1. Storage Mechanisms and Control in Carbohydrate Metabolism Apr. 7, 2016 CHEM 281

3. How does Glycogen Breakdown take place?  Glycogen is cleaved by phosphate to give  -D-glucose- 1-phosphate  Cleavage reaction is phosphorolysis and not hydrolysis  No ATP is involved in reaction  Reaction is catalyzed by glycogen phosphorylase

4. How does Glycogen Breakdown take place? (Cont’d)  In the second reaction, glucose-1-phosphate is isomerized to glucose-6-phosphate  This reaction is catalyzed by phosphoglucomutase Complete breakdown requires debranching enzymes to degrade the  (1->6) linkages

5. Mode of Action of Debranching Enzyme in Glycogen Breakdown

6. How is Glycogen formed from Glucose?  Not exact reversal of glycogen breakdown to glucose  Glycogen synthesis requires energy  Energy supplied by hydrolysis of UTP  Glucose-1-phosphate reacts with UTP to make UDPG  Pyrophosphate is also formed  UDPG is then added to a growing chain of glycogen, catalyzed by glycogen synthase

7. How is Glycogen formed from Glucose? (Cont’d)  Coupling of UDPG formation with hydrolysis of pyrophosphate drives formation of UDPG to completion

8. Reaction Catalyzed by Glycogen Synthase

9. Control of Glycogen Metabolism  Glycogen phosphorylase is a major control point in the synthesis and breakdown of glycogen  Glycogen phosphorylase activity can be allosterically controlled, as well as, controlled through covalent modification

10. Control of Glycogen Metabolism (Cont’d) The activity of glycogen synthase is subject to the same type of covalent modification as glycogen phosphorylase, however, the response is opposite In addition: Hormonal signals (glucagon or epinephrine) stimulate its phosphorylation After phosphorylation, glycogen synthase becomes inactive at the same time the hormonal signal is activating phosphorylase Glycogen synthase can be phosphorylated by several other enzymes including phosphorylase kinase Dephosphorylation is by phosphoprotein phosphatase

11. Summary  Glycogen is the storage form of glucose in animals, including humans. Glycogen releases glucose when energy demands are high  Glucose polymerizes to form glycogen when the organism has no immediate need for the energy derived from glucose breakdown  Glycogen metabolism is subject to several different control mechanisms, including covalent modification and allosteric effects

12. Gluconeogenesis Gluconeogenesis: The conversion of glucose from pyruvate Gluconeogenesis is not the exact reversal of glycolysis; that is, pyruvate to glucose does not occur by reversing the steps of glucose to pyruvate Three irreversible steps in glycolysis - Phosphoenolpyruvate to pyruvate + ATP - Fructose-6-phosphate to fructose-1,6- bisphosphate - Glucose to glucose-6-phosphate Net result of gluconeogenesis is reversal of these three steps, but by different reactions and using different enzymes

13. Oxaloacetate is an Intermediate  In first step, Pyruvate is carboxylated to oxaloacetate  Requires biotin (CO 2 carrier)  Pyruvate carboxylase is subject to allosteric control; it is activated by acetyl-CoA

14. Gluconeogenesis (Cont’d)  Next, decarboxylation of oxaloacetate is coupled with phosphorylation by GTP to give PEP  The net reaction of carboxylation/decarboxylation is Pyruvate + ATP +GTP → Phosphenolpyruvate + ADP + GDP + P i

15. Pyruvate Carboxlyase Reaction

16. Role of Sugar Phosphates  Other different reactions in gluconeogenesis relative to glycolysis involve phosphate-ester bonds bound to sugar-hydroxyl groups being hydrolyzed   G° = -16.7kJ mol -1  Fructose-1,6-bisphosphatase is an allosteric enzyme, inhibited by AMP and activated by ATP fructose-1,6-bisphosphatase is an allosteric enzyme, inhibited by AMP and activated by ATP

17. Role of Sugar Phosphates (Cont’d)  Another reaction is the hydrolysis of glucose-6-phosphate to glucose and P i  Reaction also spontaneous (  G°’ = -13.8 kJ mol -1 )  Reaction catalyzed by glucose-6-phosphatase

18. Control of Carbohydrate Metabolism  Allosteric control: fructose-2,6-bisphosphate (F2,6P)  An allosteric activator of phosphofructokinase (PFK)  An allosteric inhibitor of fructose bisphosphate phosphatase (FBPase)  High concentration of F2,6P stimulates glycolysis; a low concentration stimulates gluconeogenesis  Concentration of F2,6P in a cell depends on the balance between its synthesis (catalyzed by phosphofructokinase-2) and its breakdown (catalyzed by fructose bisphosphatase-2)  Each enzyme is controlled by phosphorylation/dephosphorylation

19. Synthesis and Breakdown of F2,6P

20. Mechanisms of Metabolic Control

21. Substrate Cycling  Substrate cycling  opposing reactions can be catalyzed by different enzymes and each opposing enzyme or set of enzymes can be regulated independently Fructose-6-Phosphate + ATP → Fructose-1,6,- bisphosphate + ADP Fructose-1,6,-bisphosphate + ADP → Fructose-6- Phosphate + P i Both Reactions are exergonic, and the net reaction is ATP +H 2 O ↔ ADP + P i

22. The Cori Cycle: How Different Organs Share Carbohydrate Metabolism  The Cori cycle  Under vigorous anaerobic exercise, glycolysis in muscle tissue converts glucose to pyruvate; NAD + is regenerated by reduction of pyruvate to lactate  Lactate from muscle is transported to the liver where it is reoxidized to pyruvate and converted to glucose  The liver shares the stress of vigorous exercise

23. The Cori Cycle

24. Control of Liver Pyruvate Kinase by Phosphorylation

25. Major Control Points in Carbohydrate Metabolism  First and last steps in glycolysis are major control points in glucose metabolism  Hexokinase  Inhibited by high levels of glucose 6-phosphate  When glycolysis is inhibited through phosphofructokinase, glucose 6- phosphate builds up, shutting down hexokinase  Pyruvate kinase (PK) is an allosteric enzyme  Inhibited by ATP and alanine  Activated by fructose-1,6-bisphosphate  PK isoenzymes have 3 different subunits  M predominates in muscle, L in liver, and A in other tissues  Native PK is a tetramer  Liver isoenzymes are subject to covalent modification

26. Summary  A number of control mechanisms operate in carbohydrate metabolism. These include allosteric effects, covalent modification, substrate cycles, and genetic control  In the mechanism of substrate cycling, the synthesis and the breakdown of a given compound are catalyzed by two different enzymes

27. Glucose is Sometimes Diverted through the Pentose Phosphate Pathway  The Pentose Phosphate Pathway (PPP) is an alternative to glycolysis, and differs in several ways  In glycolysis, ATP production is important, in PPP, this is not the case  As the name implies, five-carbon sugars, including ribose, are produced from glucose  Oxidizing agent is NADP + ; it is reduced to NADPH, which is a reducing agent in biosyntheses  Begins with two oxidation steps (NADP + ) to give ribulose-5- phosphate  Following this, a series of carbon-shuffling steps occur during which three-, four-, five-, six-, and seven-carbon monosaccharide phosphates are produced

28. The Pentose Phosphate Pathway

30. Control of the PPP  The carbon-shuffling reaction are catalyzed by:  Transketolase for the transfer of two-carbon units and  Transaldolase for the transfer of three-carbon units  Control of the pentose phosphate pathway is maintained by:  Glucose-6-phosphate (G6P) can be channeled into either glycolysis or the pentose phosphate pathway  G6P channeling into glycolysis, if ATP needed  G6P channeling into the pentose phosphate pathway, if NADPH or ribose-5-phosphate are needed

31. Group Transfer Reactions

32. Relationship between PPP and Glycolysis