Glycogen Metabolism PDF

Glycogen Metabolism  Glycogen Metabolism Glycogen is the major storage carbohydrate in animals; it’s a branched polymer of α-D-glucose It occurs mainly in liver and muscle Because of its greater mass muscle contains about three to four times as much glycogen as the liver...

Glycogen Metabolism  Glycogen Metabolism Glycogen is the major storage carbohydrate in animals; it’s a branched polymer of α-D-glucose It occurs mainly in liver and muscle Because of its greater mass muscle contains about three to four times as much glycogen as the liver Muscle glycogen is a readily available source of glucose for glycolysis within the muscle itself Liver glycogen functions to store and export glucose to maintain blood between meals Glycogen is present in the cytosol in the form of granules which contain regulatory proteins as well as enzymes that catalyze the synthesis and degradation of glycogen -After 12-18 hours of fasting the liver glycogen is almost totally depleted Glycogen storage diseases are inherited disorders characterized by deficient mobilization of glycogen or deposition of abnormal forms leading to muscular weakness or death 2 2  Glycogenesis occurs mainly in muscle and liver As in glycolysis glucose is phosphorylated to glucose 6-P by hexokinase in muscle and glucokinase in liver Glucose 6-phosphate is isomerized to glucose 1-phosphate by phosphoglucomutase (enzyme itself is phosphorylated and the phosphoryl group takes part in a reversible reaction in which glucose 1,6- bisphosphate is an intermediate) Next glucose 1-phosphate reacts with uridine triphosphate (UTP) to form an active nucleotide uridine diphosphate glucose (UDPGlc) and pyrophosphate, catalyzed by UDPGlc pyrophosphorylase. -The reaction is reversible Loading... -but the essentially irreversible hydrolysis of pyrophosphate to orthophosphate, catalysed by inorganic pyrophosphatase drives the synthesis of UDP–glucose -UDP–glucose, the glucose donor in biosynthesis of glycogen is an activated form of glucose, just as ATP and acetyl-CoA are activated forms of orthophosphate and acetate respectively. -C1 carbon of the glucosyl unit of UDP–glucose is activated cos its hydroxyl group is esterified to the diphosphate moiety of UDP. NB * Many biosynthetic reactions are driven by hydrolysis of pyrophosphate 3 3 * Nucleoside diphosphate sugars serve as glycosyl donors in the  Glucose Hexokinase or glucokinase (liver) O –O P O C C 2 OH 2 H H H O H –O H H H Phosphoglucomutase H O H H O OH OH H HO OH O P O– HO H OH H OH O– Glucose 6-phosphate Glucose 1-phosphate + O O UDP-glucose O O O Glu P P pyrophosphorylase O O O Rib Uracil P P Uracil –O P O O O Rib O– O– O– O– O – UDP-Glucose UTP Inorganic O O O pyrophosphatase 2 –O P OH –O P O P O– O – O– O– 4 Pyrophosphatase (PPi) Phosphate (Pi) 4  Glycogen synthase catalyzes the formation of a glycosidic bond between C1 of the activated glucose of UDPGlc and C4 of a terminal glucose residue of glycogen, liberating UDP. A preexisting glycogen molecule (glycogen primer) must be present to initiate this reaction The first step in glycogen synthesis is the attachment of a glucose residue on Tyr 194 of a protein glycogenin, catalyzed by the protein’s glucosyltransferase activity Glycogenin then forms a tight complex with glycogen synthase Glycogenin then autocatalyzes the addition of seven more glucose residues in a 1→4 linkage to form a primer; UDPGlc is the glucose donor in this autoglycosylation. Loading... At this point glycogen synthase which is tightly bound to glycogenin takes over addition of further glucose residues on the substrate in the 1→4 position UDP-glucose is the intermediate donor of glucose residues in a reaction catalyzed by glycogen synthase, which transfers a a glucose residue from UDP-glucose to the nonreducing end of a branched glycogen molecule Glycogen synthase is catalytically efficient only when its bound to glycogenin Consequences The number of glycogen molecules is determined by the.number of glycogenin molecules Elongation stops when glycogen synthase is no longer in contact with glycogenin which forms the core of the particle 5 5  6 C 2OH H H H O 5 H 1 4 OH H None reducing end of HO 3 2 O glycogen with n residues H OH (n>4) O P P O– UDP-glucose –O O O O Rib. Uracil C C 2OH 2OH H H H O H H O H H H 4 OH H 1 4 OH H 1 HO O O Glycogen synthase OH H OH H UDP New none reducing end C 2OH C 2OH C 2OH H H H H O H H O O H H H H H 1 H 4 1 4 OH H 4 OH H 1 OH H O HO O O H OH OH H OH H Elongated glycogen with n+1 residues 6 6  -Glycogen synthase exists in either a phosphorylated or nonphosphorylated state. -The active form is dephosphorylated (glycogen synthase a) and may be inactivated to glycogen synthase b by no fewer than six protein kinases on serine residues (multiple sites) -Two protein kinases are Ca 2+/Calmodulin dependent - cAMP-dependent protein kinase which inhibits glycogenesis -remainder are poorly characterized - Insulin also promotes glycogenesis in muscle by raising glucose 6- phosphate concentration which stimulates dephosphorylation and activation of glycogen synthase -dephosphorylation of glycogen synthase b is carried out by protein phosphatase-1 which is under control of cAMP-dependent protein 7 kinase 7  Epinephrine Adenylyl cyclase Adenylyl (Active) cyclase ATP cAMP cAMP-dep. protein cAMP-dep. protein kinase (inactive) kinase (active) Glycogen synthase a (active) Glycogen synthase b (inactive) Glycogenesis 8 8  Branching involves attachment of part of an existing glycogen chain to another. - When the chain has lengthened by at least 11 glucose residues, the branching enzyme (amylo-1,4→1,6-transglucosidase) transfers, to a more interior side, a part of 1→4 chain (at least 6 glucose residues) to a neighbouring chain to form a 1→6 linkage. -The branches grow by further additions of 1→4 –glucosyl units and further branching. -New branch must be at least four residues away from the pre-existing one - Branching is important because: -It increases solubility of glycogen -Creates a number of terminal residues, the sites of action of glycogen phosphorylase and synthase. Thus branching increases the rate of glycogen synthesis and degradation. 9 9 Glycogenolysis is a separate pathway (not reversal of glycogenesis) -Glycogen phosphorylase catalyzes the rate-limiting step in glycogenolysis by promoting phosphorylytic cleavage by inorganic phosphate (phosphorolysis) of the 1→4 linkages of glycogen to yield glucose 1-phosphate. -The terminal glucosyl residues from the outer most chains of glycogen are removed sequentially until approximately four glucose residues remain on either side of a 1→6 branch α-1,4→1,4 glucan transferase then transfers a trisaccharide unit from one branch to another, exposing the 1→6 branch point - Hydrolysis of the 1→6 linkages is effected by the debranching enzyme (amylo-1,6-glucosidase) releasing one glucose and further phosphorylase action can proceed -The action of phosphorylase, glucan transferase and debranching enzyme leads to complete breakdown of glycogen. -Glucose 6- phosphate can be formed from glucose 1-phosphate by the action of phosphoglucomutase. - In the liver and kidney (but not in muscle or brain) , glucose 6-phosphatase hydrolyzes glucose 6- phosphate to glucose that is exported leading to increased blood glucose concentration. 10 10  -Phosphorylated glucose in contrast with glucose cannot readily diffuse out of cells. The hydrolytic enzyme glucose 6- phosphatase enables glucose to leave the cells -Consequently glucose 6-phosphate is retained by muscle or brain , which need a large amount of fuel for the generation of ATP. Loading... -In contrast glucose is not the major fuel for the liver – the liver stores and releases glucose primarily for benefit of other tissues. 11 11 Cyclic AMP (cAMP) integrates the regulation of glycogenolysis and glycogenesis The principal enzymes controlling glycogen metabolism- glycogen phosphorylase and glycogen synthase are regulated by reversible phosphorylation and dephosphorylation of enzyme protein in response to hormone action cAMP- formed from ATP by the action of adenylyl cyclase in the inner surface of cell membrane - acts as an intracellular second messenger in response to epinephrine, norepinephrine, and glucagon -hydrolyzed by phosphodiesterase, terminating hormone action - in liver insulin increases the activity of phosphodiesterase Phosphorylase Differs between liver and muscle Regulated by factors that signal the energy status of the cell and reversible phosphorylation which is responsive to hormones such as epinephrine, insulin,12 and glucagon. 12 Unlike glycogen synthase the active form is phosphorylated and the inactive is In liver Phosphorylase a (active form) is phosphorylated It is inactivated by the removal of a phosphate by protein phosphatase-1 to form phosphorylase b. Reactivation requires rephosphorylation catalyzed by phosphorylase kinase Glycogenolysis in liver can be cAMP independent In addition to the action of glucagon in causing formation of cAMP and activation of phosphorylase in the liver; epinephrine and norepinephrine can mediate stimulation of glycogenolysis. This involves cAMP-independent mobilization of Ca 2+ from mitochondria into the cytosol followed by the stimulation of a Ca 2+ / calmodulin –sensitive phosphorylase kinase cAMP-independent glycogenolysis is also caused by vasopressin, oxytocin, angiotensin II acting through Ca 2+ or the phosphatidylinositol bisphosphate pathway 13 13  In the muscle Muscle lacks both glucagon receptors and glucose 6-phosphatase Therefore muscle glycogen can not be mobilized to replenish blood glucose Muscle Glycogenolysis is activated 1. In response to epinephrine through β-receptors (cAMP-dependent) This occurs during Flight and fight situation Prolonged exercise 2. Hormone independent mechanisms – The influx of Ca 2+ into the muscle cytoplasm in response to nerve stimulation activates the basal, unphosphorylated form of phosphorylase kinase by the action of Ca 2+-calmodulin complex. This is required during short bursts of exercise even in the absence of epinephrine – Direct allosteric activation of phosphorylase by AMP Increased usage of ATP during rapid burst of muscle activity leads to rapid accumulation of ADP which is converted to AMP by adenylate kinase 2 ADP → ATP + AMP AMP activates both the basal and phosphorylated forms of phosphorylase, enhancing Glycogenolysis either in the absence or presence of hormonal stimulation – The stimulatory effects of Ca 2+ and AMP assure that the muscle can respond to its energy 14 needs, even in the absence of hormonal input 14  Phosphorylase is present in two forms Phosphorylase a, which is phosphorylated Is active in either the presence or absence of AMP Is the normal physiologically active form of the enzyme Phosphorylase b, which is dephosphorylated Under most physiological conditions phosphorylase b is inactive because of the inhibitory effect of ATP and glucose 6-phosphate Active only in the presence of AMP, when the energy status is low as is the case during exercise, resulting in generation of fuel for the muscle In resting muscle nearly all the enzyme is in the inactive b form. During exercise elevated levels of AMP leads to activation of phosphorylase b cAMP activates muscle phosphorylase In response to epinephrine acting via cAMP Increased cAMP activates cAMP-dependent protein kinase ( Protein Kinase A), which catalyzes the phosphorylation by ATP of inactive phosphorylase kinase b to active phosphorylase kinase a, which by further phosphorylation activates phosphorylase b to phosphorylase a 15  Epinephrine Adenylyl cyclase Adenylyl (Active) cyclase ATP cAMP cAMP-dep. protein cAMP-dep. protein kinase (inactive) kinase (iactive) Phosphorylase kinase a Phosphorylase kinase b (active) (inactive) Phosphorylase b Phosphorylase a ( inactive) (active) Glycogenolysis 16 16  Ca 2+ synchronizes the activation of phosphorylase with muscle contraction Glycogenolysis increases in muscle several hundred- fold immediately after the onset of contraction This involves the rapid activation of phosphorylase by activation of phosphorylase kinase by Ca 2+, the same signal which initiates muscle contraction in response to nerve stimulation. Muscle phosphorylase kinase has four subunits–α β γ δ α and β units contain serine residues that are phosphorylated by cAMP-dependent protein kinase The δ subunit binds calcium and is identical to calcium binding protein calmodulin the binding of Ca 2+ activates the catalytic subunit of the γ subunit while the molecule remains in a dephosphorylated b configuration The phosphorylated form is only fully active in the presence of Ca 2+17 17  Muscle contraction Hormone-triggered Nerve cAMP cascade impulse Ca 2+ release Ca 2+ - free Phosphorylated kinase Ca 2+ - kinase kinase Inactive Partly active Fully active 18 18 Protein phosphatase-1 inactivates phosphorylase and activates glycogen synthase Both phosphorylase a and phosphorylase kinase are dephosphorylated and inactivated by protein phosphatase-1 protein phosphatase-1 is inhibited by a protein, inhibitor-1 which is active only after it has been phosphorylated by cAMP-dependent protein kinase Insulin reinforces the effect of the phosphatase by inhibiting the activation of phosphorylase b (increased glucose uptake increases formation of glucose 6-phosphate, which is an inhibitor of phosphorylase kinase). Also triggers a cascade that leads to activation of protein phosphatase-1 Protein phosphatase-1 also removes the phosphoryl group from glycogen synthase b to convert it into a much more active form. Dephosphorylation of glycogen synthase, phosphorylase kinase and phosphorylase promotes glycogen synthesis and prevents its degradation. 19 19 Regulation of glycogen metabolism (Summary) Under the influence of cAMP –dependent protein kinase, phosphorylase is activated but glycogen synthase is at the same time converted to the inactive form. Inhibition of glycogenesis enhances net glycogenolysis. Inhibition of glycogenolysis enhances net glycogenesis Dephosphorylation of phosphorylase, phosphorylase kinase a, and glycogen synthase b is catalyzed by a single enzyme of wide specificity- protein phosphatase-1. In turn protein phosphatase-1 is inhibited by cAMP –dependent protein kinase via inhibitor-1. The control of the synthesis and degradation of glucose in the liver is central to the regulation of blood glucose. The liver senses the concentration of glucose in the blood and takes up or releases glucose accordingly After glucose infusion the amount of liver phosphorylase a decreases rapidly and after a lag period the amount of glycogen synthase a increases resulting in synthesis of glycogen Phosphorylase a is the glucose sensor in the cells The binding of glucose to phosphorylase a exposes the phosphoryl group to hydrolysis by protein phosphatase -1 Conversion of phosphorylase a to phosphorylase b releases the protein phosphatase -1 which is then free to dephosphorylate and activate glycogen synthase. Removal of the phosphoryl group from the inactive synthase b converts it into the active synthase a form. Hence the activity of the synthase begins to increase only after most of the phosphorylase a is converted to phosphorylase b 20 20  Pathways of glycogenesis and of glycogenolysis in the liver Glycogen UDP (1 4 and 1 6 glycosyl units) Pi Branching enzyme (1 4 Glucosyl units)x Glyco Insulin gen primer – Glycogen Glycogen cAMP Glyco synthase Phosphorylase genin – + + Glucagon Glucan transferase Epinephrine Debranching Uridine diphosphate enzyme Inorganic glucose (UDPGlc) pyrophosphatase UDPglc pyrophosphorylase Free glucose PPi from 2Pi debranching enzyme UDP Uridine triphosphate Glucose 1-P (UTP) Phosphoglucomutase To glycolysis and pentose Glucose 6-P phosphate pathway Glucose 6 phosphatase Glucose Glucokinase Nucleoside ATP diphospho ADP 21 Kinase 21  Glycogen Storage Diseases Type Defective Glycogen Clinical Organ features Enzyme 1 von G6- Liver and Massive enlargement of liver, Normal hypoglycaemia, ketosis, Gierke’s phosphatase kidney disease hyperuracaemia, hyperlipemia II Pompe’s α-1,4- All organs Normal Cardiorespiratory disease Glucosidase Massive failure. Death usually (lysosomal) before 2 years III Cori’s Debranching Muscle & liver Short outer Like I but disease enzyme branches milder IV Liver cirrhosis. Andersen’s Branching Liver and Very long outer branches. Liver failure. disease enzyme spleen Death usually Normal amount before 2 years 22 22 V McArdle’s Phosphorylae Muscle Limited ability for strenuous Moderately exercise otherwise patient disease increased. normal Normal VI Her’s Phosphorylase Liver disease Increased Like type I but milder amount VII Tarui’s Phosphofructo Muscle and Like type V disease Increased kinase erythrocytes amount. Normal structure VIII Phosphofructo Liver Increased Mild liver kinase amount. enlargement. Normal Mild structure hypoglycaemia 23 23 En d