1. Pratt and Cornely, Chapter 13
Glucose Metabolism
Pratt and Cornely, Chapter 13

2. Glycolysis Expectations
Memorize/learn Figure 13.2
Know overall reaction and stages
Explain chemical logic of each step
Enzyme mechanisms presented in book

3. Glycolysis Ten enzymes that take glucose to pyruvate Cytosol
ATP and NADH

4. Reactions and Enzymes of Glycolysis
ATP
ATP
Pi + NAD+ 2x
ADP
ADP
NADH
ADP
ADP 2x 2x 2x
ATP
ATP
Hexose and triose phases
Energy input and payoff phases

5. Energy Input

6. Energy Payoff

7. Know...
Substrates
Co-substrates
Products
Enzyme names

8. 1. Hexokinase Previous concepts: Induced fit, kinase Utilizes 1 ATP
Chemical logic?

9. Problem 3
(Notice miswording) The DGo’ value for hexokinase is kJ/mol, but it is twice as spontaneous under cellular conditions.
What is the ratio of G-6-P to glucose under cellular conditions when the ratio of ATP:ADP is 10:1?
How high would the ratio of G-6-P to glucose have to be to reverse the hexokinase reaction by mass action?

10. 2. Phosphoglucose Isomerase
Previous concepts: Isomerization
CONCEPT: Near-equilibrium
Chemical logic:
Stereochemistry—reverse does not produce mannose!

12. 3. PFK-1 Previous concepts: Allosteric inhibition Utilizes 1 ATP
Pathway logic:
First committed step of glycolysis
Why?
regulation

13. Regulation: Bacteria vs. Human

14. 4. Aldolase
Previous concepts: Standard free energy is +23kJ, but it is a near equilibrium reaction
Pathway logic:
Beginning of triose stage

15. Aldolase Mechanism

16. 5. Triose Phosphate Isomerase
Previous concepts: Catalytic perfection
Pathway logic:
Most similar to which previous reaction?

17. 6. Glyceraldehyde-3-P DH Previous concepts: Redox and dehydrogenase
Pathway logic: Utilizes negative free energy of _____________ to drive nonspontaneous formation of ___________________

18. GAPDH Mechanism Be able to draw mechanism with full NAD+ structure

19. 7. Phosphoglycerate Kinase
Previous concepts: High energy bond
Produces 2 ATP: substrate level phosphorylation
Pathway logic?
Coupled to reaction 6 by Le Chatelier

20. Coupled Reactions GAPDH = 6.7 kJ/mol PG Kinase = -18.8 kJ/mol Overall:
A set of near-equilibrium reactions at cellular concentration

21. 8. Phosphoglycerate Mutase
Previous concepts: Covalent catalysis
Pathway logic:
Mutase—isomerization with P transfer

22. Mechanism Not a simple transfer
What happens if the bisphosphate escapes?

23. 9. Enolase Concept: Phosphoryl group transfer potential
Chemical logic?

24. 10. Pyruvate Kinase Production of 2 ATP
VERY high energy bond allows formation of _________ while still being irreversible
Regulation: F-1,6-BP can act as a feed-forward activator to ensure fast glycolysis

25. Overall Energetics Standard Free energies are up and down
Free energies under cellular conditions are downhill or near zero
Three irreversible

26. Fate of Pyruvate Amino acid and nitrogen metabolism Gluconeogenesis
Aerobic Energy
Anaerobic in
higher organisms
Anaerobic in
microorganisms

27. The Problem of Anaerobic Metabolism
With oxygen, the NADH produced in glycolysis is re-oxidized back to NAD+
NAD+/NADH is a co-substrate which means…
If there is no oxygen, glycolysis will stop because…
The solution to the problem is to…

28. The solution in Yeast Pyruvate is decarboxylated to acetaldehyde
Acetaldehyde transformed to ethanol
What type of reaction?
What cofactor?
NAD+ is regenerated to be reused in GAPDH

29. The Solution in Us Lactate formation: Draw a mechanism
Balanced equation

30. We don’t operate anaerobically...
Most energy still trapped in lactate
Back to pyruvate, then acetyl-CoA
Citric acid cycle

31. Other sugars enter glycolysis
High fructose diet puts sugars through glycolysis while avoiding major regulation step

32. Glucose Metabolism Overview
Keep the main pathway purposes distinct
But learn details of chemistry and regulation based on similarities

33. Glucose Metabolism Overview
Gluconeogenesis
Glycogen metabolism
Pentose Phosphate Pathway

34. Precursors for Gluconeogenesis
Names of compounds?
Type of reaction?
Type of enzyme?
Cofactor(s)?

35. Chemistry of Gluconeogenesis
Pyruvate transformed to glucose
But energetically costly—no perpetual motion machine!
Points of regulation

36. Glycolysis Gluconeogenesis Step 1: costs 1 ATP Step 3: costs 1 ATP
Step 7: makes 2 ATP
Step 10: makes 2 ATP
Gluconeogenesis
Step 10: no ATP back
Step 8: no ATP back
Step 3: costs 2 ATP
Step 1a/b: costs 4 ATP equivalents

37. Step 1a Pyruvate Carboxylase Biotin
Costs ATP to make driving force for next reaction
TIMES TWO!
First step in biosynthesis of glucose and many other molecules
Related to which amino acid?

38. Mechanism Mixed anhydride activates carboxyl group
Coupled through biotin coenzyme
Enolate attacks activated carboxylate

39. Step 1b PEP carboxykinase ATP cost to restore PEP CO2 loss drives rxn
TIMES TWO!
CO2 loss drives rxn

40. Step 8 Fructose-1,6-bisphosphatase No additional energy input
Phosphate ester hydrolysis is spontaneous

41. Step 10
Glucose 6-phosphatase
Liver (and others)
Not in muscle

42. Problem 34
A liver biopsy of a four-year old boy indicated that the F-1,6-Bpase enzyme activity was 20% normal. The patient’s blood glucose levels were normal at the beginning of a fast, but then decreased suddenly. Pyruvate and alanine concentrations were also elevated, as was the glyceraldehyde/DHAP ratio. Explain the reason for these symptoms.

43. Key Regulation At the committed step in glucogenic cells
Principle of Reciprocal Regulation
Local regulation vs Hormone regulation

44. Key Regulation Local regulation Hormone regulation
AMP/ATP (energy charge)
Citrate (feedback)
Hormone regulation
Fructose-2,6-bisphosphate
Gluconeogenesis is inhibited
Glycolysis is stimulated

45. Problem 39
Brazilin, a compound found in aqueous extracts of sappan wood, has been used to treat diabetics in Korea. It increases the activity of the enzyme that products F-2,6-BP and stimulates the activity of pyruvate kinase. What is the effect of adding brazilin to liver cells in culture? Why would brazilin be an effective treatment for diabetes?

46. Glucose Metabolism Overview
Gluconeogenesis
Glycogen metabolism
Pentose Phosphate Pathway

47. Glycogen Storage molecule Primer necessary Very large!
Multiple ends allow for quick synthesis and degradation

48. Chemistry of Synthesis
Step 1
Near equilibrium
The link to glucose-6-phophate, our central molecule

49. Chemistry of Synthesis
Step 2
Count high energy bonds
Pyrophosphatase
Common motiff
UDP-glucose: activated for incorporation

50. Chemistry of Synthesis
Step 3
Glycogen synthase
Growing end is non-reducing
UDP released

51. Branching

52. Energetics of Synthesis
Total cost: one ATP equivalent from G-6-P

53. Four Degradation Enzymes
Two enzyme debranching process
Glycogen phosphorylase
phophoglucomutase

54. Chemistry of Degradation
Glycogen phosphorylase
Key Regulation site
Inorganic phosphate as a nucleophile
Remake G-1-P with no ATP cost

55. Debranching
Transfer branch
a(16)glycosidase

56. Overall Energetics

57. Key Regulatory Enzymes

58. Glycogen Storage Diseases
Many disrupt glycogen breakdown in muscle and/or liver
(hypoglycemia, enlarged liver, muscle cramps...)

59. Glucose Metabolism Overview
Gluconeogenesis
Glycogen metabolism
Pentose Phosphate Pathway

60. Pentose Phosphate Pathway
Dual Purpose
Synthesis of “reducing potential”
Synthesis of 5-carbon sugars
At cost of one carbon worth of carbohydrate
Net reaction:

61. Physiological Purposes

62. Glutathione: Protection from Oxidation
Glutathione is the redox buffer of the cell
Regenerated by NADPH
PPP especially important in RBC because it is only means to generate reducing power (no mitochondria)

63. Complex, 2-Stage Process
Oxidative Stage
Generates reducing power and ribose
Non-oxidative stage
Regenerates 3- and 6-carbon sugars from 5 carbon sugars

64. Oxidative Stage Step 1:
G-6-P DH
Lactone formation

65. Oxidative Stage Step 2: Also a spontaneous hydrolysis
Practice mechanism, carbohydrate orientation

66. Oxidative Stage Step 3: Oxidative decarboxylation
Which is first, oxidation or decarboxylation?
We will see this process again

67. Biosynthesis of Ribose

68. Non-oxidative Stage
To understand purpose, realize that we generally need to make much more NADPH than ribose
Problem: stuck with C5, but need C6 and C3
Solution: “Shunt” C5 back to C6 through near-equilibrium reactions

69. PPP Reactions
Epimerase
Isomerase
Transketolase
Transaldolase

70. Epimerase

71. Transketolase
Use cofactor (B1) to overcome chemical problem

72. Mechanism

73. Transketalase, Transaldolase
Transfer of an acyl anion
Requires TPP to stabilize unstable anion
Transaldolase
Transfer of somewhat stable enolate ion
Schiff base helps
Be able to recognize need for TK vs. TA

74. Transketalase or transaldolase?

75. Different Modes for Different Purposes

76. Problem 58
A given metabolite may follow more than one metabolic pathway. List all possible fates of glucose-6-P in (a) a liver cell and (b) a muscle cell.

77. Summary
of glucose
metabolism