Glycogen metabolism

GLYCOGENESIS,GLYCOGENOLYSIS & GSD

1. GLYCOGEN METABOLISM

2. GLYCOGEN METABOLISM Carl Cori and Gerty -glycogen degradation. Glycogen synthesis, Luis Leloir (Argentina) was awarded Nobel prize in 1970. Earl Sutherland studied the role of cyclic AMP as the second messenger in glycogenolysis . Glycogen is a homopolysaccharide with glucose units linked in alpha-1, 4 linkages (straight line) and alpha-1, 6 linkages (branching point). Branching makes the molecule more globular and less space-consuming.

3. FIG. Pathways of glycogenesis and glycogenolysis in the liver.

4. FIG. Control of glycogen phosphorylase in muscle. The sequence of reactions arranged as a cascade allows amplification of the hormonal signal at each step. n, number of glucose residue

5. Fig . Coordinated control of glycogenolysis and glycogenesis by cAMP dependent protein kinase . Coordinated control of glycogenolysis and glycogenesis by cAMP -dependent protein kinase . The reactions that lead to glycogenolysis as a result of an increase in cAMP concentrations are shown with bold arrows. Inhibited by activation of protein phosphatase-1 are shown with dashed arrows. The reverse occurs when cAMP concentrations decrease as a result of phosphodiesterase activity, leading to glycogenesis .

6. Functions of Glycogen 1. storage form , liver and muscle. liver glycogen is to provide glucose during fasting. The glycogen content of liver (10 g/100 g tissue) skeletal muscle (1–2 g/100 g) 2. liver glycogen is broken down and helps to maintain blood glucose level. After taking food, blood sugar tends to rise,

7. Functions of Glycogen continue… After taking food, blood sugar tends to rise, which causes glycogen deposition in liver. About 5 hours after taking food, the blood sugar tends to fall. But, glycogen is lyzed to glucose so that the energy needs are met. After about 18 hours fasting, most of the liver glycogen is depleted, when depot fats are hydrolyzed and energy requirement is met by fatty acid oxidation . 3. The function of muscle glycogen is to act as reserve fuel for muscle contraction. 4. All the enzymes related to glycogen metabolism are cytoplasmic .

8. DEGRADATION OF GLYCOGEN (GLYCOGENOLYSIS) -Steps of Glycogenolysis

9. Step 1 -Glycogen Phosphorylase i . Glycogen phosphorylase removes glucose as glucose- 1-phosphate from glycogen ( phosphorolysis ) . It contains pyridoxal phosphate (PLP) as a prosthetic group. The alpha-1, 4 linkages in the glycogen are cleaved. ii. It removes glucose units one at a time. Enzyme sequentially hydrolyzes alpha-1, 4 glycosidic linkages,

10. Step 2.Debranching by Bifunctional (Two) Enzymes A block of 3 glucose residues ( trisaccharide unit) are transferred from the branching point to another branch. Enzyme is alpha-1, 4 → alpha-1, 4 glucan transferase . ii. Now the branch point is free. Then alpha-1, 6- glucosidase ( debranching enzyme) can hydrolyze the remaining glucose unit held in alpha-1, 6 linkage at the branch point . iii. This glucose residue is released as free glucose , the ratio of glucose-1-phosphate to free glucose is about 8:1. iv. The transferase and alpha-1, 6-glucosidase will together convert the branch point to a linear one. With the removal of the branch point, phosphorylase nzyme can proceed with its action.

11. Sep 3. Phosphoglucomutase Phosphorylase reaction produces glucose-1-phosphatem while debranching enzyme releases glucose. The glucose- 1-phosphate is converted to glucose-6-phosphate by phosphoglucomutase .

12. Step 4.Glucose-6-phosphatase in Liver Next, hepatic glucose-6-phosphatase hydrolyzes glucose- 6-phosphate to glucose. The free glucose is released to the bloodstream.

13. Muscle lacks Glucose -6-phosphatase Muscle will not release glucose to the bloodstream, because muscle tissue does not contain the glucose-6-phosphatase. Instead, in muscle, glucose-6-phosphate undergoes glycolysis to produce ATP for muscle contraction.

14. Energetics Glycogenolysis i . The energy yield from one glucose residue derived from glycogen is 3 ATP molecules, because no ATP is required for initial phosphorylation of glucose (step 1 of glycolysis ). ii. If glycolysis starts from free glucose only 2 ATPs are produced.

15. GLYCOGEN SYNTHESIS (GLYCOGENESIS)

16. GLYCOGEN SYNTHESIS (GLYCOGENESIS) The glycogen synthesis occurs by a pathway distinctly different from the reversal of glycogen breakdown, which would prevent the operation of futile cycles. The steps are:

17. Step-1. Activation of Glucose UDP glucose is formed from glucose-1-phosphate and UTP ( uridine triphosphate ) by the enzyme UDP-glucose pyrophosphorylase .

18. Step- 2.Glycogen Synthase The glucose moiety from UDP-glucose is transferred to a glycogen primer ( glycogenin ) molecule. The primer is essential to accept the glycosyl unit. The primer is made up of a protein-carbohydrate complex. It is a dimeric protein, having two identical monomers. An oligosaccharide chain of 7 glucose units is added to each monomer.

19. Step- 2.Glycogen Synthase continue…. In the next step, activated glucose units are sequentially added by the enzyme glycogen synthase . The glucose unit is added to the nonreducing (outer) end of the glycogen primer to form an alpha-1, 4 glycosidic linkage and UDP is liberated.

20. Glycogen Synthase and Primer The glycogen primer is formed by autoglycosylation of glycogenin . Glycogenin is a dimeric protein, the monomers glycosylating each other using UDP glucose till seven glucose units are added. This molecule acts as the glycogen primer to which glucose units are added by glycogen synthase .

21. Step-3.Branching Enzyme The glycogen synthase can add glucose units only in alpha-1, 4 linkage. A branching enzyme is needed to create the alpha-1, 6 linkages. ii. When the chain is lengthened to 11–12 glucose residues, the branching enzyme will transfer a block of 6 to 8 glucose residues from this chain to another site on the growing molecule. The enzyme amylo - [1, 4]→[1, 6]- transglucosidase ( branching enzyme) forms this alpha-1, 6 linkage. iii. To this newly created branch, further glucose units can be added in alpha-1, 4 linkage by glycogen synthase .

22. Regulation of Glycogen Metabolism i . The synthesis and degradation pathways are reciprocally regulated to prevent futile cycles. ii. The phosphorylated form of glycogen phosphorylase is active; but glycogen synthase becomes inactive on phosphorylation . The covalently modified phosphorylase is active even without AMP . Active ( dephosphorylated ) glycogen synthase is responsive to the action of glucose-6- phosphate . Covalent modification modulates the effect of allosteric regulators. The hormonal control by covalent modification and allosteric regulation are interrelated . iii. These hormones act through a second messenger, cyclic AMP ( cAMP ) iv . The covalent modification of glycogen phosphorylase and synthase is by a cyclic AMP mediated

23. Regulation of Glycogen Metabolism iv. The covalent modification of glycogen phosphorylase and synthase is by a cyclic AMP mediated cascade. Specific protein kinases bring about phosphorylation and protein phosphatases cause dephosphorylation .

24. Regulation of Glycogen Metabolism Generation of Cyclic AMP ( cAMP ) Protein Kinase Activation Phosphorylase Kinase Activation Glycogen Phosphorylase in Liver and Muscle Glycogen Synthase

25. Generation of Cyclic AMP ( cAMP ) i . Both liver and muscle phosphorylases are activated by a cyclic AMP mediated activation cascade triggered by the hormonal signal. ii. The hormones epinephrine and glucagon can activate liver glycogen phosphorylase but glucagon has no effect on the muscle. iii. When the hormone binds to a specific receptor on the plasma membrane, the enzyme adenyl cyclase is activated which converts ATP to cyclic AMP ( cAMP ). iv. When level of cyclic AMP rises, it will activate a protein kinase .

26. Protein Kinase Activation The protein kinase is inactive when the catalytic (C) and regulatory (R) subunits are associated with each other. Earl Sutherland studied the role of cyclic AMP as the second messenger in glycogenolysis . The cAMP combines with the R subunit so that the C subunit is free to have its catalytic activity . PKA is an enzyme that can phosphorylate serine and threonine residues of several enzyme proteins and is activated by cAMP which combines with the regulatory subunit of PKA. In the absence of cAMP , PKA is an inactive tetramer.

27. Protein Kinase Activation

28. Protein Kinase Activation The intracellular concentration of cAMP therefore decides the level of active PKA. cAMP level depends on the activity of adenylate cyclase and phosphodiesterase . Cyclic AMP level is increased by glucagon and decreased by insulin.

29. Phosphorylase Kinase Activation The active protein kinase can now convert the phosphorylase kinase to an active phosphorylated form, which converts phosphorylase -b to phosphorylase -a . Phosphorylase kinase itself is a tetrameric enzyme (alpha, beta, gamma , delta). The gamma subunit has the catalytic site and the other 3 subunits have regulatory effects . Phosphorylase kinase is activated by Ca++ and phosphorylation of alpha and beta subunits by PKA . Phosphorylation of alpha and beta subunits relieves autoinhibition of catalytic activity of gamma subunit . Binding of Ca++ to the delta subunit which is identical to calmodulin ( CaM ) is also necessary for full activity of delta subunit. since it also has a role in dysregulating the

30. Phosphorylase Kinase Activation continue.. I t also has a role in dysregulating the gamma subunit . Calcium triggers muscle contraction as well as glycogen breakdown through the action of phosphorylase kinase . The rate of glycogenolysis is linked to rate of muscle contraction.

31. Phosphorylase Kinase Activation continue.. The dephosphorylation of the active form by protein phosphatase 1 (PP1) involves removal of phosphate group from phosphorylase a and alpha and beta subunits of phosphorylase kinase . The activity of PP1 is controlled differently in liver and muscle. The catalytic subunit of PP1 in muscle is active only when it is bound to glycogen through the glycogen binding GM subunit . The phosphorylation of PP1 by an insulin stimulated protein kinase (site1) activates the enzyme where as phosphorylation at site 2 by PKA makes its action ineffective. When cAMP level is high, PP1 is inhibited by inhibitor1 which is activated by phosphorylation by PKA. The effect of cyclic AMP is not only by increasing the phosphorylation of enzymes, but also by decreasing dephosphorylation .

32. Glycogen Phosphorylase in Liver Liver: The liver phosphorylase -b is the inactive form. It becomes active on phosphorylation . The active enzyme is denoted as phosphorylase -a. The enzyme is inhibited by ATP and glucose-6-phosphate. In the liver the PP1 is regulated differently through the intermediary of glycogen binding subunit (GL). GL complex can bind to R and T forms of phosphorylase a. The phosphate group attached to ser14 is exposed only in the T state, so that PP1 can convert phosphorylase a to phosphorylase b. Glucose is an allosteric inhibitor of phosphorylase a. Insulin favors this effect by promoting the uptake and phosphorylation of glucose.

33. Glycogen Phosphorylase in Muscle b. Muscle: Skeletal muscle glycogen is degraded only when the demand for ATP is high. The regulation of glycogenolysis in skeletal muscle is by epinephrine. Glucagon has no effect on muscle glycogenolysis . AMP formed by degradation of ATP during muscle contraction is an allosteric activator of phosphorylase b. The active form of phosphorylase is referred to as ‘a’ (active, phosphorylated ) and the relatively inactive dephosphorylated form as ‘b’. The active glycogen synthase (a) is dephosphorylated and phosphorylated (b) is relatively inactive. Glycogen phosphorylase is activated by phosphorylation by phosphorylase kinase that adds phosphate group to a specific serine residue of phosphorylase b (ser14). This phosphorylase kinase , in

34. FIG. Control of glycogen phosphorylase in muscle. The sequence of reactions arranged as a cascade allows amplification of the hormonal signal at each step. n, number of glucose residue

35. Glycogen Phosphorylase in Muscle This phosphorylase kinase , in turn, is activated by protein kinase A or cyclic AMP dependent protein kinase . Phosphoprotein phosphatase I dephosphorylates both phosphorylase kinase and phosphorylase b. Phosphorylase b is sensitive to allosteric effectors like AMP but phosphorylase a is not sensitive. High concentration of ATP and glucose-6-phosphate in the cell will inhibit phosphorylase b.

36. Glycogen Synthase i . Glycogen synthase and phosphorylase activities are reciprocally regulated . ii. The same protein kinase , which phosphorylates the phosphorylase kinase would also phosphorylate glycogen synthase . iii. The activity of the glycogen synthase is markedly decreased on phosphorylation . Insulin promotes glycogen synthesis by favouring dephosphorylation . Glycogen synthase is active in the dephosphorylated state. Phosphorylase kinase can phosphorylate glycogen synthase and inactivate the enzyme. PKA can also inactivate the enzyme by phosphorylation . Ca++ and calcium dependent cam kinase also phosphorylate the enzyme.

37. Glycogen Synthase continue…. Relative rates of glycogen synthesis and breakdown are controlled by the action of PKA, phosphorylase kinase and PP1. PP1 can activate glycogen synthase only after dephosphorylating and inactivating phosphorylase a. The regulation of glycogen phosphorylase and synthase is a typical example of multisite phosphorylation (primary and secondary sites) for metabolic regulation. Control by allosteric effectors is superimposed on covalent modification. Glucose-6-phosphate can activate glycogen synthesis. Insulin activates PP1 and PDE which decreases cAMP level and increases G6P level to promote glycogen synthesis.

38. Glycogen Synthase continue…. The reciprocal regulation of glycogenolysis and glycogenesis is by covalent modification ( phosphorylation and dephosphorylation ). Insulin and glucagon are the major regulatory hormones, although epinephrine has stimulatory effect on glycogenolysis in both liver and muscle. They bring about alterations in the activity of protein kinases and phosphatases by varying the level of cAMP .

39. Summary of Glycogen Regulation i . The key enzyme for glycogenolysis is phosphorylase , which is activated by glucagon and epinephrine, under the stimulus of hypoglycemia. ii. The key enzyme for glycogen synthesis is glycogen synthase , the activity of which is decreased by glucagon and epinephrine but is enhanced by insulin, under the stimulus of hyperglycemia . Glycogen metabolism is regulated by coordinated regulation of glycogen synthase and glycogenphosphorylase . The regulatory mechanisms include: Allosteric control as well as hormonal control by covalent modification of enzymes. The allosteric effectors are ATP, glucose-6-phosphate and AMP.

40. GLYCOGEN STORAGE DISEASES These are inborn-errors of metabolism.

41. Glycogen Storage Disease Type-I It is also called Von Gierke’s Disease. Most common type of glycogen storage disease is type I. 2. Incidence is 1 in 100,000 live births. 3. Glucose-6-phosphatase is deficient. 4. Fasting hypoglycemia that does not respond to stimulation by adrenaline. The glucose cannot be released from liver during over night fasting 5. Hyperlipidemia , lactic acidosis and ketosis. 6. Glucose-6-phosphate is accumulated, so it is channeled to HMP shunt pathway producing more ribose and more purine nucleotides. 7. Purines are then catabolized to uric acid, leading to hyperuricemia . 8. Glycogen gets deposited in liver. Massive liver enlargement may lead to cirrhosis. 9. Children usually die in early childhood. 10. Treatment is to give small quantity of food at frequent intervals. Other types : 1 in 1 million births

42. Pompe’s disease infantile-onset Pompe’s disease Myozyme (recombinant human acid alpha glucosidase , rhGAA ) is administered intravenously.