How do you convert glucose to two molecules of pyruvate?

Reactions needed:
Carbon-carbon bond cleavage
Reduction/oxidation
Dehydration
Cleavage via retro aldol most likely, but glucose yields a C4 and C2 product
The Aldol Reaction Enolate formation

B:
Chemical strategy of glycolysis
1. Add phosphoryl groups to glucose

2. Isomerize glucose (aldose sugar) to fructose (ketose sugar) so retro

aldol will yield two C3 units

3. Chemically convert phosphorylated intermediates into compounds

with high phosphate group-transfer potentials

4. Chemically couple the subsequent hydrolysis of reactive substances to

ATP synthesis
(adapted from Voet & Voet)
Metabolic Oxidation of Glucose
Glycolysis
Net Reaction:

Glucose + 2 ADP + 2 NAD+ + 2 Pi

2 pyruvate + 2 ATP + 2 NADH + 2 H+ + 2 H2O

Preparative
stage

Payback
stage
Hexokinase (glucokinase) Structure of Hexokinase (1GLK)

ΔG°’ = -16.7 kJ/mol

Hexokinase is the prototypical example of an enzyme that uses

induced fit for substrate recognition

Random Bi Bi mechanism

Major control point for glycolysis

Regulated by Glc-6-P (product inhibition)

ATP = -1
NADH = 0
Phosphorylation of glucose ensures that it cannot diffuse out of cell
Phosphoglucose isomerase Structure of PGI (2PGI)

Reaction is an aldose --> ketose conversion ΔG°’ = 1.7 kJ/mol

Highly stereoselective

Occurs under near-equilibrium conditions

ATP = -1
NADH = 0
Reaction pathway goes through linear form
PGI mechanism

Enzyme deprotonates C2 of linear Stereospecific protonation

G6P forming enediolate intermediate of C1 leads to isomerization
Phosphofructokinase (PFK-1) Structure of PFK (1CFY)

ΔG°’ = -14.2 kJ/mol

Primary control point for glycolysis

overall rate determining step for glycolysis

ATP is a substrate, but it is also an allosteric inhibitor

other effector molecules include AMP and citrate

ATP = -2
NADH = 0
 Structure of aldolase (1ADO)
Aldolase
 Structure of aldolase (1ADO)
Aldolase

ΔG°’ = +23.9 kJ/mol (GAP) (DHAP)

This bond
will be broken

Where is the bond to be broken :B

in the open form?

Reaction is base catalyzed. What group

must be deprotonated? ATP = -2
NADH = 0

Reaction is very unfavorable thermodynamically, but is driven by PFK-1 and

glyceraldehyde-3-phosphate dehydrogenase
In the payback phase, C6 and C1 from original glucose are
equivalent
Triose Phosphate Isomerase Structure of TPI (1TIM)

(GAP)
(DHAP)
ΔG°’ = +7.6 kJ/mol

Isomerization is an aldose <----> ketose shift like that of G6P --> F6P

Interconversion between DHAP and GAP is extremely rapid

as GAP consumed in subsequent steps of glycolysis, DHAP is
converted to GAP

Proposed mechanism involves TS‡ stabilized by low-barrier hydrogen bonds

ATP = -2
NADH = 0
Triose phosphate isomerase mechanism
 Structure of GAPDH (1b7g)
Glyceraldehyde 3-phosphate dehydrogenase

Since there are 2 GAP produced per molecule of glucose - all reactions of the
triose stage occur in duplicate!!

Reaction occurs under near-equilibrium conditions

Produces both NADH and a high energy phosphate group

Reaction proceeds through a thioacyl intermediate to drive phosphorylation

ATP = -2
NADH = 2
Vitamine B3 (Niacin)
Nicotinamide
(NAD+, NADP+, NADH, NADPH)

Involved in 2e- redox reactions

H+, 2e-
Mechanism of NAD+ dependent oxidation of lactic acid

Overall reaction:

Enzyme aligns substrate and

cofactor for transfer of hydride
equivalent (H+ and 2 e-)

Often activated by deprotonation

(as shown for oxidation of lactate
to pyruvate) or nucleophilic attack.
Mechanism of GAPDH
Phosphoglycerate kinase Structure of PGK (13PK)

ΔG°’ = -18.8 kJ/mol

Example of substrate level phosphorylation

BPG is a high-energy phosphate compound (ΔG°’ = -49.4 kJ/mol)
can donate a phosphate to ADP

This reaction drives the slightly unfavorable oxidation of GAP to BPG

ΔG°’ = -18.8 + 6.7 = -12.1 kJ/mol

Occurs under near-equilibrium conditions ATP = 0

NADH = 2
 Structure of PGM (1BQ3)
Phosphoglycerate mutase

ΔG°’ = +4.4 kJ/mol

H H

Occurs under near-equilibrium conditions

Phosophohistidine in active site
ATP = 0
Bisphosphoglycerate made transiently NADH = 2
 Structure of enolase (1EBH)
Enolase

+ H2O

H ΔG°’ = +1.7 kJ/mol

B: Phosphoenolpyruvate

-C

B-H+
Mg2+

Occurs under near-equilibrium conditions

Dehydration reaction ATP = 0
NADH = 2
PEP is the highest energy phosphate compound in biochemistry
ΔG°’ = -62 kJ/mol
 Structure of PK (1A3W)
Pyruvate Kinase

ADP + H+ ATP

ΔG°’ = -31.4 kJ/mol

Mg2+
K+

H+

Metabolically irreversible

Major control point for glycolysis ATP = 2

NADH = 2
Fate of pyruvate

NADH must be reoxidized to NAD+ to allow glycolysis to continue.

Fate of pyruvate

YEAST MAMMALS
NADH + H+
In muscle
Fermentation: Lactate + NAD+
Pyruvate H+
decarboxylase
Lactate transported
CO2 + acetaldehyde to liver via blood
NADH + H+ NAD+
Alcohol
dehydrogenase

Ethanol and NAD+ Pyruvate + NADH + H+ In liver

NADH must be reoxidized to NAD+ to allow glycolysis to continue.