Berg • Tymoczko • Stryer

Biochemistry
Sixth Edition

Chapter 21:
Glycogen Metabolism

Copyright © 2007 by W. H. Freeman and Company

Glycogen Metabolism
 OUTLINE
• Glycogen breakdown requires the interplay of several enzymes
• Phosphorylase is regulated by allosteric interactions and reversible
phosphorylation
• Epinephrine and glucagon signal the need for glycogen
breakdown
• Glycogen is synthesized and degraded by different pathways
• Glycogen breakdown and synthesis are reciprocally regulated

 Glycogen synthesis: glycogenesis

 Degradation of glycogen: glycogenolysis.
Glycogen

 Glycogen is a highly branched, very large polymer of

glyc mols linked  1 4
 Branches arise by  1 6 at about every 8-10th residue
 It is found in the cytosol.
 It is the storage form of Glc.
 Liver and muscle are the major sites for the storage of
glycogen.
 Surface: nonreducing ends
At the core:
glycogenin

Degradation: takes place at the surface!

Degradation of Glycogen
 Is not a simple reversal of the synthetic pathway
 Other enzymes involved
Glycogen  G-1-P
 Shortening of chains:
– Glycogen phosphorylase (-1  4)
– It is an exoglucosidase
– Degrades the gly. chains at their non-reducing ends
until four glucosyl units remain on each chain before
the branch point
– The resulting structure  a limit dextrin
– Phosphorylase cannot degrade this any further!
Degradation of Glycogen continued

Removal of branches:

 Branches are removed through two enzymatic activities

of the debranching enzyme
a. Glucosyl 4:4 transferase removes the outer 3 of 4 glucosyl
residues
b. Single glucose residue attached in an  16 linkage is then
removed by the -amylo (1:6) glucosidase activity of the
debranching enzyme, releasing free glucose
Regulation phosphorylase
Regulation of glycogen metabolism is different in muscle
and liver.
 In muscle, the end served by glycolysis is ATP
production and the rate of glycolysis increases as
muscle works more, demanding more ATP.
 The liver has a different role in whole-body metabolism
and glucose metabolism in the liver is different. The
liver makes sure that glucose level is constant in the
blood, producing and exporting Glc.
 Regulation of Glycogen Breakdown
Glycogen represents the most immediately available large-scale
source of metabolic energy. Therefore, it is important that
animals be able to activate glycogen mobilization very rapidly.
 Glycogen breakdown is an hormone-controlled process.

Structure of glycogen phosphorylase:

 Dimer; exists in two forms.
– Active phosphorylase a
– Inactive phosphorylase b
 Activation  by phosphorylase kinase
 Deactivation  phosphorylase phosphatase

Control of phosphorylase activity:

 Phosphorylase kinase is activated by c-AMP protein kinase
 Muscle phosphorylase
 In muscle, phosphorylase has 2 forms.
1. Phosphorylase a:
– active form
– 2 subunits, in each Ser residue at position 14 is Plated (Phosphorylase kinase
does this)
2. Phosphorylase b:
– inactive form
– in resting muscle, all enzyme is its inactive form
– structurally identical except that Ser residues are not Plated. It is active when
AMP is high! It is inactive when ATP and Glc 6-P are high! So, muscle
phosphorylase b is active only when the energy charge of the muscle is low.

 The rate of glycogen breakdown is due to the a/b which is controlled by

hormones especially by epinephrine.
 Phosphorylase a  phosphorylase b by dephosphorylation catalyzed by
phosphorylase a phosphatase.
Muscle phosphorylase
Phosphorylase b
Control of
Glycogen Phosphorylase in Muscle
Muscle phosphorylase regulation

 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.
 AMP dependency of phosphorylase b
 Phosphorylase a is AMP-independent
 Phosphorylase b is AMP-dependent.
– The stimulation of phosphorylase b by AMP can be prevented by high
ATP concentrations.
 AMP binds its allosteric site and stabilizes the conformation of phosphyrylase
b in the R state.
 ATP acts as a negative allosteric modulator by competing with AMP and so
favors the T state.

Intensive exercise AMP/ATP goes up, Phosphorylase b (active)

In resting muscle AMP/ATP goes down, Phosphorylase b (inactive)

 Exercise will also result in hormone release (epinephrine) that

generates the phosphorylated a form of the enzyme.
Liver phosphorylase
 Liver phosphorylase and muscle phosphorylase are 90%
identical in amino acid sequence.
 Liver phosphorylase a, but not b, has the most responsive
T-to-R transition.
 The binding of Glc shifts the allosteric equilibrium of the
a form from the R to the T state, deactivating the enzyme.
 Why would Glc function as a negative regulator of liver
phosphorylase a?
– When there is plenty of Glc, no need to breakdown liver
glycogen!
 Liver Glycogen phosphorylase
is regulated by hormones and blood glucose.
 Liver glycogen has a different role in our system.
– When blood glucose is low (lower than 4-5 mM)
Glycogen  Glc-1-P  Glc-6-P  Glc
– So, when blood glucose is low, glucose is released into the blood
stream and carried to the needy tissues.
 Glycogen phosphorylase of liver is under hormonal control.
– Glucagon is the hormone.
– When glucose is low, glucagon is released.
– Liver phosphorylase is allosterically regulated by Glc not AMP.
• When Glc is high in the blood, it enters hepatocytes and binds regulatory
sites of the enzyme, causing conformational changes (favoring the T
state).
• Therefore, glycogen phosphorylase is a glucose sensor.
• When Glc is high, it stops its own FORMATION.
 Phosphorylase kinase
is activated by phosphorylation and calcium ions
The phosphorylase enzyme.
 Has a fully active form and an inactive form
 Has a mass of 1200 kd
 Consist of 4 subunits (abgd)
– The subunit g is the source of catalytic activity.
– The other subunits are regulatory subunits.
 Is under dual control
1. Regulated by phosphorylation
– The b subunit is phosphorylated by cAMP dependent PKA).
2. Partly activated by calcium levels of the order of 1 mM.
– The d subunit is calmodulin, a calsium sensor that stimulates many enzymes.
 Phosphorylase kinase has the highest activity only after both the
phosphorylation of the b subunit and the activation of the d subunit by Ca
binding.
Epi and Glucagon signal the need for glycogen
breakdown
 PKA  activates phosphorylase kinase
 Glycogen phosphorylase activated
 Glc 1-P is made
 What activates PKA?
– HORMONES
• Glucagon
• Epinephrine
 Signal transduction
– Epi
– GTP-bound G proteins
– Increased cAMP
– PKA increases
 cAMP amplifies the effects of hormones
What shuts off glycogen breakdown?
 This signal transduction pathway is shut down
by the same pathway.

 How?
• GTP is deactivated by its inherent GTPase
activity
• cAMP is converted to AMP (not a second
messenger) by phosphodiesterase enzyme.
Steps in Glycogen Synthesis

A. UDPG synthesis:
G-6-P  G-1-P
G-1-P + UTP  UDPG + PPi

B. A primer is required for glycogen synthesis:

1. A fragment of glycogen can serve as a primer in cells whose
glycogen stores are not totally depleted.
2. If a glycogen fragment is not present, glycogenin, a
glycosyltransferase, serves as the primer.
Steps in Glycogen Synthesis continued

C. Elongation of glycogen chains

– Glucose is transferred from UDPG to the non-reducing end of the
growing chain.
• New glycosidic bond between C-1 of the activated sugar and C-4 of the
accepting glucosyl residue
• Enzyme: glycogen synthase
– If no other enzyme acts on the chain, the resulting structure is a linear
molecule of glucosyl residues attached by  1-4.
• Such a compound, called amylose, is found in fruits.
– The UDP released when the new bond is made can be convert back.
UDP + ATP  UTP + ADP
Steps in Glycogen Synthesis continued

D. Creating branches in glycogen:

– Amylose  unbranched
Glygogen  branches (~8)
 The branches are made through the “branching enzyme”, glucosyl 4:6
transferase (amylo  1, 4 -  1,6 transglycosylase
– This enzyme transfers 5 to 8 glucosyl residues from the non-
reducing end to another residue by an  1,6 link.
– Further elongation
– Branches have two important functions
a) increases the solubility of the glycogen molecule.
b) The number of non-reducing ends to which new glucosyl residues can
be added and thereby greatly accelerating the rate at which glycongen
synthesis and degradation can occur.
Initiation of Glycogenesis By Glycogenin
Summary of the synthesis
 UDP-Glucose synthesis UDP-glucose phosphorylase
 A primer is required for glycogen synthesis (glycogenin or
a fragment of glycogen)
 Glc units are added to the either the existing glycogen
chains or glycogenin (enzyme glycogen synthase).
• C-4 is the non-reducing end of glycogen chain. New glucose
molecules are always added to this non-reducing terminus.
 Elongation of glucose chains
 Creating branches in glycogen (enzyme transferase)
 Branches have 2 functions:
1. Increase the solubility of the glycogen molecule
2. Increase the rate of glycogen synthesis
How Is Glycogen Synthesis Regulated?
 Glucagon and Epi promote glycogenolysis, at the same
time they inhibit glycogen synthesis.
– Both effects are mediated by cAMP and cAMP dependent
protein kinase.
– Regulated enzyme: glycogen synthase
• a form: active (not phosphorylated)
• b form: inactive (phosphorylated)
• PKA and other kinases phosphorylate the enzyme.
– Protein kinase (Ser – phosphorylated)
 Steps after the binding of the hormones:
– Epi  liver cell recep.
– Adenylate cyclase activity
– cAMP
– cAMP  Pkinase which phosphorylates and inactivates
glycogen synthase
 Coordinate Control of Glycogen Breakdown and Synthesis by cAMP Cascades

Glycogen
degradation
Glycogen synthesis

Inactive forms are shown in red, active forms are shown in green.
 Glycogen
degradation

Inactive forms are shown in red,

active forms are shown in green.
 Glycogen
synthesis

Inactive forms are shown in red,

active forms are shown in green.
Breakdown and synthesis are reciprocally
regulated
 Hormone -triggered cAMP cascade acting through PKA
 Glycogen breakdown and synthesis are reciprocally
regulated.
• Phosphorylase kinase also inactivates glycogen synthase.
 PP1(protein phosphatase 1) reverses the regulatory effects
of glycogen metabolism.
– PKA action is reversed by phosphatases
– PP1 inactivates phosphorylase kinase and phosphorylase a by
dephosphorylating these enzymes.
– PP1 also removes P groups from the glycogen synthase b to the
glycogen synthase a form (more active)
 PP1 has 3 components:
– P1
– Rgl
– I
 How is phosphatase activity of PP1 regulated?
– Rgl phosphorylation by PKA prevents its binding to PP1, therefore
activation of cAMP cascade leads to the inactivation of PP1 because
it cannot bind to its substrate.
– Phosphorylation of inhibitor 1 by protein kinase A blocks catalysis
by PP1.
 Thus, Epi increases glycogen breakdown by making
phosphorylase a and decreases glycogen synthesis by
making inactive phosphatase.
Insulin stimulates glycogen synthesis
by activating protein phosphatase 1

 When blood glucose is high, insulin is stimulated.

 Activated insulin-sensitive protein kinase makes activated
protein phosphatase

The consequent dephosphorylation of glycogen

synthase, phosphorylase kinase, and phosphorylase
promotes glycogen synthesis and blocks its
degradation!
Phosphorylation of the enzymes is regulated
by hormones

 Phosphorylated groups can be removed by

phosphatases; therefore, the action of phosphatases
always opposes kinases.
– If kinases activity is greater than activity of phosphatase, the
enzyme is in the phosphorylated mode.

 Insulin, Glucagon, and Epi are three important

hormones which affect glycogen metabolism!
Glycogen metabolism in the
liver regulates the blood-glucose
level

 After a carbohydrate-rich meal blood glucose increases.

 Insulin is the primary signal for glycogen synthesis.
 Blood glucose level 80-120 mg/dL (4.4-6.7 mM)
 The liver senses the concentration of blood glucose and
either release or takes up glucose.

 Glucose infusion changes the enzymes involved in

glycogen metabolism
Glucose regulation of glycogen metabolism

 Phosphorylase a is the glucose sensor in liver cells

 Glucose is high
 Binding of Glc converts R T
 PP1 is released
 Inactivation of glycogen breakdown and the activation of
glycogen synthesis take place..
Glucose regulation of glycogen metabolism

[Glycogen breakdown inhibited]

[Glycogen synthesis favored]

 Summary of the Regulation of
Glycogen Synthesis and Degradation
 Synthesis and degradation are regulated by the same
hormonal signals!
 An increase in insulin stimulates glycogen synthesis
 An increase in glucagon or Epi stimulates glycogen degradation
 cAMP production increases in response to the release ofEpi and
glucogon
 cAMP production decreases in the presence of insulin!
 Key enzymes are phophorylated by a family of kinases,
some of which are cAMP dependent.
 Phosphorylation of an enzyme causes 3D change that
affects the active site. It may either increase or decrease
its activity depending on the type of enzyme.
Glycogen storage diseases

 Glycogen metabolism is a finely controlled system.

– It is not surprising that genetically determined enzyme deficiencies
result in disease state.
– Genetic diseases are in fact valuable research tools for us.
 There are 8 glycogen storage diseases but we will only
cover Type I and Type V
– Type I
• Von Gierke Disease
• Glc 6-phosphatase is missing.
– Type V
• McArdle Disease
• Phosphorylase is missing.
Problem 31.
Purified from two samples of human liver, glycogen was either treated or not treated
With a-amylase and subsequently analyzed by SDS-PAGE
Berg • Tymoczko • Stryer and western blotting with
the use of antibodies to glycogenin. The results are presented in next slide.

1. Why are no proteins without amylase?

Biochemistry
Glycogen is too large to enter the gel. Antibody to glycogenin was used so we only see
Glycogenin by western blotting.

Seventh Edition
2. What is the reason using amylase?
Amylase degrades glycogen, releases glycogenin
3. Why don’t we see other proteins?
CHAPTER
1. Antibody against glycogenin 21 P-lase, glycogen synthase,
was used (glycogen
Protein phosphatase-1 can be seen if we used Abs for them)
Glygogen Metabolism

Copyright © 2012 by W. H. Freeman and Company

Problem for students. The gene for glycogenin was transfected into a cell line that normally sto
small amounts of glycogen. The cells were then manipulated according to the following
protocol, and glycogen was isolated and analyzed by SDS-PAGE and western blotting by using
An antibody to glycogenin with and without amylase treatment. The results are presented in the
Berg • Tymoczko • Stryer
next slide. The protocol: Cells cultured in growth medium and 25 mM glucose (lane1) were
Switched to medium containing no glucose for 24 hours (lane 2). Glucose-starved cells were
refed with medium containing 25 mM glucose for 1 hour (lane 3) or 3 hours (lane 4). Samples
(12 microg of protein) were either treated or not treated with amylase, before being loaded on
the gel.
Biochemistry
a. Why did the western analysis produce a “smear”-that is, the high molecular-weight
staining in lane 1(-)? Seventh Edition
b. What is the significance of the decrease in HMW-staining in lane 2(-)?
c. What is the significance of the difference between lanes 2(-) and 3(-)?
d. Suggest a plausible reason why there is essentially no difference between lanes 3(-)
and 4(-)? CHAPTER 21
e. Why are the bands at 66 kd the same in the lanes treated with amylase, despite the
fact that the cells Glygogen Metabolism
were treated differently?

Copyright © 2012 by W. H. Freeman and Company