Regulation of Glycogen
Metabolism
•
•
•
•
•
•
Protein Kinases
Protein Phosphatases
cAMP
G proteins
Calcitonin
Insulin, glucagon, and epinephrine
Biochemical Definitions
Consider
A
B
C
1. Equilibrium: (Kinetics) When the rate of the
forward reaction matches the rate of the reverse
reaction. Does not specify quantity at equilibrium.
2. Equilibrium constant: (Thermodynamics) The
quantity [B]/[A] at equilibrium. Does not specify rate.
3. Flux: Net carbon flow in one direction
4. Steady-state: A dynamic condition that allows flux
without changing the concentration of components in
the pathway.
Connecting Kinetics with Thermodynamics
Chemistry
o
∆G
= RT ln Keq
Biochemistry
pH = 7.0 (10-7 M [H+] )
∆G = RT ln Keq
o’
Practically Speaking…..
1. A negative ∆Go’ means [P] > [S] at equilbrium.
A
B
’
o
∆G = RT ln Keq
[B]/[A]
2
10
100
1,000
10,000
100,000
∆Go’ (kJ/mol)
-1.8
-5.9
-11.9
-17.8
-23.7
-29.7
Steady-State vs Equilibrium Reactions
Rule: An equilibrium reaction occurs in a closed system
whereby two components, a reactant and a product, achieve
a constancy of concentration based on chemical potentials
A
B
Rule: A steady-state reaction occurs in an open system
whereby two or more components achieve a constancy of
concentration based on similar rates of components entering
and leaving the system
Y
A
B
Z
See Strategies
p 163-167
Representing a Steady-State
k1
A
k2
When
k1 = k2
A is constant
When
k 1 < k2
A decreases
When
k 1 > k2
A increases
Basics of Metabolic Homeostasis
Rule: A shift away from a dynamic steady-state
evokes factors to restore the steady-state.
Rule: Restoring steady-state requires modulating the
activity of a rate-controlling enzyme(s) in the pathway
Enzyme activity can be modulated by:
1. Covalent modification
2. Changes in pathway [S] or enzyme cofactors
3. Allosterism (Vmax or Km)
4. Hormonal intervention
5. Enzyme turnover
Regulation of Carbon Flux
• Net carbon flux through an individual step in a
pathway is defined as the difference between the
forward and reverse reaction velocities
J = VF - VR (these are rates of change)
• At equilibrium: VF= VR; J = 0; i.e., there is no net
flux even though VF and VR could be large
• When VF >> VR, J = VF
• At steady state, J = k (constant)
• At steady state, J depends on the rate determining
(slowest) step in the pathway
A
B
C
D
Rate-determining
Textbook p591
Meaning of Flux
Glucose
J=0
VF > VR
Rate-controlling
Step
Lactate
Flux varies with equilibrium
position
AA
A
A
A
A
BB B
1. If [A] = 100mM; [B] = 10mM, an increase in [A] by 10 mM
would increase flux would by 10%. (from 10:1 to 11:1)
2. If [A] = 20mM; [B] = 10mM, an increase in [A] by 10 mM
would increase flux by 33% . (2:1 to 3:1)
3. If [A] = 10mM; [B] = 10mM, an increase in [A] by 10 mM
would increase flux by 100% (1:1 to 2:1).
Rule: Regulators with the lowest steady-state
concentration have the greatest impact on regulation
AMP (0.1 mM)
Adenylate Kinase
ATP + X
5
0.5
ATP (5 mM)
ADP (1 mM)
2ADP = AMP + ATP
ATP + X~P +
4.5
0.5
ATP + AMP
2ADP
0.5
0.5
0.5
Final Talley:
Before
After
ATP
5 mM
5 mM
ADP
1 mM
1 mM
AMP
0.1 mM
0.6 mM
ADP
0.5
No change
No change
6 times
Why are there 2 enzymes in glycogen
metabolism?
Glycogen
Glycogenase
Glycogen
Phosphorylase
Glucose 1-PO4
How can glycogen
synthesis and degradation
be tuned to the needs
of the cell? Only glycogen
or glucose 1-PO4 concentration
can affect VF or VR
Stimulating or inhibiting the
enzyme cannot control direction
of flux
Synthase
Glucose 1-PO4
UDP-Glucose
Because separate enzymes
control synthesis and
degradation, VF and VR
can vary depending on the
enzyme that controls the
direction. This opens the
way to allosteric and covalent
control of glycogen
cAMP as a Regulator of Glycogen
ADP
Phos a
Syn b
Pkinase
Ptase
Phos b
PO4
Pkinase
Ptase
ATP
PO4
ADP
Syn a
ATP
Protein Kinase targets of cAPK
cAMP Protein kinase (cAPK)
Adenyl cyclase
Glucagon
Epinephrine
Textbook p586
ATP
cAMP
cAMP Protein kinase (cAPK)
R
R
C
C

+ 4 cAMP
 

+
C
P
C

2ATP
cAMP
R cAMP
cAPK
Calmodulin
Inactive Kinase
cAMP
R cAMP
C
2ADP
P
 

Active Kinase
Catalytic site
Example would be
phosphorylase b kinase
Now its ready to
phosphorylate
phosphorylase b
Inactivation of Glycogen Synthase
Priming
phosphate
Inactive
P P
H2O
P
P
B
PP1
Glycogen
Synthase
3Pi
A
ADP
Casein kinase II
(primer)
ATP
Glycogen synthase
kinase 3
(GSK3)
Phosphoprotein Phosphatase-1 (PP1)
Phos a
ADP
Pkinase
PP1
Phos b
PO4
ATP
Syn b
Pkinase
PP1
PO4
Syn a
Phosphorylase Kinase a
H2O
ATP
ADP
cAPK
PP1
PO4
ADP
Phosphorylase Kinase b
ATP
Remember, this is the “stripper” enzyme. It takes “off”
phosphate groups on proteins
Insulin stimulates glycogen synthesis in liver and muscle
by blocking the action of GSK3 and activating PP1
Insulin
3 ATP
3 ATP
X
GSK3
CKII
P P
P
ATP
Glycogen
Synthase b
Glycogen
Synthase a
Inactive
3Pi
PP1
Insulin
Glucagon
epinephrine
X
Active
Glucose
Text p586
Glucose6-phosphate
Insulin
Insulin-stimulated
protein kinase
Phosphoprotein Phosphatase-1
(Muscle)
(site 1 phosphorylation)
Glycogen
ADP
more active
P
Glycogen synthesis
stimulated,
breakdown blocked
G subunit PP1
OH
K1
ATP
OH
Glycogen
Epinephrine,
glucagon
K2
G subunit PP1
OH
Less active
ATP (site 2 phosphorylation)
ADP
Glycogen
Breakdown
stimulated,
synthesis blocked
cAPK
K2
P
2ATP
2ADP
Glycogen
G subunit
(site 1 and 2 phosphorylation)
Text p588
P
+
PP1
Inhibitor
P
inactive
Insulin regulation
of GSK3
Shut down of
GSK3 by PKB
Text p 587
Summary of Catabolic Hormonal
Effects on Glycogen
• Glucagon and Epinephrine stimulate synthesis of
cAMP
• cAMP activates phosphorylase b kinase that
converts phosphorylase b to phosphorylase a
• Phosphorylase a breaks down glycogen at an
accelerated rate
• cAMP also inactivates PP1, prolonging the action
of phosphorylase a
Summary of Anabolic Effects on
Glycogen
• Insulin activates glycogen synthase by stimulating
phosphorylation of GDK3
• GDK3 inactivates itself allowing phosphoprotein
phosphatase (PP1) to remove phosphate and
activate glycogen synthase
• PPI also removes phosphate from phosphorylase a
thereby shutting off glycogen breakdown
See Tutorial on cAMP-dependent Protein
Kinases available on the web
Adenylcyclase
cAMP dependent
protein kinase
R
R
C C
Phosphorylase
kinase
P
Phosphorylase
 
 
P
Liver
Text p446-447
R State
T State
P
Glucose
P
High Glucose
Phosphoprotein
phosphatase-1
Phosphorylase a
Low Glucose
5’-AMP
+
Binds weakly
to b form
Phosphorylase b
Active
Low glucose a form (R state) P is resistant to phosphatase
High glucose a form (T state) P is vulnerable to phosphatase
Summary of Phosphoprotein Phosphatase-1
(Liver)
1. The Phosphatase binds to phosphorylase a (T or R form)
(It does not bind to glycogen or a G protein)
2. The T form has a serine exposed to allow hydrolysis
of -PO4
3. The R form of phosphorylase a -PO4 cannot be hydrolyzed
4. Glucose converts R to T form which causes hydrolysis of
-PO4 and dissociate the Phosphatase.
(glucose is an allosteric effector of phosphorylase in liver)
5. The liberated Phosphatase is free to activate glycogen
synthase, thereby stimulating glycogen synthesis