochemistry and other processes
within cells, and between cells, are
coordinated in a systematic way to
support the function of the organism as
a whole. On a biochemical level
carbohydrate and protein are equally
suppressive of ketogenesis.
That's not true on the physiological
level. In order to understand the
physiological suppression of ketogenesis
by carbohydrate and insulin we need to
review lipoprotein lipase, which we first
introduced in lesson 23 on insulin
secretion, and hormone-sensitive lipase,
which we first introduced
in lesson 25 on how insulin shuts down fat
burning. We then revisited lipoprotein
lipase again in lesson 26 on the
question of whether insulin makes you
fat. We're going to now come back to
these same effects of insulin to look at
how insulin suppresses ketogenesis on a
physiological level. To briefly review
lipoprotein lipase and hormone-sensitive
lipase, they're shown on the screen.
The left panel shows lipoprotein lipase.
Lipoprotein lipase is secreted by an LPL-
producing cell, LPL is lipoprotein lipase,
and that cell will send the LPL out to
be embedded in the capillary endothelial
cells of the capillaries that feed that tissue.
For the moment since we're mostly
concerned with adipose fat let's think
mostly about the adipocyte. So the
adipocyte secretes LPL into the
capillaries that feed the adipose tissue,
and the LPL goes into the capillary
endothelial bed. Chylomicrons carry
triglycerides from the fat in the meal
that you had just eaten, and the
triglycerides are hydrolyzed by LPL to
glycerol and free fatty acids leaving
behind chylomicron remnants that later
get taken up mostly by the liver, and the
glycerol and fatty acids come into
adipose tissue. There are other enzymes
that will then resynthesize triglycerides
from the glycerol and fatty acids.
Meanwhile the adipose tissue,
as well as many other cells, but the
adipose tissue for purposes of releasing
fatty acids into the blood has hormone-
sensitive lipase or HSL, and this is
responsible for hydrolyzing
triglycerides inside the adipocyte. They
become glycerol and fatty acids for
release into the blood. Now HSL and other
tissues besides adipose tissue serves
the internal needs of that tissue.
But in adipose tissue we have
LPL and HSL that determine the
flux of free fatty acids.
The more LPL activity we have the more
fat will get taken up into adipose tissue.
LPL activity can release fatty
acids into the blood if they exceed the
ability of the adipose tissue to take
them up, but its main function is to get
fat into the adipose tissue. HSL serves
the opposite function. So we retain fat
in adipose tissue when we have high LPL
activity at adipose and low HSL activity
at adipose. We have a lot of free fatty
acids moving from adipose tissue into
the blood when we have the opposite
conditions. Insulin promotes the
retention of fat in adipose tissue. Now
we talked about why that doesn't mean
that carbohydrate intrinsically makes
you fat in lesson 26. But for the
purposes of discussing the physiological
suppression of ketogenesis it's
important to note that fatty acids being
released into the blood will become the
major source of acetyl CoA that can't
enter the citric acid cycle. So it's very
relevant that insulin promotes the
retention of fat in adipose tissue.
Remember under conditions of high
insulin signaling LPL is reduced in
heart and skeletal muscle, favoring fat
not being taken up into those tissues.
Insulin increases LPL at adipose tissue,
favoring taking up fat there. It also
suppresses HSL at adipose tissue
favoring the retention of that fat
inside the adipocyte instead of allowing
its release into the blood. It's also
important to understand the influence of
insulin and glucagon on gluconeogenesis
because gluconeogenesis affects the
supply of oxaloacetate. Remember that
insulin and glucagon as well as energy
status regulate it. When you have
conditions of high glucagon signaling
and low insulin signaling and when you
have lots of energy inside the liver
rather than low energy in the liver
that's when you get a lot of
gluconeogenesis. When you have low energy
status in the liver or you have a high
insulin-to-glucagon ratio, that's when
you suppress gluconeogenesis.
The conditions that favor maximal
ketogenesis in the liver are the conditions that
allow a high rate of entry of fatty
acids into the liver and a high degree
of gluconeogenesis. Fatty acids when they
reach the liver are undergoing beta-
oxidation and generate acetyl CoA.
If that acetyl CoA was generated by
carbohydrate or protein it would be
relatively easy for anaplerosis, introduced
in lesson 16, to supply the oxaloacetate
that would be needed to take that acetyl
CoA into the citric acid cycle. But
since fatty acids generate acetyl CoA
without a very quantitatively
significant way of also supplying
anaplerosis, it's fatty acids that will generate the
acetyl CoA that can't enter the citric
acid cycle. Under conditions of low
insulin signaling when you have lots of
fatty acids entering into the liver in
this way, those are the same conditions
that have oxaloacetate, whether derived
from aspartate, from other precursors in
the citric acid cycle, or from pyruvate,
oxalocetate leaves the citric acid
cycle for gluconeogenesis under
conditions of low insulin signaling. So the
conditions that favor fatty acids
reaching the liver are the same
conditions that favor oxaloacetate
leaving the citric acid cycle, which all
the more enhances the likelihood that
that acetyl CoA will not be
able to enter the citric acid cycle and
will therefore generate ketones. Now keep
in mind that if you have fatty acids
being released from adipose tissue and
it's the liver that's bearing the brunt
of metabolizing those fatty acids for
the needs of the rest of the body that
also means that the fatty acids can
supply the liver with energy and that
energy can be used for gluconeogenesis.
So this diagram is not meant to imply
that none of the acetyl CoA from the
fatty acids enters the citric acid cycle.
Instead some of it does and what does is
used to supply the high energy status in
the liver that can contribute to the
synthesis of glucose. But the synthesis
of glucose requires depletion of
oxaloacetate from the citric acid cycle.
And there's tons of acetyl groups from
all this incoming fatty acid
beta-oxidation that are now left over
that after the needs
for energy status are met cannot
enter the citric acid cycle because even
that energy that they supplied helped
the oxaloacetate leave the cycle for
gluconeogenesis. All of this together is
what contributes to the biochemical
event, *the* biochemical event that initiates
ketogenesis, which is the accumulation of
acetyl CoA that can not enter the
citric acid cycle because of a relative
lack of oxaloacetate. When thinking about
the physiological suppression of
ketogenesis and distinguishing between
the effects of carbohydrate and protein
we need to think about the effects of
these nutrients on insulin and glucagon
that was discussed briefly in lesson 23.
So remember that carbohydrate leads to
more insulin production than protein
does, but protein still stimulates insulin.
By contrast carbohydrate suppresses
glucagon and protein raises it.
Now remember we briefly
talked about this as a way of preventing
hypoglycemia when you eat protein.
Because if protein stimulates some
insulin, which helps the protein's amino
acids be taken up into cells and undergo
metabolism, and that protein just makes
insulin, it's going to cause hypoglycemia
because the insulin also drives
carbohydrate into cells.
So by also stimulating glucagon we could see that
as a way of preventing the hypoglycemia
that protein would cause if it only
stimulated the secretion of insulin.
But now we can look at how this plays
into ketogenesis.
When we're talking about the release of
fatty acids from adipose tissue, it's insulin
that's almost by itself relevant rather
than glucagon. But when talking
about gluconeogenesis
it's the insulin-to-glucagon ratio
that's key. So when we have carbohydrate
restriction we get a lot of glucagon and
less insulin. The less insulin is the main
thing driving up free fatty acids
in the blood.The low insulin-to-glucagon
ratio is the main thing driving
increased gluconeogenesis. The influx of
free fatty acids in the blood mostly
goes to the liver, or at least the lion's
share of those fatty acids go to the
liver, and that generates a lot of acetyl
CoA in the liver. Meanwhile the
gluconeogenesis occurs
from oxaloacetate leaving the citric
acid cycle. And so gluconeogenesis
contributes to the relative deprivation
of oxaloacetate inside the liver. That
means that the hepatic acetyl CoA-to-
oxaloacetate ratio goes through the roof.
And that leads to ketogenesis.
So when talking about physiological
suppression it's mainly carbohydrate
that's suppressing ketogenesis. And what
we mean by physiological suppression is
the prevention of free fatty acids from
reaching the liver and the suppression
of hepatic gluconeogenesis.
By contrast it's still the case that at a
biochemical level its carbohydrate and
protein that are equally suppressive of
ketogenesis, and by biochemical
suppression we mean anaplerosis,
covered in lesson 16, the provision of
oxaloacetate or one of its precursors.
We'll look in the future how this might
play out on a quantitative level in the
diet, but what it ultimately means is
that because your diet influences your
physiology and your biochemistry then
dietary protein will surpress
ketogenesis, just not as much as dietary
carbohydrate will. So, mechanistically, why
is it that the accumulation of acetyl CoA
that can't enter the citric acid cycle
in the liver generates ketone bodies?
The reason is because of the biochemical
pathway shown on the screen. What we're
doing in this biochemical pathway is
we're ultimately joining
acetyl CoA in multiple units to make
acetoacetate. As its name implies
acetoacetate is essentially two acetyl
groups joined together, but in the
process we're actually joining three and
we're cleaving apart the intermediates in
a way that leaves us with what appears
to be two structurally joined units.
So the first thing that happens is acetyl CoA
condenses with another acetyl CoA
using the enzyme beta-ketothiolase,
which cleaves the CoA from one of the acetyl CoA,
but not the other. And so you can see this
color-coded where one acetyl CoA is in
pink the other is in blue, they join
together, but in the process the enzyme
beta-ketothiolase takes away this CoA
on top that allows this one join to that one,
that CoA leaves and what you have
left over is acetoacetyl CoA and
you can see the blue from the acetyl
group that came in on the bottom and the
pink from the one that came in on top.
So we still have this acetoacetyl unit, two
acetyl groups joined together that are
also joined to CoA. A third acetyl CoA
shown in green comes in and the enzyme
hydroxymethyglutaryl CoA synthase
hydrolyzes that acetyl CoA so that CoA
leaves and its acetyl group is joined to
the bottom of acetoacetyl CoA. That
generates 3-hydroxy-3-methyl-glutaryl CoA.
Then hydroxymethylglutaryl CoA
cleavage enzyme cleaves off
from the end what is now one unit of
acetyl CoA to generate the final product, and
first ketone body, acetoacetate.
The names of these molecules are
important and the
enzymes in this pathway are also
important. But we're going to come back
to look at the intermediates of
ketogenesis later when we talk about how
the metabolism of certain amino acids
intersects with this pathway during
protein metabolism. For now we're going
to show the details, but focus on why
acetyl CoA enters this pathway, which
we've already done, and then what happens
to acetoacetate once it's made, which
we'll now go on to do now.
Acetoacetate is the first
of the three ketone bodies.
It has two major fates. One is to be
converted to D-3-hydroxybutyrate and
the other is to be converted to acetone.
These three together make up the ketone bodies.
Now if acetoacetate gets converted to
D-3-hydroxybutyrate, it will be because
the hepatic ratio of NADH to NAD+ is
high. This conversion is enzymatic and
completely reversible.
D-3-hydroxybutyrate dehydrogenase is
what reduces the keto group of
acetoacetate -- remember the carbonyl
that's situated between two other
carbons is a keto group -- it reduces the
keto group to a hydroxyl group. And so
actually D-3-hydroxybutyrate is not a
ketone, chemically,
even though it's one of the 3 ketone bodies.
If you look at the name of this
compound it's called 3-hydroxybutyrate
because if we start with the carboxyl
group that makes this an acid, which is
the most important functional group, we
can count the carbons 1, 2, 3, and the
hydroxyl group is on carbon 3.
We call it D-3 because once we reduce
the keto group to a hydroxyl group we now have
stereoisomerism. We get stereoisomerism
when there's a chiral carbon. A chiral
carbon is a carbon that's attached to
four different things. We look at one as
the hydrogen, one is an OH group, one
is a methyl group, one is the rest of
this molecule, there's four different
things attached to that carbon; that's
not true in the carbon before it was
reduced. Now that it is, that means that
with a chiral carbon the way those
different pieces are oriented around the
carbon can make the difference between
one isomer and another and
we use letters like D and L to distinguish
between the possible confirmations. And
so the full name of this is a D-3-hydroxybuyrate.
However, we could also call this
beta-hydroxybutyrate because if we look
at the carboxyl group we care about the
next carbon is alpha to it and the
carbon after that is beta to it. So this
is beta-hydroxybutyrate, and in fact
it's much more common in even scientific
papers let alone pop science culture to
call this beta-hydroxybutyrate.
The other fate of acetoacetate is a slow
and spontaneous decarboxylation to form
acetone. That happens when the carboxyl
group shown in purple comes off, forming
acetone. The reduction of acetoacetate to
beta-hydroxybutyrate is very analogous
to the reduction of pyruvate to lactate,
covered in lesson 15. Acetoacetate is a
beta-keto acid, and beta-hydroxybutyrate
is a beta-hydroxy acid. That's because if
we count up from the carboxyl group,
alpha, beta, we have a beta-keto group on
acetoacetate and we have a beta-hydroxy
group on beta-hydroxybutyrate.
When we go from pyruvate, an alpha-keto
acid, to lactate, an alpha-hydroxyacid,
we're just doing the same thing. The only
difference is that if we count from the
carboxyl group on pyruvate the keto
group is in the alpha position and in
lactate the OH group that's derived from
the keto group of pyruvate is in the
alpha position as well. Other than that
these reactions are almost identical.
We have beta-hydroxybutyrate dehydrogenase
on the one hand, and lactate
dehydrogenase on the other hand.
In both cases we have
NADH reducing the keto group to the
hydroxy group. And in both cases doing
that liberates NAD+ and going
in the opposite direction
sequesters NAD+ as NADH. This is
interesting for two reasons. From just
the perspective of what we call these
molecules we're talking about
ketogenesis and we're associating it
with fat and yet beta-hydroxybutyrate,
which is sold as an exogenous
"ketone," is not a ketone. It's a "ketone
body" because historically we've referred
to the process of ketogenesis as what
happens in uncontrolled diabetes or on
ketogenic diets and beta-hydroxybutyrate
pops up into the blood under those
conditions. So we call it a ketone body.
But chemically it's not a ketone.
Acetoacetate is a ketoacid, and pyruvate,
derived mostly from glucose, is a
ketoacid, and it generates lactate, which is
not a ketone in the same exact way that
beta-hydroxybutyrate is not a ketone.
That's interesting just for the sake of
curiosity. What's interesting from a
practical perspective is that in later
lessons we can talk about how the
conversion of acetoacetate to beta-
hydroxybutyrate can be a way of
rescuing mitochondrial NAD+ in just the
same way that converting pyruvate to
lactate is a way of rescuing cytosolic
NAD+. And that can be an important
part of how ketogenesis could help with
certain aspects of energy metabolism.
The slow and spontaneous
decarboxylation of
acetoacetate to form acetone is
analogous to the decarboxylation of
oxalosuccinate to form alpha-ketogutarate
in the citric acid cycle that we talked
about in lesson 6.
If you look at acetoacetate
and you count the carbons from the
carboxyl group it decarboxylates, and you
count up alpha, beta, you can see that the
ketone is in the beta position to that
carboxyl group, which makes it a beta-
keto acid. In oxalosuccinate we have
the carboxyl group that's going to
decarboxylate shown in purple and if we
count the carbons next to it alpha, beta,
we see the keto group is in the beta
position, again making oxalosuccinate a
beta-keto acid. For the reasons that we
described in exhaustive detail in lesson 6,
beta-ketoacids are unstable and tend
to decarboxylate. The same thing that
makes oxalosuccinate decarboxylate to
alpha-ketoglutarate in
a rapid enzymatic fashion
because it's catalyzed makes acetoacetate,
which doesn't have an enzyme to
catalyze this, slowly but spontaneously
decarboxylate to form acetone. Whereas
acetoacetate and oxalosuccinate
are ketoacids, and in fact even
alpha-ketoglutarate is a ketoacid, acetone is
a simple ketone. It has a keto group, a
carbonyl between two carbons, but it's
not an acid. Even more recently in lesson
29 we saw this slide about how we get
around the irreversibility of the
pyruvate kinase reaction in
gluconeogenesis by forming oxaloacetate
from pyruvate, which is an unstable
beta-ketoacid and that helps us
form phosphoenolpyruvate,
which is an even more
unstable enol. We talked about the
rationale behind this in lesson 29 so
we're not going to talk about it again,
but we'll briefly note here that
oxaloacetate is a beta-keto acid and
that confers instability that makes it easier
to form phosphoenolpyruvate
in the energy-intensive way
described, catalyzed by PEPCK, in
gluconeogenesis. One final point about
acetone. Because it's not an acid like
acetoacetate is it's very volatile.
Now we first talked about the volatility of
acetic acid in lesson 3 when we
introduced cellular respiration and
talked about how one of the roles, not
the only role by any means, but one of
the roles of coenzyme A is to weigh down
small molecules like acetyl groups
because acetic acid would be volatile.
But then in lesson 13 we talked about
how we don't form acetaldehyde in
significant amounts in metabolism not
only because it would be toxic
but because acetaldehyde has even weaker
reactions with water than acetate does
or acetic acid does, making it even more
volatile than acetic acid. Out of all of
these, acetone is the most volatile.
And we can see why by the slide on the
screen that compares the interactions
with water. Remember interactions with
water are facilitated by polar bonds and
especially by charged ions. In acetate we
have small size, which makes it more
volatile, but we have this full negative
charge on the oxygen, which provides
incredibly strong interactions with
water. The polarity around the carbonyl
group also provides interactions with
water but nowhere near as strong as
those conferred by the full negative
charge on the oxygen. In acetone we do
have some polarity around the carbonyl,
but we don't have any other polar bonds
and we don't have any full charges.
So acetone is also
very small like acetate, but it has very
little causing it to interact with water.
If it's floating through your blood,
which is mostly water, and there's
nothing holding it in that aqueous
solution because of interactions with
that water, then it's small size and the
fact that it doesn't really care about
any of the water in its surroundings
makes it very easily evaporate through
the lungs. That's what we call ketone breath.
Ladies will recognize acetone as
the smell of nail polish remover. Dudes
will recognize the smell of acetone as
the smell of paint thinner.
On a ketogenic diet, when there's a large
generation of acetone, someone can
develop that characteristic smell in
their breath and that's what we call
ketone breath.
The audio of this lesson was generously
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Bob Davodian of Taurean Mixing,
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All right this is how ketogenesis works.
In future lessons coming up shortly we'll
see the various benefits and pitfalls
that we can get from a ketogenic diet.
I hope you found this useful.
Signing off, this is Chris Masterjohn if
chrismasterjohphd.com. You've been
watching Masterclass with Masterjohn.
And I will see you in the next lesson.
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