Lecture 18: Glycogen And Metabolism

Question 1

Describe the structure of glycogen

Answer 1

• Osmotically inactive, easily mobilized
• 12 branched layers of glucose
• Linked together with alpha 1-4 bonds, branched points at alpha 1-6 bonds.
• Non reducing ends have a terminal hydroxyl at C4.
• Reducing end is connected to glycogenin
• Stored in liver and muscles as granules

Question 2

Describe glycogen metabolism

Answer 2

• Regulated storage and release of glucose
• Synthesis and degradation of glycogen involve different pathways
• Both pathways regulated independently
• Regulation
• • Allosteric control
• • Covalent modification through reversible phosphorylation of key enzymes
• • Hormonal control

Question 3

Describe the products of glycogenolysis (the basic version)

Answer 3

• Glycogen broken down to release glucose-1phosphate
• Glycogen remnant remodeled to permit further degradation
• Glucose -1-phosphate converted to glucose-6 phosphate
• Glycolysis
• Free glucose for release into blood stream
• Pentose phosphate pathway – NADPH and ribose derivative

Question 4

What are the basic steps in glycogenolysis?

Answer 4

1. Glycogen + Glycogen Phosphorylase =
2. 5: Glucose 1-Phosphate
3. G-1-P + Phosphoglucomutase =
4. 5: G-6-P
5. G-6-P can go one of several ways. Glucose 6-Phosphatase for conversion to Glucose

Question 5

What are the Four Key Enzymes of Glycogenolysis?

Answer 5

• one to degrade glycogen (chain shortening) (Glycogen Phosphorylase)
• two to remodel glycogen remnants (Glycogen Transferase and alpha1-6 Glucosidase)
• one to convert glycogen breakdown product suitable for further metabolism (?)

Question 6

Describe Glycogen Chain Shortening

Answer 6

• Glycogen phosphorylase (GP) (rate limiting enzyme) catalyzes the cleavage of glycogen.
• Chain shortening occurs at the non-reducing end of the polymer
• GP adds an orthophosphate and releases a glucose residue as glucose-1-phosphate
• Uses pyridoxal phosphate (vitamin B6) as a cofactor
• Phosphorolysis of glucose residues continues till the GP gets within 4 residues of the α-1,6 linkage of a branch point.

Question 7

Describe the branch transfer and the release of glucose

Answer 7

• Transferase transfers a block of 3 of the remaining 4 glucose to the non-reducing end of the main chain forming an α-1,4 bond.
• Debranching enzyme or α-1,6 glucosidase cleaves the α1,6 bond of the single remaining glucose residue to release the free glucose. Glucose phosphorylated by hexokinase
• Transferase and α-1,6 glucosidase convert branched glycogen into a linear structure for further action by GP

Question 8

Describe the phosphoglucomutase step

Answer 8

• Converts Gluc-1-phosphate to Gluc-6-phosphate
• A phosphoryl group is transferred from the enzyme to the substrate, and a different phosphoryl group is transferred back to restore the enzyme to its initial state.

Question 9

Describe the glucose 6-phosphatase step

Answer 9

• Gluc-6-phosphate cannot get out of the cell
• Only the liver has glucose 6-phosphatase
• Converts it to glucose

Question 10

How is glycogen phosphorylase regulated?

Answer 10

GP regulated by:

• several allosteric effectors (signal energy state of the cell)
• reversible phosphorylation (responsive to hormones)

Question 11

What are the two states of Glycogen Phosphorylase?

Answer 11

Exists in 2 forms:

• Active “a” form (R relaxed state) – in liver
• Inactive “b” form (T tense state) – in muscle
• Both isozymes exist in equilibrium between R and T

Question 12

What inhibits phosphorylase a in the liver?

Answer 12

The binding of glucose to Phosphorylase a shifts it into the T state.

Question 13

What can inhibit and activate phosphorylase b in the muscle?

Answer 13

• Phosphorylase B switches to it’s R state in the presence of AMP
• It is usually held stable in it’s T state by the presence of ATP and Glucose 6-Phosphorylase

Question 14

Describe the allosteric muscle regulation of Glucose Phosphorylase in Muscle

Answer 14

• Default “b” form or inactive form
• Activated by AMP
• Binds to active site and stabilizes conformation of b in the active R state
• During muscle contraction ATP converted to AMP by myosin and adenylate kinase signaling the GP to breakdown glycogen
• ATP and Gluc-6-phosphate are negative allosteric regulators
• Under normal physiological conditions GP inactive because of inhibitory effect of ATP and Gluc-6-phosphate

Question 15

What is the general role of phosphorylation?

Answer 15

• Phosphorylation of a single serine residue converts b to a.
• Conversion initiated by hormones
• Phosphorylation carried out by phosphorylase kinase (PK)

Question 16

In rough terms, describe what must occur for Phosphorylase to be activated

Answer 16

1. Phosphorylase Kinase starts off in it’s inactive state
2. PK is partially activated by the presence of Calcium ions.
3. Partly activated PK is fully activated by the presence of PKA Hormones.
4. Fully activated phosphorylase kinase uses ATP to transfer phosphorylase from phosphorylase B to phosphorylase A

Question 17

Describe the hormonal control of GP

Answer 17

• Low blood sugar levels release glucagon (acts on liver)
• Muscle activity releases epinephrine (effects are on muscle)
• Effects of both hormones mediated via G protein coupled receptors (GPCR)
• Epinephrine and glucagon signal glycogen breakdown

Question 18

Describe the GPCR-mediated pathway

Answer 18

Glucagon and epinephrine Work through GPCR - do not cross membrane like steroid hormones

• Glucagon and epinephrine signal glycogen breakdown.

Question 19

What biochemical reaction is generally regarded as the “off switch” for glycogen breakdown.

Answer 19

• Shuts down when secretion of hormone stops
• PK and GP are dephosphorylated and inactivated
• Breakdown of glycogen stops
• Synthesis of glycogen promoted

Question 20

Compare Phosphorylase in the Muscle and Liver

Answer 20

• Liver and muscle forms of GP are products of separate genes. Called isozymes.
• Differ in their sensitivities to regulatory molecules.
• Both forms activated by phosphorylation by PK and inhibited by ATP and G6P.
• Muscle form is allosterically activated by AMP (measure of low energy status of cell)
• Liver enzyme is inactivated by free glucose (indicator of blood sugar levels). Unaffected by AMP.
• Mutation in liver GP causes Hers disease
• Mutation in muscle GP causes McArdle syndrome

Question 21

What do glucagon and epinephrine do in a fasting state, and in a period of exercise?

Answer 21

• In the liver, hormones signal glycogen to break down into glucose, and for lactose to be converted back into glucose
• In the muscle cells, Both glucose and glycogen are broken down into G-6-P.

Question 22

How is glucose built back into glycogen?

Answer 22

1. Glucose
2. 5. Add Hexokinase and (ATP -> ADP) =
3. Glucose 6-P
4. 5: Add Phosphoglucomutase =
5. Glucose 1-P
6. 5 Add UDP-Glucose pyrophosphorylase (UTP->2Pi) =
7. UDP-Glucose
8. 5: Add Glycogen Synthase (Rate limiting step) (Primer->UDP) =
9. (Doesn’t really have a name. Non-branched glycogen)
10. 5 Add Glucosyl 4:6 transferase
11. Glycogen

Question 23

What is the first of 3 key events of glycogenesis?

Answer 23

• Trapping and Activation of Glucose:
• Glucokinase/hexokinase in cytosol of hepatocytes and muscle cells catalyze phosphorylation of glucose to glucose-6-phosphate
• This traps the glucose in these cells
• Phosphoglucomutase then reversibly isomerizes glucose-6phosphate to glucose-1-phosphate
• Uridine diphosphate(UDP)-glucose pyrophosphorylase then transfers the glucose-1-phosphate to uridine triphosphate (UTP) which generates UDP-glucose (active form of glucose)
• Breakdown of pyrophosphate to Pi generates energy and drives the reaction forward

Question 24

What is the second of the three key steps involved in glycogen synthesis?

Answer 24

• Elongation of a glycogen primer:
• Preexisting short glycogen polymer serves as a primer to which glucose units are added
• Primer formation done by Glycogenin, a Mn requiring protein
• Glycogen synthase (rate limiting enzyme). Catalyzes the transfer of glucose from UDPglucose onto the non-reducing end of glycogen chain. Forms α-1,4 glycosidic bonds between glucose molecules

Question 25

What is the third of the three key steps of glycogen synthesis?

Answer 25

• Branching of glycogen chains:
• When glycogen chain reaches 11 residues, a fragment of the chain (about 7 residues long) is broken off at an α -1, 4 link and reattached elsewhere through an α -1, 6 link by glucosyl (4:6) transferase.
• The new branch point must be at least 4 residues away from a preexisting branch
• Branching increases solubility of glycogen and increases number of terminal non-reducing ends.
• Increases rate at which glycogen can be synthesized and degraded

Question 26

Describe the branching enzyme of glycogen synthesis

Answer 26

See Slide 42

• The branching enzyme removes an oligosaccharide of approximately seven residues from the nonreducing end and creates an internal alpha 1-6 linkage.
• Synthase extends non-reducing ends followed by more branching

Question 27

Describe how glycogen synthesis is regulated

Answer 27

• Glycogen synthase: Key enzyme
• Exists in 2 forms, one form present in liver and second in muscle and other tissues
• Active non-phosphorylated “a” form
• Inactive phosphorylated “b” form
• Interconversion mediated by covalent modification (finetuning role)
• Phosphorylated by several kinases most importantly glycogen synthase kinase (GSK)
• GSK under the control of insulin and PKA
• Allosteric regulation – gluc-6-phosphate powerful activator, stabilizes R state

Question 28

Provide an overview for the regulation of glycogen metabolism

Answer 28

• Regulation very important to maintain blood sugar and provide energy to muscles
• Pathways of synthesis and degradation are independent
• Allows for independent regulation
• Two key enzymes: glycogen phosphorylase and glycogen synthase, the rate limiting steps of degradation and synthesis, respectively.
• Both enzymes are regulated by reversible phosphorylation, but effects are in opposite directions
• Big one: Glucagon and Epinephrine are able to control PKA synthesis via PKA

Question 29

Name the cascades that stimulate glycogen breakdown and the cascades that prevent glycogen synthesis.

Answer 29

Stimulate Glycogen Breakdown:
Activated cAMP activates Protein Kinase A, which activates Phosphorylase Kinase which activates Phosphorylase into it’s A form.

Prevents Glycogen Synthesis:
Glycogen Synthase Kinase joins with activated Protein Kinase A, to DEactivate Glycogen Synthase into it’s B state.

Question 30

How does PPI inhibit and activate other enzymes after a full meal

Answer 30

PPI does pretty much three things.

1. Inhibits Glycogen Breakdown by inactivating Phosphorylase Kinase (directs it towards it’s B state)
2. Inhibits Phosphorylase (directs it towards it’s B state)
3. Stimulates Glycogen Synthesis by activating Glycogen Synthase

Question 31

Describe in further detail the regulation of glycogen metabolism

Answer 31

• Glycogenesis is favored in fed state (when glucose and insulin high and cellular ATP high - signal of high energy)
• When glycogen synthesis favored, the depospho form glycogen synthase (active) and glycogen phosphorylase (inactive) are predominant
• Glycogenolysis favored in fasting state (when blood glucose levels are low and glucagon levels are high and cellular calcium and AMP are elevated (in exercising muscles)
• When glycogen degradation is favored phosphorylated forms of glycogen synthase (inactive) and glycogen phosphorylase (active) are predominant

Question 32

Describe the mechanism of regulation by insulin

Answer 32

• High blood glucose
• Release of insulin by β cells of pancreas
• Binding of insulin to its receptor tyrosine kinase
• Activation of signaling cascade
• Four key proteins:
  –GLUT 4 (glucose transporter)
  –Protein kinase B (PKB)
  –Protein phosphatase 1 (PP1)
  –Glycogen synthase kinase 3 (GSK3)

Question 33

Describe the outcome of insulin regulation

Answer 33

• Formation of the insulin receptor complex
• Activation of PKB
• Translocation of GLUT to membrane
• PKB phosphorylates PP1 (activate) and GSK3 (inactivate)
• Active PP1 dephosphorylates glycogen synthase (activate) and dephosphorylates glycogen phosphorylase (inactivate)
• Net result - activation of glycogen synthase and inactivation of glycogen phosphorylase

Question 34

Describe Type 2 Diabetes

Answer 34

• Reduced sensitivity to insulin
• Called insulin resistance
• Mutations in insulin receptor and/or downstream signaling proteins
• Down-regulation in receptor levels triggered by elevated insulin (leading to endocytosis and degradation of the insulin receptor). Not replaced by translation

Question 35

What acts as a glucose sensor in liver cells?

Answer 35

Glycogen Phosphorylase

• See Slides 54 & 55

Question 36

Again, describe glycogen phosphorylase in both the liver, and in muscle

Answer 36

• Liver and muscle forms of GP are products of separate genes. Called isozymes.
• Differ in their sensitivities to regulatory molecules.
• Both forms activated by phosphorylation by PK and inhibited by ATP and G6P.
• Muscle form is allosterically activated by AMP (measure of low energy status of cell)
• Liver enzyme is inactivated by free glucose (indicator of blood sugar levels). Unaffected by AMP.
• Mutation in liver GP causes Hers disease
• Mutation in muscle GP causes McArdle syndrome
• In McArdle’s Syndrome, there is limited ability to perform strenuous exercise because of painful muscle cramps. Can live relatively normal life apart from that.
• In Her’s Syndrome, there is enlargement of the liver, moderate to severe hypoglycemia, ketosis, hyperuricemia, and hyperlipemia.