Human diseases of carbohydrate metabolism

1. Human diseases of carbohydrate metabolism Inherited enzyme deficiencies Mutations that change enzyme function or abolish enzyme activity Most are recessive since only one functional copy of gene is sufficient for needed activity Diabetes Lactose intolerance Galactosemia Glycogen storage disease

2. Monosaccharides - Aldoses # Isomers = 2n where n = # of chiral carbons Enantiomer Distant chiral C From most oxidized Epimers – differ in configuration at only one chiral carbon Not all made in nature

3. Monosaccharides - Ketoses # Isomers = 2n where n = # of chiral carbons

4. Cyclization - aldohexose • Draw most oxidized carbon (C1 aldose and C2 ketose) on right and number C clockwise • In ring most oxidizes carbon new chiral center (anomeric C) • Transfer information from Fisher projections • -OH on right then down in Haworth • -OH on left then up in Haworth • Bulky substituent on highest numbered carbon points up Anomers rapid equilibrium

5. Cyclization - aldopentose Equilibrium Anomeric C Hemiacetal Haworth projection Anomers

6. Glycolysis: Steps 2 and 3 Opens the chain during the rxn PFK-1 CH2OH OH utilizes 100% -anomer Stereospecific: uses -Glc; produces 100% -D-fructose-6-phosphate 36% -fructose 64% -fructose

7. Glycoside Bonds – Disaccharides No open chain equil Hemiacetals -a reactive carbonyl that can be oxidized. reducing non-reducing b anomer: refers to free C1 OH non-reducing sugar

8. Glycoside Bonds – Disaccharides epimer Most abundant disacc. in nature (plants)

9. Humans don’t have b-glucosidases Microbe that live in ruminants do Polysaccharides –Structure Amylose • Cellulose • b-(1-4) linkage 180 deg rotation 300- 15,000 Glc residues Rigid extended conformation H-bonding Forms bundles or fibrils Plant cell walls, stems and branches termites

10. Plant starch – mixture of amylose and amylopectin Animals glycogen Polysaccharides –Glucose Storage • Homoglycans- one type of monosaccharide Amylose • 100-1000 glucose residues (maltose units) Amylopectin and Glycogen Amylopectin: branch every 25 residues Glycogen: branch every 8-12 residues 10% mass of liver • No template (ie no gene)

11. Humans digest starch via two enzymes: α -amylase - endoglycosidase of α-(1-4) linkages (random) debranching enzyme (cleaves limit dextrans) Higher plants have β- amylase exoglycosidase of α- (1-4) linkages, releasing the disaccharide maltose Polysaccharides -Starch Degradation Know how starch is broken down ! Single reducing end multiple non-reducing end

12. Glycogen Metabolism Synthesis: Different enzymes for syn and degradation Driven by PPi hydrolysis Major regulatory step

13. Key regulation by phosphorylation (hormonally regulated) Pre-existing Glycogenin primer Amylo-(1,4 1,6)-transglycolase catalyzes the branch point. (Alpha 1-6 link)

14. Degradation: Two subunits, two catalytic sites, allosteric sites. AMP- activator; ATP & Glc-6-P – inhibitor. Phosphorylation: active (phosphorylase a). Dephosphorylated: less active (phosphorylase b). Phosphorolysis rxn. Generates phosph-sugar not free glc Primary regulation

15. Branching inc speed of syn and degradation phosphorolytic Reg by ATP and G-6-P Sequential removal of Glc From non-reducing end Stops 4 Glc from branch pt Primarily by phosphorylation Consequences of branch Reducing vs non-reducing ends solubility Rate of syn/degradation Energy yield from glycogen Higher than from glc

16. Human diseases of carbohydrate metabolism Inherited enzyme deficiencies Mutations that change enzyme function or abolish enzyme activity Most are recessive since only one functional copy of gene is sufficient for needed activity Diabetes Lactose intolerance Galactosemia Glycogen storage disease

17. α -amylase lactase Glc + Gal

18. Absorbed from intestine Major source of energy for nursing animals In liver 20% of caloric intake of infants Glc-6-P

20. Glucose metabolism lactase Glc + Gal Glc glucogen Glc glucogen Glc 1-P Glc 6-P Fruc 6-P Fruc 1,6-P

21. X Absorbed from intestine Major source of energy for nursing animals In liver 20% of caloric intake of infants X Two inherited metabolic errors Glc-6-P Hypolactasia (lactose intolerance) Galactosemia

22. Lactose Intolerance X Single gene defect Normal decrease in enzyme by 6 yrs old 10% of original activity (Northern Europeans are lactase producing adults) Mutation in chromosome 2 Lactase deficient people: Lactose passes intact into colon bacteria in colon ferment to lactic acid, methane and H gas Bloated/ gas and diarrhea Can also hinder absorption of other nutrients Avoid dietary lactose Take enzyme substitute

23. Galactosemia X Galactose 1-P accumulates in liver cells (high galactose in blood and urine) Decrease liver function and cataracts death CNS damage and mental retardation (even if avoid milk) cataracts Cataracts (clouding) due to high galactose in eye. Converted to galactol allowing diffusion of water into eye

24. Glucose metabolism glucogen Glc Glc glucogen Glc 1-P Glc 6-P Fruc 6-P Fruc 1,6-P

25. Glycolysis and Cancer Defined: Glucose is converted anaerobically to the three carbon acid pyruvate Net Reaction: Glucose + 2 ADP + 2 NAD+ + 2 Pi 2 Pyruvate + 2 ATP + 2 NADH + 2 H+ + 2 H2O Generates ATP at higher rate than Oxid Phosp Oxidative phosphorylation: allows more energy extracted from Glc Otto Warburg-cancer cells utilize glycolysis even in presence of O2 Aerobic glycolysis: Warburg effect Initial stages of tumor growth vessels grow at slower rate: cells deprived of O2 Cells switch to reliance to glycolysis Max energy when pyruvate from glycolysis enters Citric Acid Cycle

26. Glycolysis and Cancer Continue to rely of Glycolysis even when O2 restored to tumor Can visualize tumors based on inc sugar uptake (PET scan) Treatment?? Blocking lactose dehydrogenase (block NAD regeneration turn off Glycolysis)

27. Glucose ATP Phosphorylation Hexokinase ADP Glucose-6-phosphate Know key reg steps! Isomerization Glucose-6-phosphate isomerase Fructose-6-phosphate ATP Phosphorylation Phosphofructokinase-1 ADP Fructose-1,6-bisphosphate Cleavage Aldolase Dihydroxyacetone phosphate Isomerization Trios phosphate isomerase Glyceraldehyde-3-phosphate Glyceraldehyde-3-phosphate NAD+ + Pi Oxidation and Phosphorylation Glyceraldehyde-3-phosphate dehydrogenase NAD+ + Pi NADH + H+ NADH + H+ 1,3-bisphosphoglycerate 1,3-bisphosphoglycerate ADP Substrate Level Phosphorylation ADP Phosphoglycerate kinase ATP ATP 3-phosphoglycerate 3-phosphoglycerate Phosphoglycerate mutase Rearrangement 2-phosphoglycerate 2-phosphoglycerate Enolase H2O Dehydration H2O phosphoenolpyruvate phosphoenolpyruvate ADP ADP Substrate Level Phosphorylation ATP Pyruvate kinase ATP pyruvate pyruvate

28. Enzymatic Regulation of Glycolysis Not moving forward, stop converting ATP Cellular rxns are converting ATP and ADP, make more ATP You’ve committed! Bi-phosphated furanoses, keep pathway moving CAC intermediates, slow down, there is already adequate supply of energy

29. Glycolysis: Hexokinase Isozymes Hexokinases (I-III) -regulated negatively by Glc-6-P -if later steps slow down, Glc-6P builds up Glucokinase (IV) in Liver -regulated negatively by Fru-6-P -pulls glucose out of bloodstream until equil -liver can produce more Glc-6-P -converts Glucose to Glycogen storage I-III IV Isozymes Different inhibition profiles Location, Km Control point Glc Glc glucogen Glc 1-P Glc 6-P Can’t leave the cell with negative charge Fruc 6-P Fruc 1,6-P

30. in Liver Hormones Involved High blood [Glc], insulin released Low blood [Glc], glucagon released Insulin Dependent Uptake Muscle Adipose Major function of liver: maintain constant level of Glc in blood Release Glc (from glycogen) during muscle activity and between meals

31. Glucose 6-phosphatase Most cases Glc-6-P is end product---used in other pathways - glycogen, starch, pentose, hexose synthesis Enzyme only found in liver, kidney, small intestines Bound to ER with active site towards lumen Hydrolysis of phosphate irreversibly forms glucose Secretory pathway exports to blood stream for other tissues

32. Body does not transfer pyruvate Lactate- produced in RBC and Muscle Lactate to pyruvate in liver Cori cycle Major function of liver: maintain constant level of Glc in blood Release Glc during muscle activity and between meals Breakdown of glycogen to Glc 6-P (does not leave the cell) Liver contains glc 6-phoshatase enzyme Glc not major fuel in liver

33. Regulation of Phosphofructokinase-1 Large oligomeric enzyme bacteria/mammals - tetramer yeast - octamer ATP - product of pathway - allosteric inhibitor AMP - allosteric activator - relieves inhibition by ATP Citrate - feedback inhibitor - regulates supply of pyruvate - links Glycolysis and CAC Fru-2,6-bisphosphate - strong activator - produced by PFK-2 when excess fru-6-phosphate - indirect means of substrate stimulation or feed forward activation

34. Regulation of Pyruvate Kinase + F 1,6 BP Inactivation by covalent modification -blood [Glc] drops, glucagon released -liver protein kinase A (PKA) turned on -PKA phosphorylates pyruvate kinase Allosteric (feed-forward) activation Fructose-1,6-bisphosphate -allosterically activates -produced in step three -links control steps together Allosteric inhibition by ATP -product of pathway and CAC Low blood [Glc] High blood [Glc]

35. Regulation of Glycogen Metabolism Hormonal Regulation: Fed state fasting Via cAMP Via PIP3 Insulin: secreted by pancreas when Glc high inc rate of transport into cell and glycogen syn GLUT4 Glucagon: secreted when Glc low Epi: released by adrenal gland in response to neural signal (flight or flight) Sudden energy response

36. Gluconeogenesis Liver glycogen PPP - 3 places differ- control points in glycolysis - 4 new enzymes ATP energy, NADH reducing equivalents consumed Glc also syn from pyruvate (lactate and amino acids) Liver/kidney Glc needed in brain/muscle

37. Gluconeogenesis: Regulation Modulate one enzyme and affect 2 opposing pathways Sensitive regulatory point Low [Glc]: glucagon increases protein kinase A (activates Fru-2,6-bisP phosphatase) lowering [Fru-2,6-bisP]. Activate Glc syn and Loss of glycolysis stim

38. Regulation of Glycogen Metabolism Hormonal Regulation: Fed state fasting Via cAMP Via PIP3 Insulin: secreted by pancreas when Glc high inc rate of transport into cell and glycogen syn GLUT4 Glucagon: secreted when Glc low Epi: released by adrenal gland in response to neural signal (flight or flight) Sudden energy response

39. Intracellular Regulation of Glycogen Metabolism by Interconvertible Enzymes: Low glc activate kinase and breakdown Simultaneous effect Low [Glc]

40. High [Glc]

41. Human diseases of carbohydrate metabolism Inherited enzyme deficiencies Mutations that change enzyme function or abolish enzyme activity Most are recessive since only one functional copy of gene is sufficient for needed activity Diabetes Lactose intolerance Galactosemia Understand how enzyme deficiency leads to accumulation of glycogen Glycogen storage disease Other symptoms Treatment, if any

42. Glycogen storage disease glucogen Glc X V II, III, VI X X Glc glucogen Glc 1-P Glc 6-P I IV Fruc 6-P All defects lead to glycogen accumulation X VII Fruc 1,6-P I Glucose-6-Phosphatase in liver (von Gierk’s disease): hypoglycemia (low blood glc) when fasting Liver enlargement IV Branching enzyme in organs (liver) (Andersen’t disease) Liver dysfunction and early death V Glycogen phosphorylase in muscle (McArdle’s disease) Muscle cramps with exercise VII Phosphofructokinase in muscle Glycogen accumulation and Inability to exercise

43. Glycogen storage disease Type I: Glucose-6-Phosphatase deficiency in liver (von Gierk’s disease): Glc not released into blood No response to Epinephrine or Glucagon hypoglycemia (low blood glc) between meals infant in convulsions Large amounts of glycogen in liver (G-6-P inhibits breakdown) Liver enlargement Glc-6-P increases glycolysis inc lactate/pyruvate in blood (Lactic acidosis) Delayed puberty, short stature Continuous feedings of cornstarch (intragastric feeding) Drug induced inhibition of Glc uptake by liver Surgical transplant of portal vein (normally intestine-liver) Glc to peripheral tissues before liver

44. Glycogen storage disease Type IV: Most severe disease Branching enzyme deficiency in organs (liver) (Andersen’s disease) Accumulate abnormal glycogen Reduced solubility of glycogen Foreign body immune response?? Liver dysfunction Failure to thrive----- death 2-5 yrs old

45. Glycogen storage disease Type V: Glycogen phosphorylase deficiency in muscle (McArdle’s disease) No breakdown of glycogen Exercise indices muscle cramps otherwise normal Effective utilization of muscle glycogen not essential to life Can’t provide fuel for glycolysis to keep up Demand for ATP NMR on forearm muscle Muscle cramps correlate with inc ADP Vasodialation-muscle now has access to Glc and fatty acids in blood