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Glycogen - the glucose polymer

Glycogen is a branched polymer of glucose residues
glycogen serves as a buffer to maintain blood-glucose levels
Glycogen is not as reduced as fatty acids are and consequently not as energy rich.
Why do animals store any energy as glycogen? Why not as fatty acids?
Glycogen is an important fuel reserve for several reasons.
The controlled breakdown of glycogen and release of glucose increase the amount
of glucose

glucose is the only fuel used by the brain (except during prolonged starvation).
glucose from glycogen is readily mobilized and is therefore a good source of energy
for sudden activity.
Unlike fatty acids, the released glucose can provide energy in the absence of oxygen and
can thus supply energy for anaerobic activity.

The two major sites of glycogen storage are the liver and
skeletal muscle.
The concentration of glycogen is higher in the liver than in
muscle (10% versus 2% by weight), but more glycogen is stored in skeletal muscle
overall because of its much greater mass.

In the liver, glycogen synthesis and degradation are regulated

to maintain blood-glucose levels as required to meet the needs
of the organism as a whole.
In muscle, these processes are regulated to meet the energy
needs of the muscle itself.

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Most of glycogen are linked by a-1,4-glycosidic bonds. Branches

at about every tenth residue are created by a-1,6-glycosidic bonds

The glycogen-branching enzyme, amylo (14) to (16)

transglycosylase, catalyzes the transfer of a terminal fragment of
6-7 glucose residues from a nonreducing end to the C-6 hydroxyl
group of a glucose residue deeper into the interior of the glycogen
molecule

Electron Micrograph of a Liver Cell.

Glycogen is present in the cytosol in the form of
granules ranging in diameter from 10 to 40 nm

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Glycogen Is Synthesized and Degraded by Different Pathways

Separate pathways afford much greater flexibility, both in

energetics and in control.
Like glycolysis and gluconeogenesis.

Glycogen synthesis

Glycogen synthesis requires an activated form of glucose, uridine

diphosphate glucose (UDP-glucose).

UDPglucose
pyrophosphorylase.
PPi

This reaction is readily reversible. However, pyrophosphate is rapidly hydrolyzed in vivo to

orthophosphate by an inorganic pyrophosphatase. The essentially irreversible hydrolysis of
pyrophosphate drives the synthesis of UDP-glucose.
2 Pi

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UDP

The activated glucosyl unit of UDPglucose is transferred to the hydroxyl group at a C-4 terminus
of glycogen to form an a-1,4-glycosidic linkage. In elongation, UDP is displaced by the terminal
hydroxyl group of the growing glycogen molecule. This reaction is catalyzed by glycogen
synthase, the key regulatory enzyme in glycogen synthesis.

Glycogen Is an Efficient Storage Form of Glucose

One ATP is hydrolyed incorporating glucose 6-phosphate into glycogen. The energy yield from the
breakdown of glycogen is highly efficient. About 90% of the residues are phosphorolytically cleaved to
glucose 1-phosphate, which is converted at no cost into glucose 6-phosphate. The other 10% are branch
residues, which are hydrolytically cleaved. One molecule of ATP is then used to phosphorylate each of
these glucose molecules to glucose 6-phosphate. The complete oxidation of glucose 6-phosphate yields
about 31 molecules of ATP, and storage consumes slightly more than one molecule of ATP per molecule of
glucose 6-phosphate; so the overall efficiency of storage is nearly 97%.

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A Branching Enzyme Forms a-1,6 Linkages

(by the enzyme Glycogenin)

A branch is created by the breaking of an a-1,4 link and the formation of an a-1,6 link:
this reaction is different from debranching. A block of residues, typically 7 in number, is
transferred to a more interior site. The block of 7 or so residues must
(a) include the nonreducing terminus
(b) come from a chain at least 11 residues long.
(c) the new branch point must be at least 4 residues away from a preexisting one.

Branching is important because:

(a) It increases the solubility of glycogen.
(b) creates a large number of terminal residues, the sites of action of glycogen phosphorylase
and synthase.
Thus, branching increases the rate of glycogen synthesis and degradation.

Glycogen branching requires a single transferase activity.

Glycogen debranching requires two enzyme activities: transferase and an a-1,6 glucosidase.

Glycogenin

G=glycogenin.

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Glycogen degradation

The efficient breakdown of glycogen to provide glucose 6-phosphate

for further metabolism requires four enzyme activities:
1) degrades glycogen
2+3) remodel glycogen so that it remains a substrate for degradation
4) converts the product of glycogen breakdown into a form suitable
for further metabolism.

Glycogen degradation

Glycogen phosphorylase, the key enzyme in glycogen breakdown,

cleaves its substrate by the addition of orthophosphate (Pi) to yield
glucose 1-phosphate. The cleavage of a bond by the addition of
orthophosphate is referred to as phosphorolysis.

Phosphorylase catalyzes the sequential removal of glycosyl residues from the nonreducing ends
of the glycogen molecule (the ends with a free 4-OH groups). Orthophosphate splits the
glycosidic linkage between C-1 of the terminal residue and C-4 of the adjacent one.

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Glycogen is phosphorylated to be broken down.

Why not hydrolyses?

The glucose released is already phosphorylated, ready to be

used by other enzymes.
The p-glucose is (-) charged, therefore cannot defuse out of the
cell, or be transported by GLUT.

The a-1,6-glycosidic bonds at the branch points are not susceptible to

cleavage by phosphorylase. Indeed, phosphorylase stops cleaving a-
1,4 linkages when it reaches a terminal residue four residues away
from a branch point. (about 1 in 10 residues is branched)

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First, a-1,4-glycosidic bonds

on each branch are cleaved
by phosphorylase, leaving
four residues along each
branch. The transferase shifts
a block of three glycosyl
residues from one outer
branch to the other. In this
reaction, the a-1,4-glycosidic
link between the blue and the
green residues is broken and
a new a-1,4
link between the blue and the
yellow residues is formed.
The green residue is then
removed by a-1,6-glucosidase,
leaving a linear chain with all
a-1,4 linkages, suitable for
further cleavage by The transferase shifts a block of three glycosyl residues from
phosphorylase. one outer branch to the other

Phosphoglucomutase Converts Glucose 1-phosphate into

Glucose 6-phosphate

The phosphoryl group is transferred from the serine residue to the C-6 hydroxyl group
of glucose 1-phosphate to form glucose 1,6-bisphosphate. The C-1 phosphoryl group of
this intermediate is then shuttled to the same serine residue, resulting in the formation
of glucose 6-phosphate and the regeneration of the phosphoenzyme.
NOTE- same mechanism like (2,3-BPG) in the interconversion of 2-phosphoglycerate and 3-
phosphoglycerate in glycolysis. A phosphoenzyme intermediate participates in both reactions.

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Liver Contains Glucose 6-phosphatase, a Hydrolytic Enzyme

Absent from Muscle (and absent from most other tissues)

glucose 6-phosphatase is located on the lumenal side of the smooth ER membrane (the
same enzyme that releases free glucose at the conclusion of gluconeogenesis).
Meaning that glucose 6-phosphate is transported into the ER; glucose and orthophosphate
formed by hydrolysis are then shuttled back into the cytosol

The absence of glucose 6-phosphatase in muscle ensures that G6P derived from
glycogen remains within the cell for energy transformation.

Note-glucose is not a major fuel for the liver.

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Glycogen Phosphorylase

Regulation - allosteric
- phosphorylation

Distribution - muscle
- liver

We will examine the differences in the control of glycogen

metabolism in two tissues: skeletal muscle and liver.

The transition from the T state (phosphorylase b) to the R state (phosphorylase a) entails a 10-degree
rotation around the twofold axis of the dimer. This transition is associated with structural changes in a
helices that move a loop out of the active site of each subunit. Thus, the T state is less active because the
catalytic site is partly blocked.

The equilibrium for phosphorylase a favors the R state whereas the equilibrium for phosphorylase b favors the T state.
Phosphorylase a is phosphorylated on serine 14 of each subunit. This modification favors the structure of the more active R state. One
subunit is shown in white, with helices and loops important for regulation shown in blue and red. The other subunit is shown in yellow,
with the regulatory structures shown in orange and green. Phosphorylase b is not phosphorylated and exists predominantly in the T
state.

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Mainly in the Mainly in the

muscle liver

active

Both phosphorylase b and phosphorylase a

exist as equilibria between an active R state
and a less-active T state. Phosphorylase b is
usually inactive because the equilibrium
favors the T state. Phosphorylase a is usually
active because the equilibrium favors the R
state. Regulatory structures are shown in blue
and green.

inactive Phosphorylase a and phosphorylase b differ by a

single phosphoryl group in each subunit.
Phosphorylase b is converted into phosphorylase
a when it is phosphorylated at a single serine
residue (serine 14) in each subunit.

Muscle Phosphorylase Is Regulated by the Intracellular

Energy Charge

Muscle phosphorylase b is active only in the presence of high concentrations of AMP, which
binds to a nucleotide binding site and stabilizes the conformation of phosphorylase b in the R state.
ATP acts as a negative allosteric effector by competing with AMP and so favors the T state. Thus,
the transition of phosphorylase b between the T and the R state is controlled by the energy charge of
the muscle cell. Glucose 6-phosphate also favors the T state of phosphorylase b, an example of
feedback inhibition.

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Under most physiological conditions, phosphorylase b is inactive

because of the inhibitory effects of ATP and G6P.

phosphorylase a is fully active, regardless of the levels of AMP, ATP,

and G6P.

In resting muscle, nearly all the enzyme is in the inactive b form.

During exercise AMP increases activation of phosphorylase b.

Exercise hormone release, ba.

In human beings, liver phosphorylase and muscle phosphorylase

isozymes are approximately 90% identical in amino acid sequence.

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In contrast with the muscle enzyme, liver phosphorylase a but not b

exhibits the most responsive T-to-R transition.

The binding of glucose shifts the allosteric equilibrium of the a form

from the R to the T state, deactivating the enzyme.

Unlike the enzyme in muscle, the liver phosphorylase is insensitive to

regulation by AMP because the liver does not undergo the dramatic
changes in energy charge seen in a contracting muscle.

This is an example of the use of isozymic forms of the same enzyme

to establish the tissue-specific biochemical properties of muscle and
the liver.

Activation of Phosphorylase Kinase

Phosphorylase kinase is activated by hormones that lead to the phosphorylation of the b subunit and by
Ca2+ binding of the d subunit. Both types of stimulation are required for maximal enzyme activity.

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Phosphorylase Kinase Is Activated by Phosphorylation and Calcium Ions

phosphorylase kinase, a very large protein with a subunit composition in skeletal muscle
of (a b g d) and a mass of 1200 kd.
The catalytic activity resides in the g subunit, whereas the other subunits serve a
regulatory function.
This kinase is under dual control. Like its own substrate, phosphorylase kinase is
regulated by phosphorylation: the kinase is converted from a low-activity form into a
high-activity one by phosphorylation of its b subunit.
The enzyme catalyzing the activation of phosphorylase kinase is protein kinase A (PKA),
which is switched on by a second messenger, cyclic AMP.
Hormones such as epinephrine induce the breakdown of glycogen by activating a cyclic
AMP cascade.
Phosphorylase kinase can also be partly activated by Ca2+ levels of the order of 1 mM. Its
d subunit is calmodulin, (a calcium sensor that stimulates many enzymes in eukaryotes).
This mode of activation of the kinase is important in muscle, where contraction is triggered by the release of Ca2+ from the SR.
Phosphorylase kinase attains maximal activity only after both phosphorylation of the b
subunit and activation of the d subunit by Ca2+ binding.

Hormonal control

Glucagon (liver) and epinephrine (muscle) trigger the breakdown of glycogen.

Epinephrine markedly stimulates glycogen breakdown in muscle and, to a
lesser extent, in the liver.

Muscular activity leads to the release of epinephrine (adrenaline), from

the adrenal
The liver is more responsive to glucagon,

Epinephrine is derived from tyrosine

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epinephrine and glucagon bind to specific 7TM receptors in the plasma membranes of muscle (b-adrenergic receptor) and liver
cells (glucagon receptor).

cAMP binds to the regulatory subunits of PKA,

which then dissociate from the catalytic subunits.
The free catalytic subunits are active.

Protein kinase A phosphorylates the b subunit of

phosphorylase kinase,

glycogen

Glycogen Synthase Is the Key Regulatory Enzyme in Glycogen

Synthesis
Glycogen synthase is phosphorylated at multiple sites by protein
kinase A and several other kinases. The resulting alteration of the
charges in the protein lead to its inactivation.

Phosphorylation has opposite effects on the enzymatic activities of

glycogen synthase and phosphorylase:
Phosphorylation converts the active a form of the synthase into a
usually inactive b form. The phosphorylated b form requires a high
level of the allosteric activator G6P for activity,
whereas the a form is active whether or not G6P is present.

Charge before phosphorylation

After phosphorylation
Inactive b form

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Glycogen Breakdown and Synthesis Are Reciprocally Regulated

by a hormone-triggered cAMP cascade acting through protein kinase A

Red=inactive

In addition to phosphorylating and activating phosphorylase kinase, protein kinase A adds

a phosphoryl group to glycogen synthase, which leads to a decrease in enzymatic
activity. This important control mechanism prevents glycogen from being synthesized at
the same time that it is being broken down.

Protein Phosphatase 1 (PP1) Reverses the Regulatory Effects of Kinases on Glycogen

Metabolism

PP1 inactivates phosphorylase kinase and phosphorylase a by dephosphorylating these enzymes.

PP1 decreases the rate of glycogen breakdown:

1) reverses the effects of the phosphorylation cascade (as stated above).
2) removes the phosphoryl group from glycogen synthase b to convert it into the much more
active a form.

The complete complex of PP1 consists of three components:

1) PP1 itself, a 37-kd catalytic subunit;
2)123-kd RGl subunit that confers high affinity for glycogen;
3) inhibitor 1, a small regulatory subunit that, when phosphorylated, inhibits PP1.
The importance of the RGl subunit is that it brings PP1, which is active only when
associated with glycogen molecules, into proximity with its substrates.

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How is the phosphatase activity of PP1 itself regulated?

When glycogen degradation is predominant, PKA is active. Two components

of PP1 are themselves substrates for protein kinase A.
Phosphorylation of the RGl component by protein kinase A prevents RGl from
binding the catalytic subunit of PP1.
Consequently, activation of the cAMP cascade leads to the inactivation of PP1
because it can no longer bind its substrates.
Phosphorylation of inhibitor 1 by protein kinase A blocks catalysis by PP1.
Thus, when glycogen degradation is switched on by cAMP, the accompanying
phosphorylation of inhibitor 1 keeps phosphorylase in the active a form and
glycogen synthase in the inactive b form.
The epinephrine-induced phosphorylation of the RGl subunit and inhibitor 1
are complementary devices for sustaining glycogen degradation.

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Insulin Stimulates Glycogen Synthesis by Activating PP 1

The binding of insulin to its receptor leads to the activation of an insulin-sensitive

protein kinase that phosphorylates the RGl subunit of PP1 at a site different from that
modified by protein kinase A. This phosphorylation leads to the association of the RGl
subunit with PP1 and the glycogen molecule. The consequent dephosphorylation of
glycogen synthase, phosphorylase kinase, and phosphorylase promotes glycogen
synthesis and blocks its degradation.
Once again we see that glycogen synthesis and breakdown are coordinately controlled.

Blood Glucose Regulates Liver Glycogen Metabolism

The infusion of glucose into the bloodstream leads to the inactivation of

phosphorylase, followed by the activation of glycogen synthase, in the liver.

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The binding of glucose to phosphorylase a shifts its allosteric equilibrium from

the active R form to the inactive T form. This conformational change renders
the phosphoryl group on serine 14 a substrate for protein phosphatase 1.
It is significant that PP1 binds tightly to phosphorylase a but acts catalytically
only when glucose induces the transition to the T form.
The R T transition of muscle phosphorylase a is unaffected by glucose and
is thus unaffected by the rise in blood-glucose levels

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Glycogen-Storage Diseases

Edgar von Gierke described the first glycogen-storage disease in 1929. A patient with
this disease has a huge abdomen caused by a massive enlargement of the liver. There is
a pronounced hypoglycemia between meals. Furthermore, the blood-glucose level does
not rise on administration of epinephrine and glucagon. An infant with this
glycogen-storage disease may have convulsions because of the low blood-glucose
level. The enzymatic defect in von Gierke disease was elucidated in 1952 by Carl and
Gerty Cori. They found that glucose 6- phosphatase is missing from the liver of a
patient with this disease. This was the first demonstration of an inherited
deficiency of a liver enzyme. The liver glycogen is normal in structure but present in
abnormally large amounts. The absence of glucose 6-phosphatase in the liver causes
hypoglycemia because glucose cannot be formed from glucose 6-
phosphate. This phosphorylated sugar does not leave the liver, because it cannot cross
the plasma membrane. The presence of excess glucose 6-phosphate triggers an increase
in glycolysis in the liver, leading to a high level of lactate
and pyruvate in the blood. Patients who have von Gierke disease also have an
increased dependence on fat metabolism. This disease can also be produced by a
mutation in the gene that encodes the glucose 6-phosphate transporter. Recall
that glucose 6-phosphate must be transported into the lumen of the endoplasmic
reticulum to be hydrolyzed by phosphatase. Mutations in the other three essential
proteins of this system can likewise lead to von Gierke disease.
Seven other glycogen-storage diseases have been characterized

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