Gluconeogenesis

Definition

The pathway that makes glucose from non-glucose precursors. (neo – new;
genesis – creation)

What starting materials are used?

The main ones are:

lactate – end product of lactate fermentation;

amino acids – from the breakdown of proteins;
glycerol – from the breakdown of fats.

Note that fatty acids, which form the main part of fat molecules, cannot be
converted to sugars in animals, though plants are able to do this.

Why do we need to make glucose?

Two main reasons.

1. When glucose is broken down by lactate fermentation the ATP yield is

very poor compared with that from glucose respiration (2 ATPs against
about 30 ATPs). A large part of this difference is because of the
amount of energy still locked up in lactate. To simply lose the lactate by
excretion would be a great waste of dietary resources. Instead the
lactate, produced largely by skeletal muscle is carried in the blood
stream to the liver, which converts it back to glucose. The glucose can
then be exported, via the blood stream, back to the muscle or other
tissues. This is known as the Cori cycle after Professors Cori (husband
and wife) who discovered it.

2. In conditions of starvation the body’s store of carbohydrate is quickly

depleted. Carbohydrate is stored in the liver as the polysaccharide,
glycogen, and only about 24 hours supply is kept there. Depletion of
the glycogen supply would lead to a fall in blood sugar. Some cells,
notably the nerve cells of the brain, are almost entirely dependent on a
supply of sugar from the blood. They don’t store their own
carbohydrate and they can’t metabolise fat. Clearly a fall in blood sugar
level would be very dangerous and the body must create new sugar to
maintain the blood concentration. This sugar is largely made from
amino acids, obtained by breaking down bodily protein, and from the
glycerol component of fats.

The gluconeogenetic pathway

We’ll start by considering the biosynthesis of glucose from lactate. We’ve

already studied lactate fermentation. One way of making glucose from lactate
would be simply to use exactly the same reactions, catalysed by exactly the
same enzymes, in the opposite direction. We can do this as long as all the
reactions of the lactate fermentation pathway are readily reversible. If you look
at the outline of the pathway given in the handout on glycolysis and glucose
fermentation, you’ll see that all the reactions have reversible arrows except for
three of them; those catalysed by hexokinase, phosphofructokinase and
pyruvate kinase. If we could find ways of bypassing these irreversible steps,
we could use all of the other fermentation reactions in the reversed direction
and we would have a biosynthetic pathway. We’ll look at each of these
irreversible steps in turn, to see why they are irreversible, and how we can
bypass them.

Pyruvate kinase reaction

This reaction is the conversion of phosphoenolpyruvate to pyruvate with the

generation of ATP from ADP:

PEP + ADP  Pyruvate + ATP + HEAT ENERGY

Finer detail of the structures of the intermediates is available on the glycolysis

handout so I’ve not included it in the equation above, but you’ll notice that I’ve
included another “product”: heat energy.

When PEP loses its phosphate, energy is released as the phosphate is

attached by a high-energy bond. This energy is used to make ATP and
“trapped” within that molecule. However, the energy released from PEP is
actually more than required to make ATP so the rest is lost as heat energy.
This reaction is not irreversible. All chemical reactions are reversible. But to
run it “backwards” would require the heat energy lost in the forward reaction to
be supplied. As we’ve seen before, heat can’t be used as an energy source in
biochemical reactions as they take place at constant temperature. To create
PEP from pyruvate, then, we need to find a way to use another source of
chemical energy, in addition to that made available by conversion of ATP to
ADP, which would be available in a straightforward reversal. The cell does
this by using two reactions to bypass the single reaction of pyruvate kinase,
each of which uses a high-energy compound. The first of these reactions is
catalysed by pyruvate carboxylase:

This reaction adds an extra carbon to the pyruvate from CO2, creating a new
carboxyl group. Creating the new C-C bond requires energy, which comes
from the breakdown of ATP to ADP. The energy from ATP is now “stored”
within the new C-C bond. Notice that the product, oxaloacetate, is the same
compound that is involved at the start of the TCA cycle. We’ll see the
importance of that later. Pyruvate carboxylase requires two co-factors:
magnesium ions and biotin, a small organic compound that acts as a carbon
dioxide carrier.

The second of our two reactions is catalysed by an enzyme called

phosphoenolpyruvate carboxykinase:

This reaction uses GTP as an energy source, and also a source of the
phosphate that’s added to create PEP. In addition the carbon added in the
previous reaction is released again, making a second parcel of energy,
originally from ATP in the last reaction, available to create the very high-
energy phosphate bond in PEP. This reaction also requires magnesium ions.

Between the two of them these reactions create PEP from pyruvate,
overcoming the energy difficulty by making use of two high energy
compounds.

Phosphofructokinase

Fructose 6-phosphate + ATP  Fructose 1,6-bisphosphate + ADP + HEAT

ENERGY

Notice again the addition of heat energy to the right of this equation. The
energy from the conversion of ATP to ADP is used to bond the phosphate of
ATP to the sugar, but ATP provides more energy than is required so the rest
is lost as heat. As in the last reaction, to reverse the reaction exactly would
require the presence of that heat energy to drive the reaction backwards. Heat
energy, which is not available. The energy would be needed in the reverse
reaction to create the ATP which provided it in the first place. We can convert
fructose 1,6-bisphosphate to fructose 6-phosphate without the need of an
input of heat energy if we don’t insist on using the lost phosphate to make
ATP. We can do this by simply removing the phosphate by hydrolysis.

Fructose 1,6-bisphosphate + H2O  Fructose 6-phosphate + Pi

Enzymes that remove phosphate groups by hydrolysis are called

phosphatases. This one is fructose 1,6-bisphosphatase.

Hexokinase

Glucose + ATP  Glucose 6-phosphate + ADP + HEAT ENERGY

The difficulty with reversing this reaction is identical to that discussed under
phosphofructokinase and the solution is also identical. The phosphate is
removed by hydrolysis using the enzyme glucose 6-phosphatase:

Glucose 6-phosphate + H2O  Glucose + Pi

Summary of gluconeogenesis

We now have a pathway to make glucose from lactate using the reversed
glycolysis/ fermentation reactions together with the by pass reactions
discussed above:

lactate to pyruvate by reversed lactate dehydrogenase;

pyruvate to PEP via oxaloacetate using the bypass enzymes;
PEP to fructose 1,6-bisphosphate by reversed glycolysis reactions;
fructose 1,6-bisphosphate to fructose 6-phosphate by the bypass
enzyme;
fructose 6-phosphate to glucose 6-phosphate by reversed glycolysis;
glucose 6-phosphate to glucose by the bypass enzyme.

Biosynthesis of glucose from amino acids

We haven’t studied amino acid metabolism, and we won’t in any detail in this
module, but all amino acids can be broken down by pathways that ultimately
feed into the TCA cycle. The TCA cycle is the final breakdown pathway for
amino acids as well as sugars. Because the twenty amino acids found in
proteins have quite varied structures, the pathways to break them down are
also quite varied, resulting in amino acids entering the TCA cycle at various
different points. We’ll look at that shortly.

Remember that in the gluconeogenesis bypass reactions we’ve just studied,

pyruvate was converted to PEP by firstly converting it to oxaloacetate (OAA).
It follows that OAA can be converted to glucose, and anything that can be
converted to OAA can therefore be converted to glucose.

The following table shows how amino acids that enter the TCA cycle at
different points can be converted to glucose.

Amino acid is Route to glucose

converted to:
Pyruvate Converted to OAA by pyruvate carboxylase, then to
glucose
-Ketoglutarate Converted to OAA by normal reactions of TCA cycle,
then to glucose
Succinyl-CoA Converted to OAA by normal reactions of TCA cycle,
then to glucose
Fumarate Converted to OAA by normal reactions of TCA cycle,
then to glucose
Oxaloacetate To glucose
Acetyl-CoA Cannot be converted to OAA, and no route to glucose
Perhaps the last item in the table needs a little more explanation. Some amino
acids are broken down to acetyl-CoA, which can enter the TCA cycle for
further degradation in the same way as acetyl-CoA from glucose. Remember,
from our discussion on the TCA cycle, that the cycle cannot create new OAA
molecules from acetyl-CoA. It follows that the acetyl-CoA from these amino
acids is not converted to OAA and, therefore, cannot be converted to glucose.
The only way around this problem would be if the acetyl-CoA could be
converted to pyruvate by the reverse of the pyruvate dehydrogenase reaction.
This reaction is irreversible and, in animals, there is no bypass process that
can avoid this irreversibility. Plant biochemistry is a little different and plants
do possess enzymes to carry out this bypass and are able to convert acetyl-
CoA to sugar.

Amino acids which can be converted to sugar are called glucogenic amino
acids. Ones that can’t are called ketogenic. Some of the larger amino acids
are broken into two parts during breakdown and one part is glucogenic and
the other part ketogenic. Nearly all protein amino acids are fully, or partially,
glucogenic; only two, leucine and lysine, are completely ketogenic.

Biosynthesis of glucose from fats

Fats are large molecules whose structure we’ll look at in more detail later on.
In simple terms, though, they are made up of a molecule of glycerol, a small
three carbon structure, and three molecules of fatty acid. Fatty acids are large
molecules, typically of 16 to 20 carbons in size so, obviously, the bulk of a fat
is fatty acid, with glycerol making up only a small component of it. In
catabolism of fats these components are separated and metabolized
separately so we’ll consider each of them in turn

Glycerol

Glycerol can be converted, by two steps, to dihydroxyacetone phosphate. I’ve

left out some of the detail and the enzyme names. You should be able to
complete these yourself without looking them up in a text book. DHAP is an
intermediate on the glycolysis/gluconeogenesis pathways so can either be
converted to pyruvate (glycolysis) for further degradation by the TCA cycle, or
it can go in the other direction (gluconeogenesis) and be converted to
glucose.

Fatty acids
These are broken down to acetyl-CoA so, like ketogenic amino acids, they
cannot be converted to glucose in animals, though plants are capable of doing
this.