1. Glycolysis: Energy Generation Without an Oxygen Requirement
Glucose Biofuel Prominence:
Low-reactive ring-form minimizes protein glycosylation

2. Glycolysis: A Three Step Process
Glucose trapping and destabilization (priming)
Three carbon unit generation (cleaving)
Energy generation

3. Induced Fit in Hexokinase
Glucose induces a large enzyme conformational change
Substrate-induced cleft
closing prevents ATP hydrolysis
Kinases require a divalent metal ion
What function does Mg+2 play in hexokinase?

4. Hexokinase Closed Around Substrates
What mechanisms of catalysis are operative?

5. Hexokinase Reaction Mechanism
What is the Nu:, electrophile, and leaving group in this reaction?

6. Phosphoglucose Isomerase: Aldose to
Ketose Conversion

7. PGI Reaction Mechanism
Phosphoglucose Isomerase (PGI) G6P Conversion via Acid-Base Catalysis
PGI Reaction Mechanism

8. Phosphoglucose Isomerase (PGI) G6P Conversion via Acid-Base Catalysis
Base catalyzed
bond formation

9. Phosphoglucose Isomerase (PGI) G6P Conversion via Acid-Base Catalysis
Acid catalyzed
ketal formation

10. Base catalyzes ring closure
Phosphoglucose Isomerase (PGI) G6P Conversion via Acid-Base Catalysis
Base catalyzes ring closure H+

11. Phosphofructokinase: Trapping the Fructose Isomer
What is the mechanism for this reaction?

12. Glycolysis Stage I: Glucose Trapping and Destabilization (priming)

13. Six Carbon Sugar Cleaved to Two Three Carbon Units
What is the bond to be cleaved?
Which alcohol becomes an aldehyde?

14. Haworth and Fischer Projections
Equivalency
The functional group that is down in a Haworth projection is positioned how in a Fischer structure?

15. Aldolase Reaction Mechanism
Fructose-1,6-bisphosphate binds to the aldolase enzyme for covalent catalysis

16. Aldolase Reaction Mechanism
What is lost when the Schiff base forms?

17. Aldolase Rxn Mechanism
Aldolase Reaction Mechanism
Aldolase Rxn Mechanism
Compare and contrast a Schiff base with a carbonyl group.

18. Aldolase Reaction Mechanism
What is the process for Schiff base to carbonyl conversion?

19. Aldolase Reaction Mechanism
H2O
Aldolase cleaves FBP into GAP and DHAP

20. Triose Phosphate Isomerase (TIM)
Reversible and driven towards GAP due to product depletion
Which previous glycolytic step is similar to TIM?

21. Glycolysis: Step #5 Triose Phosphate Isomerase
Triose Phosphate Isomerase Reaction Mechanism
Glycolysis: Step #5 Triose Phosphate Isomerase
TIM- or α,β-barrel with 8 parallel β-strands surrounded by 8 α-helices.
DHAP conversion to GAP necessary to proceed through glycolysis

22. Stoichiometry: Stages 1-2 of Glycolysis
Two ATPs are initially invested.
One glucose is metabolized into two GAP molecules.

23. Road Map for Energy Harvest (Stage 3)

24. Glyceraldehyde-3-Phosphate Dehydrogenase: Covalent Catalysis

25. Glyceraldehyde-3-Phosphate Dehydrogenase: a 2 Step Process
What amino acid will serve as a nucleophile to form a thioester?

26. Glyceraldehyde-3-Phosphate Dehydrogenase:
Reaction
Mechanism

27. Actual coupled reaction
Glyceraldehyde-3-Phosphate Dehydrogenase: Catalysis Energetics
Hypothetical reaction with no coupling
Actual coupled reaction

28. Phosphoglycerate Kinase
What is the Nu:, electrophile and leaving group for this reaction? (hint: consider hexokinase in reverse)

29. Glycolysis: the Three Final Steps

30. Pyruvate Kinase
What is the Nu:, electrophile and leaving group for this reaction? (hint: consider phosphoglycerate kinase)

31. Glycolysis Energetic ∆G°ʹ ∆G Enzyme (kcal/mol) (kcal/mol)
1near equilibrium means that ∆G is about zero
What is the relationship between ∆G and ∆G°ʹ?
When can ∆G and ∆G°ʹ diverge?

32. Regulating Glycolysis: A Pictorial Analogue
Water represents
metabolite flux
Water amount in
flask represents
intermediate
abundance
Flasks connections
are enzymes
Vertical drop represents
decrease in free energy
ΔG° = height difference between flask bottoms
ΔG = height difference between water levels

33. Metabolic Regulation
Irreversible reactions are potential regulatory sites (e.g. hexokinase, phosphofructokinase and pyruvate kinase)
What duel role does ATP play in PFK-1 catalysis?
In what direction does ATP regulate phosphofructokinase?

34. Energy Status Regulates Glycolytic Flow
Elevated [ATP] sufficient energy; elevate [AMP] low energy
ADP + ADP ↔ ATP + AMP <adenylate kinase>
Muscle Tissue

35. Fructose-2,6-Bisphosphate an Allosteric Regulator of Phosphofructokinase-1
PFK-2
Liver Tissue
Front activation by fructose-6P
F-2,6-BP amplifies or diminishes PFK-1 activity?

36. Fructose-2,6-Bisphosphate Reduces ATP Inhibition of Phosphofructokinase-1
PFK-2
Liver Tissue
ATP is a substrate and inhibitor of PFK-1

37. Fructose Entry Points for Glycolysis
Glycerol-3P
Glucose + Fructose
Major dietary sugars: sucrose (table sugar) and fructose (high-fructose corn syrup)

38. Fructose Metabolism
How is
this
different
than
glucose metabolism?

39. Fructose Metabolism
Glycerol 3-phosphate a precursor to triacylglycerol
Fructose catabolism bypasses
phosphofructokinase regulation
Glycerol
3-Phosphate
Lipid Synthesis

40. Alternative Fates for Pyruvate

41. Anaerobic Recycling of NADH for Glycolysis

42. Microbial Recycling of NADH for Glycolysis

43. Pyruvate Dehydrogenase: the Bridge between Glycolysis and Citric Acid Cycle

44. Standard Free Energy Change Comparisons for Glucose Catabolism
With and Without Oxygen

45. Pathogenic Obligate Anaerobes

46. Pyruvate Targeted for Anabolism
The biotin prosthetic group serves as a CO2 carrier
What reaction links biotin to the protein?

47. Pyruvate Carboxylase: an Endergonic Reaction
Oxaloacetate

48. Glucose Metabolism: Both Catabolic and Anabolic

49. Glucose Metabolism: Both Catabolic and Anabolic

50. Problems: 1, 3, 5, 7, 13, and 21