Glycolysis and the Pentose Phosphate Pathway

1. To be lectured by Professor Zengyi Chang on September 19, 2007 Chapter 14Glycolysis and the Pentose Phosphate PathwayGlycolytic pathway;Fermentation;substrate channeling;Pentose phosphate pathway.

2. The chemistry of glucose catabolism was elucidated over about 100 years • The anaerobic breakdown (fermentation) of glucose to ethanol and CO2 by yeast has been exploited for many centuries in baking and winemaking. • Scientific investigation of the chemistry of glucose breakdown began in the mid-19th century. • The complete pathway was described around 1940.

3. D-glucose occupies a central position in metabolism (fat) The metabolic fates of glucose involves hundreds or thousands of chemical transformations. Other biomolecules Biosynthesis D-glucose G'o = –2840 kJ/mole ATP NADPH + CO2 + H2O

4. Understanding sugar catabolism occupied a central position in the development of biochemistry • Wherecell-free fermentationwith yeast extract was first observed by Buchner (1897): the vitalistic dogma was shaken and metabolim became chemistry. • Wherephosphorylated intermediates were first discovered (Harden and Young, 1900s). • Heat-labile, nondialyzable enzymes were distinguished from the heat-stable, dialyzable coenzymes (e.g., NAD+). • Muscle enzymes involved in the production of lactic acid were found to be astonishingly similar to the yeast enzymes involved in alcohol production. • ATPwas revealed to be an energy currency.

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8. The whole pathway of glycolysis (arbitrarily defined as from glucose to pyruvate) containing ten steps of chemical reactions with each catalyzed by a specific enzyme, was elucidated by the 1940s.

9. Glucose is first phosphorylated at C-6: consuming ATP Mg2+ATP2- is the actual substrate 己糖激酶 (Present in all cells of all organisms) Glucose is thus trapped and destabilized. An exergonic group transferring reaction

10. Inactive conformation Hexokinase exhibits induced fit property: The binding of glucose in the active site causes a major conformational change.

11. Active conformation Glucose Substrate-induced cleft closing is a general feature of all kinases!

12. Glucose 6-P is then converted to fructose 6-P: via isomerization A necessary prelude for the next two steps of reactions (phosphorylation and C-C cleavage). A ketose An aldose 磷酸己糖异构酶 A reversible Isomerization reaction (intramolecular redox reaction)

13. Fructose 6-P is then phosphorylated at C-1: ATP consuming (PFK-1) 磷酸果糖激酶-1 Another exergonic group transferring reaction

14. The six-carbon fructose 1,6-bisphosphate is then cleaved into two different triose phosphates 4 1 2 5 6 3 醛缩酶 The “lysis” step

15. Interconversion between dihydroxyacetone phosphate and glyceraldehyde 3-P then occurs Other hexoses (e.g., Fru., Man., Gal.) are also converted to glyceraldehyde 3-phosphate to enter glycolysis. An aldose (C3 or C4) A ketose (C5 or C2) (C1 or C6) 磷酸丙糖异构酶 A reversible Isomerization reaction (intramolecular redox reaction)

16. Glyceraldehyde 3-P is then oxidized to form 1,3-bisphosphoglycerate Tetrameric An acyl phosphate (a type of anhydride) 3H 3H 32P 32P 3-磷酸甘油醛脱氢酶 Oxidation (dehydrogenation)followed by phosphorolysis

17. A Cys residue is involved in the catalysis of glyceraldehyde 3-P dehydrogenase Inactivation of the enzyme Phosphorolysis step Oxidation step NADH exchanged for NAD+ HB HB HB (An acyl-enzyme intermediate) :B 3-磷酸甘油醛脱氢酶

18. The phosphoryl group linked to the carboxyl group of 1,3-bisphosphoglycerate is then transferred to ADP to form ATP substrate-level phosphorylation (vs. respiration-linked phosphorylation) ATP ADP 磷酸甘油酸激酶 (This enzyme was named for the reverse reaction)

19. 3-phosphoglycerate is then converted to 2-phosphoglycerate 磷酸甘油酸变位酶 Mutase: catalyzes group transferring from one position to another in a molecule.

20. The phosphoglycerate mutase acts as a transient carrier of phosphoryl groups Phosphoglucomutase acts in a similar fashion! (Initially phosphorylated by using 2,3-BPG)

21. 2-phosphoglycerate is then dehydrated to produce phosphoenolpyruvate A Redistribution of energy A super high-energy phosphate compound 烯醇酶 (PEP) • G`0for the hydrolysis of the phosphate group is changed from –17.6 to –61.9 kJ/mol.

22. The phosphoryl group of PEP is transferred to ADP to form ATP at the end ADP ATP 丙酮酸激酶 This enzyme was also named for the reverse reaction Nonenzymatic conversion The second substrate-level ADP phosphorylation!

23. Major catabolic fates of pyruvate Pyruvate also acts as precursors in many anabolic reactions.

24. The first stage of the glycolytic pathway: The preparatory phase priming isomerization priming cleavage Glucose is activated (or primed), with 2 ATP molecules invested.

25. The second stage of the glycolytic pathway: The payoff phase Only ~5% of the potential energy of the glucose molecule is released during glycolysis. All the enzymes catalyzing glycolysis have been found in the cytosol. The net production of ATP per glucose in glycolysis is 4-2=2.

26. The pathways of carbons, phosphoryl groups, and electrons for glycolysis:

27. Glycolytic enzymes may form multienzyme complexes within cells • When proteins are purified from extracts of broken cells in diluted solutions, noncovalent interactions between proteins could be destroyed (i.e., higher level organization destroyed). • Kinetic and physical evidences suggest that the enzymes act to catalyze the ten reactions of glycolysis pathway (as enzymes act in other metabolic pathways) may assemble into multienzyme complexes, where intermediates are directly channeled from one enzyme to another, without entering the aqueous solutions, a phenomenon called “substrate channeling”.

28. Substrate channeling The glycolytic enzymes very likely form multienzyme complexes. Dilution dissociation

29. Fermentation: pyruvate is converted to lactic acid or ethanol under anaerobic conditions

30. Lactic acid Fermentation: Pyruvate is reduced to lactate, leading to NAD+ regeneration when O2 lacks (thus allowing glycolysis to continue); occurring in very active skeleton muscle, some bacteria like lactobacilli 乳酸脱氢酶

31. Ethanol Fermentation: Pyruvate be decarboxylated and reduced to form ethanol in some microorganisms 丙酮酸脱羧酶 Present only in those alcohol fermentative Organisms 乙醇脱氢酶 Present in many organisms including human

32. Thiamine pyrophosphate (TPP) is involved in the cleavage of C-C bonds adjacent to a carbonyl group. Vitamin B1 (thiamine) is used to synthesize TPP. A lack of Vitamin B1 in humans will result in a condition known as Beriberi

33. Many carbohydrates meet their catabolic fate in glycolysis • Polysaccharides such as starch and glycogen are first degraded into glucoses via hydrolysis in extracellular spaces (catalyzed by a-amylases and other enzymes), but into glucose 1-phosphate via phosphorolysis inside cells (catalyzed by phosphorylases). • Oligosaccharides (e.g., sucrose, lactose, trehalose etc) are degraded into monosaccharides before further transformed. • Hexoses other than glucose can also be catabolized via the glycolytic pathway after being converted to a phosphorylated derivative.

35. HOH a-amylase OH

37. Pentose phosphate pathwayconverts glucose to specialized products needed by cell biosynthesis: NADPH and Ribose 5-phosphate This is primarily an anabolic pathway.

38. To counter the damaging effects of oxygen radicals For Reductive biosynthesis Some biological roles served by PPP Nucleotide biosynthesis

39. Glucose Glycogen PPP in rapidly dividing cells Cyclic ester

40. The Non-oxidative phase of PPP Isomerization & epimerization of Ru5P Ribulose 1,5-bisphophate is regenerated also via this pathway in the Calvin cycle. Carbon-carbon bond cleavage and formation TPP TPP Glycolysis 转二羟丙酮基酶 转羟乙醛酶

41. Two other ways of displaying the PPP

42. TPP is involved in the C-C cleavage and formation reaction catalyzed by transketolase: transferring two carbons from a ketose to an aldose.

43. Transaldolase catalyzes a reaction in which a three-carbon unit is transferred from a ketose to an aldose without being helped by any cofactor.

44. G-6-P is partitioned between glycolysis and PPP

45. Summary (1) • D-glucose is a commonly used fuel and versatile precursor in almost all organisms. • The study of glucose degradation has a rich history in biochemistry (especially for enzymology). • Glucose is first converted into two three-carbon pyruvates via the ten-step glycolysis pathway without directly consuming O2 and with a net production of two ATP molecules by substrate-level phosphorylation. • Limited amount of energy can be released by oxidizing glucose under anaerobic conditions by fermentation.

46. Summary (2) • The enzymes participating glycolysis may form multiple enzyme complexes, where substrate is channeled from one enzyme to another. • The sugar units on glycogen is converted to glucose 1-phosphate via phosphorolysis, which is catalyzed by glycogen phosphorylase. • Glucose 6-phosphate can also be oxidized to form ribose 5-phosphate and NADPH via the pentose phosphate pathway.