1. Chapter 8 Carbohydrate Metabolism Overview Metabolism and Jet Engines
Section 8.1: Glycolysis
Section 8.2: Gluconeogenesis
Section 8.3: The Pentose Phosphate Pathway
Section 8.4: Metabolism of Other Important Sugars
Section 8.5: Glycogen Metabolism
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

2. Metabolism and Jet Engines
Catabolic pathways with a turbo step are optimized and efficient
Energy is fed back into the system to accelerate the fuel input step
Figure 8.1 Glycolysis and the Turbo Jet Engine
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

3. FIGURE 14-1 Major pathways of glucose utilization
FIGURE 14-1 Major pathways of glucose utilization. Although not the only possible fates for glucose, these four pathways are the most significant in terms of the amount of glucose that flows through them in most cells.

4. Chapter 8: Overview
Figure 8.2 Major Pathways in Carbohydrate Metabolism
Energy transforming pathways of carbohydrate metabolism include glycolysis, glycogenesis, glycogenolysis, gluconeogenesis, and pentose phosphate pathway
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

5. Glycolysis (anaerobic process) occurs in almost every living cell
Section 8.1: Glycolysis
Figure 8.2 Major Pathways in Carbohydrate Metabolism
Glycolysis (anaerobic process) occurs in almost every living cell
Ancient process central to all life
Splits glucose into two three-carbon pyruvate units
Catabolic process that captures some energy as 2 ATP and 2 NADH
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

6. Following the pathway
Carbons
H/electrons
Phosphate

7. Glycolysis is an anaerobic process
Section 8.1: Glycolysis
Glycolysis is an anaerobic process
Two stages (stage 1 and 2): energy investment and energy producing
Glycolytic Pathway: D-Glucose + 2 ADP + 2 Pi + 2 NAD+  2 pyruvate + 2 ATP + 2 NADH + 2 H+ + 2 H2O
In eukaryotes, the enzymes for this pathway are in the cytosol. They are all homodimers or homotetramers.
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press 7

8. Section 8.1: Glycolysis Figure 8.3 Glycolytic Pathway
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press 8

9. Section 8.1: Glycolysis Figure 8.3 Glycolytic Pathway
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press 9

10. Table 17-1 Standard Free Energy Changes (DG°¢), and Physiological Free Energy Changes (DG) in Heart Muscle, of the Reactions of Glycolysisa.
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11. FIGURE 14-3 Three possible catabolic fates of the pyruvate formed in glycolysis. Pyruvate also serves as a precursor in many anabolic reactions, not shown here.

12. Reactions of the Glycolytic Pathway
Section 8.1: Glycolysis
Reactions of the Glycolytic Pathway
1. Synthesis of glucose-6-phosphate
Phosphorylation of glucose (kinase) prevents transport out of the cell and increases reactivity
2. Conversion of glucose-6-phosphate to fructose-6-phosphate
Conversion of aldose to ketose
Figure 8.3a Glycolytic Pathway
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press 12

14. The nucleophilic attack of the C6—OH group of glucose on the g phosphate of an Mg2+–ATP complex.
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15. The phosphoryl-transfer reaction catalyzed by hexokinase.
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16. This same change in conformation is observed for ALL kinases!
Conformation changes in yeast hexokinase on binding glucose. (a) Space-filling model of a subunit of free hexokinase. (b) Space-filling model of a subunit of free hexokinase in complex with glucose (purple).
This same change in
conformation
is observed for
ALL kinases!
It also accounts for the
fact that water cannot be
used for hydrolysis of ATP
unless we fool the enzyme
by using xylose instead of glc. \

18. Phosphoglucose isomerase (PGI)
pKs for active site: 6.7 and 9.3
(determined by rate vs. pH)
Which aa’s??
Actually Glu (!!!) and His with stabilization of His+ by a Glu
(remember the ser protease mechanism!)

20. FIGURE 14-4 The phosphohexose isomerase reaction
FIGURE 14-4 The phosphohexose isomerase reaction. The ring opening and closing reactions (steps 1 and 4) are catalyzed by an active-site His residue, by mechanisms omitted here for simplicity. The proton (pink) initially at C-2 is made more easily abstractable by electron withdrawal by the adjacent carbonyl and nearby hydroxyl group. After its transfer from C-2 to the active-site Glu residue (a weak acid), the proton is freely exchanged with the surrounding solution; that is, the proton abstracted from C-2 in step 2 is not necessarily the same one that is added to C-1 in step 3.

21. Phosphofructokinase (PFK)
Works exactly like HK.
Inhibited by hi [ATP]
or citrate
Activated by [AMP] even in the presence of hi [ATP].

22. Reactions of the Glycolytic Pathway Continued
Section 8.1: Glycolysis
Reactions of the Glycolytic Pathway Continued
3. Phosphorylation of fructose-6-phosphate
This step is irreversible due to a large decrease in free energy and commits the molecule to glycolysis
4. Cleavage of fructose-1,6-bisphosphate
Aldol cleavage giving an aldose and ketose product
Figure 8.3a Glycolytic Pathway
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

24. FIGURE 14-5 The class I aldolase reaction
FIGURE 14-5 The class I aldolase reaction. The reaction shown here is the reverse of an aldol condensation. Note that cleavage between C-3 and C-4 depends on the presence of the carbonyl group at C-2. A and B represent amino acid residues that serve as general acid (A) or base (B).

25. Reactions of the Glycolytic Pathway Continued
Section 8.1: Glycolysis
Reactions of the Glycolytic Pathway Continued
5. Interconversion of glyceraldehyde-3-phosphate and dihydroxyacetone phosphate
Conversion of aldose to ketose enables all carbons to continue through glycolysis
Figure 8.3a Glycolytic Pathway
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

26. Figure 17-10 Proposed enzymatic mechanism of the TIM reaction: General Acid Catalysis.
pKs = 6.5 and 9.5
Like PGI
But pK1 is for GLU! Normal pk?
4.1
GluAsp  activity by 1000!
Reaction rate is
diffusion limited!!

27. End of Glycolysis Collection Phase
Net result so far?
ATP
NAD+
Carbon

28. FIGURE 14-6 Fate of the glucose carbons in the formation of glyceraldehyde 3-phosphate. (a) The origin of the carbons in the two three-carbon products of the aldolase and triose phosphate isomerase reactions. The end product of the two reactions is glyceraldehyde 3-phosphate (two molecules). (b) Each carbon of glyceraldehyde 3-phosphate is derived from either of two specific carbons of glucose. Note that the numbering of the carbon atoms of glyceraldehyde 3-phosphate differs from that of the glucose from which it is derived. In glyceraldehyde 3-phosphate, the most complex functional group (the carbonyl) is specified as C-1. This numbering change is important for interpreting experiments with glucose in which a single carbon is labeled with a radioisotope. (See Problems 6 and 9 at the end of this chapter.)

29. Reactions of the Glycolytic Pathway Continued
Section 8.1: Glycolysis
Reactions of the Glycolytic Pathway Continued
In Step 2 (reactions 6-10), each reaction occurs in duplicate
6. Oxidation of glyceraldehyde-3-phosphate
Creates high-energy phosphoanhydride bond for ATP formation and NADH
7. Phosphoryl group transfer
Production of ATP via substrate-level phosphorylation
Figure 8.3b Glycolytic Pathway (Stage 2)
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

30. FIGURE NAD and NADP. (a) Nicotinamide adenine dinucleotide, NAD+, and its phosphorylated analog NADP+ undergo reduction to NADH and NADPH, accepting a hydride ion (two electrons and one proton) from an oxidizable substrate. The hydride ion is added to either the front (the A side) or the back (the B side) of the planar nicotinamide ring (see Table 13-8). (b) The UV absorption spectra of NAD+ and NADH. Reduction of the nicotinamide ring produces a new, broad absorption band with a maximum at 340 nm. The production of NADH during an enzyme-catalyzed reaction can be conveniently followed by observing the appearance of the absorbance at 340 nm (molar extinction coefficient ε340 = 6,200 M–1cm–1).

32. Some reactions employed in elucidating the enzymatic mechanism of GAPDH. (a) The reaction of iodoacetate with an active site Cys residue. (b) Quantitative tritium transfer from substrate to NAD+.
Page 596
32Pi also incorporated

33. Section 8.1: Glycolysis
Figure 8.4Glyceraldehyde-3-Phosphate Dehydrogenase Reaction
Oxidation of glyceraldehyde-3-phosphate (G-3-P) is a 2-step process (reaction 6)
G-3-P undergoes oxidation and phosphorylation
G-3-P interacts with the sulfhydryl group in the enzyme’s active site
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

34. Section 8.1: Glycolysis
Figure 8.4 Glyceraldehyde-3-Phosphate Dehydrogenase Reaction
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

35. Section 8.1: Glycolysis
Oxidation of glyceraldehyde-3-phosphate (G-3-P) is a complex process (reaction 6)
Substrate oxidized after interaction with sulfhydryl
Bound NADH exchanged for NAD+
Enzyme displaced by addition of inorganic phosphate
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

36. Space-filling model of yeast phosphoglycerate kinase showing its deeply clefted bilobal structure ,3 BPG + ADP →3 PGA + ATP
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37. Reactions of the Glycolytic Pathway Continued
Section 8.1: Glycolysis
Reactions of the Glycolytic Pathway Continued
8. Interconversion of 3-phosphoglycerate and 2-phosphoglycerate
First step in formation of phosphoenolpyruvate (PEP)
9. Dehydration of 2-phosphoglycerate
Production of PEP, which has a high phosphoryl group transfer potential (tautomerization), locks it into the highest energy form
Figure 8.4b Glycolytic Pathway (Stage 2)
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

38. FIGURE 14-8 The phosphoglycerate mutase reaction.

39. The pathway for the synthesis and degradation of 2,3-BPG in erythrocytes is a detour from the glycolytic pathway.
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41. Figure 17-21 Proposed reaction mechanism of enolase.
F- binds Pi + Mg+2
Potent inhibitor
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44. Reactions of the Glycolytic Pathway Continued
Section 8.1: Glycolysis
Reactions of the Glycolytic Pathway Continued
10. Synthesis of pyruvate
Formation of pyruvate and ATP
Produces a net of 2 ATP, 2 NADH, and 2 pyruvate
Figure 8.3b Glycolytic Pathway (Stage 2)
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

45. Figure 17-22 Mechanism of the reaction catalyzed by pyruvate kinase.
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46. Let's sing!!

47. Lyrics

49. The product’s composition, 3-phosphoglycerate
From 3 to 2 position can readily mutate
And now 2 phosphoglycerage does something rather strange—
Electrons on and 3 proceed to rearrange.
Thus, redox-dehydration, catalysed by enolase
Gives PEP formation and bond energy raise
So phosphoenolphruvate reacts with ADP
The kinase making ATP but NOT reversibly.
In anaerobiosis, pyruvate’s not the end;
The problem we suppose is not hard to comprehend’
The dehydrogenation to phosphoglycerate
Would grind to halt if NAD+ could not regenerate.
The answer is quite subtle, pryruvayte is reduced,
Instead of malate shuttle, L-lactate is produced;
Lactate dehydrogenase performs that noble feat,
NADH is oxidised; the pathway is complete.
The balance sheet you’ll see shows transfer of energy,
Two ATPs from glucose, and three from G1P
That’s good, but oh to use the way where pyruvate’s reduced—
With decarboxylation first, then ethanol produced!!!!!

50. Section 8.1: Glycolysis The Fates of Pyruvate
Figure 8.6 The Fates of Pyruvate
The Fates of Pyruvate
Pyruvate is an energy-rich molecule
Under aerobic conditions, pyruvate is converted to acetyl-CoA for use in the citric acid cycle and electron transport chain
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

51. The Fates of Pyruvate Continued
Section 8.1: Glycolysis
The Fates of Pyruvate Continued
Under anaerobic conditions pyruvate can undergo fermentation: alcoholic or homolactic
Regenerates NAD+ so glycolysis can continue
Figure 8.7 Recycling NADH during Anaerobic Glycolysis
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

52. Energetics of Glycolysis
Section 8.1: Glycolysis
Figure 8.8 Free Energy Changes during Glycolysis in Red Blood Cells
Energetics of Glycolysis
In red blood cells, only three reactions have significantly negative DG values
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

53. Regulation of Glycolysis
Section 8.1: Glycolysis
Regulation of Glycolysis
The rate of the glycolytic pathway in a cell is controlled by the allosteric enzymes:
Hexokinases I, II, and III
PFK-1
Pyruvate kinase
Allosteric enzymes are sensitive indicators of a cell’s metabolic state regulated locally by effector molecules
The peptide hormones glucagon and insulin also regulate glycolysis
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

54. Regulation of Glycolysis Continued
Section 8.1: Glycolysis
Regulation of Glycolysis Continued
High AMP concentrations activate pyruvate kinase
Fructose-2,6-bisphosphate, produced via hormone- induced covalent modification of PFK-2, activates PFK-1
Accumulation of fructose-1,6-bisphosphate activates PFK-1 providing a feed-forward mechanism
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

55. Section 8.1: Glycolysis
Figure 8.9 Fructose-2,6-Bisphosphate Level Regulation
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

56. Section 8.2: Gluconeogenesis
Gluconeogenesis is the formation of new glucose molecules from precursors in the liver
Precursor molecules include lactate, pyruvate, and a-keto acids
Gluconeogenesis Reactions
Reverse of glycolysis except the three irreversible reactions
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

57. Section 8.2: Gluconeogenesis
Figure 8.10 Carbohydrate Metabolism: Gluconeogenesis and Glycolysis
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

58. Section 8.2: Gluconeogenesis
Figure 8.10 Carbohydrate Metabolism: Gluconeogenesis and Glycolysis
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

59. Section 8.2: Gluconeogenesis
Gluconeogenesis Reactions Continued
Three bypass reactions:
1. Synthesis of phosphoenolpyruvate (PEP) via the enzymes pyruvate carboxylase and pyruvate carboxykinase
2. Conversion of fructose-1,6-bisphosphate to fructose-6-phosphate via the enzyme fructose-1,6-bisphosphatase
3. Formation of glucose from glucose-6-phosphate via the liver and kidney-specific enzyme glucose-6-phosphatase
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

60. Section 8.2: Gluconeogenesis
Gluconeogenesis Substrates
Three of the most important substrates for gluconeogenesis are:
1. Lactate—released by skeletal muscle from the Cori cycle
After transfer to the liver lactate is converted to pyruvate, then to glucose
2. Glycerol—a product of fat metabolism
Figure 8.11 Cori Cycle
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

61. Section 8.2: Gluconeogenesis
Figure 8.12 The Glucose Alanine Cycle
Gluconeogenesis Substrates Continued
3. Alanine—generated from pyruvate in exercising muscle
Alanine is converted to pyruvate and then glucose in the liver
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

62. Section 8.2: Gluconeogenesis
Gluconeogenesis Regulation
Substrate availability
Hormones (e.g., cortisol and insulin)
Figure 8.13 Allosteric Regulation of Glycolysis and Gluconeogenesis
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

63. Section 8.2: Gluconeogenesis+
Gluconeogenesis Regulation Continued
Allosteric enzymes (pyruvate carboxylase, pyruvate carboxykinase, fructose-1,6-bisphosphatase, and glucose-6-phosphatase)
Figure 8.13 Allosteric Regulation of Glycolysis and Gluconeogenesis
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

64. Section 8.3: Pentose Phosphate Pathway
Alternate glucose metabolic pathway
Products are NADPH and ribose-5-phosphate
Two phases: oxidative and nonoxidative
Glucose-6-phosphate dehydrogenase
Gluconolactonase
Figure 8.14a The Pentose Phosphate Pathway (oxidative)
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

65. Section 8.3: Pentose Phosphate Pathway
6-phosphogluconate dehydrogenase
Pentose Phosphate Pathway: Oxidative
Three reactions
Results in ribulose-5-phosphate and two NADPH
NADPH is a reducing agent used in anabolic processes
Figure 8.14a The Pentose Phosphate Pathway (oxidative)
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

66. Section 8.3: Pentose Phosphate Pathway
Pentose Phosphate Pathway: Nonoxidative
Produces important intermediates for nucleotide biosynthesis and glycolysis
Ribose-5-phosphate
Glyceraldehyde-3-phosphate
Fructose-6-phosphate
Figure 8.14b The Pentose Phosphate Pathway (nonoxidative)
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

67. Section 8.3: Pentose Phosphate Pathway
If the cell requires more NADPH than ribose molecules, products of the nonoxidative phase can be shuttled into glycolysis
Figure 8.15 Carbohydrate Metabolism: Glycolysis and the Phosphate Pathway
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

68. Section 8.4: Metabolism of Other Important Sugars
Figure 8.16 Carbohydrate Metabolism: Galactose Metabolism
Fructose, mannose, and galactose are also important sugars for vertebrates
Most common sugars found in oligosaccharides besides glucose
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

69. Fructose Metabolism Section 8.4: Metabolism of Other Important Sugars
Second to glucose in the human diet
Can enter the glycolytic pathway in two ways:
Through the liver (multi-enzymatic process)
Muscle and adipose tissue (hexokinase)
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

70. Section 8.4: Metabolism of Other Important Sugars
Figure 8.16 Carbohydrate Metabolism: Other Important Sugars
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

71. Section 8.5: Glycogen Metabolism
Glycogenesis
Synthesis of glycogen, the storage form of glucose, occurs after a meal
Requires a set of three reactions (1 and 2 are preparatory and 3 is for chain elongation):
1. Synthesis of glucose-1-phosphate (G1P) from glucose-6-phosphate by phosphoglucomutase
2. Synthesis of UDP-glucose from G1P by UDP-glucose phosphorylase
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

72. Section 8.5: Glycogen Metabolism
Figure 8.17a Glycogen Synthesis
Glycogen synthase
Glycogenesis Continued
3. Synthesis of Glycogen from UDP-glucose requires two enzymes:
Glycogen synthase to grow the chain
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

73. Glycogenesis Continued
Section 8.5: Glycogen Metabolism
Branching enzyme
Glycogenesis Continued
Branching enzyme amylo-a(1,41,6)-glucosyl transferase creates a(1,6) linkages for branches
a(1,6) Glycosidic Linkage is formed
Figure 8.17b Glycogen Synthesis
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

74. Glycogenolysis Section 8.5: Glycogen Metabolism
Glycogen degradation requires two reactions:
1. Removal of glucose from nonreducing ends (glycogen phosphorylase) within four glucose of a branch point
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

75. Section 8.5: Glycogen Metabolism
Figure 8.18 Glycogen Degradation
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

76. Glycogenolysis Cont. Section 8.5: Glycogen Metabolism
Glycogen degradation requires two reactions:
2. Hydrolysis of the a(1,6) glycosidic bonds at branch points by amylo-a(1,6)-glucosidase (debranching enzyme)
Amylo-a(1,6)-glucosidase
Figure 8.19 Glycogen Degradation via Debranching Enzyme
Amylo-a(1,6)-glucosidase
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

77. Section 8.5: Glycogen Metabolism
Amylo-a(1,6)-glucosidase
Figure 8.19 Glycogen Degradation via Debranching Enzyme
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

78. Section 8.5: Glycogen Metabolism
Regulation of Glycogen Metabolism
Carefully regulated to maintain consistent energy levels
Regulation involves insulin, glucagon, epinephrine, and allosteric effectors
Figure 8.21 Major Factors Affecting Glycogen Metabolism
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

79. Section 8.5: Glycogen Metabolism
Glucagon activates glycogenolysis
Insulin inhibits glycogenolysis and activates glycogenesis
Epinephrine release activates glycogenolysis and inhibits glycogenesis
Figure 8.21 Major Factors Affecting Glycogen Metabolism
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

80. Biochemistry in Perspective
Saccharomyces cerevisiae and the Crabtree effect
S. cerevisiae is the only yeast that can produce ethanol and CO2 in such large quantities
S. cerevisiae ferments carbohydrates efficiently and dominates its environment due to the Crabtree effect
The Crabtree Effect
Unlike most fermenting organisms S. cerevisiae can also ferment sugar in the presence of O2
As glucose and/or fructose levels rise pyruvate is diverted away from the citric acid cycle into ethanol synthesis
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

81. Biochemistry in Perspective
The phenomenon, in which glucose represses aerobic metabolism, is the Crabtree effect
Rapid production of ethanol has the effect of eliminating microbial competitors
Once glucose levels are depleted and O2 is available the yeast reabsorbs the ethanol and converts it to acetaldehyde for use as an energy source
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press

82. Biochemistry in Perspective
Figure 8A Ethanol Metabolism in S. cerevisiae
From McKee and McKee, Biochemistry, 5th Edition, © 2011 Oxford University Press