You are on page 1of 11

10/28/10

Glycolysis 1

Lecture 23
Chapter 15
Sections 1 and 2

Chapter 14 Opener

History of Glycolysis
•  1850’s: Pasteur showed yeast can ferment sugar to
CO2 and ethanol, but he cannot replicate with
extracts, proposes “vitalism”.
•  1898: Hans and Eduard Buchner demonstrate yeast
extracts ferment sugar to CO2 and ethanol, proving
vitalism false. Birth of biochemistry.
•  1905: Harden and Young dialyze yeast extract into
two fractions:
–  Retained (high molecular weight), called “zymase”.
–  Dialyzed (low molecular weight), called “co-zymase”.
–  Need to mix together to recover ability to ferment sugar.
–  Lose activity if boil zymase, but not if boil co-zymase. Zymase
Co-zymase
Figure 2-14

1
10/28/10

What were Zymase and Co- NAD+: Nicotinamide Adenine


zymase? Dinucleotide
•  Zymase: high molecular weight, heat- •  Mobile 2 e- carrier.
labile enzymes. •  Carries 2e- in the form of hydride ion:
H+ + 2e- => H-
•  Co-zymase: low molecular weight, •  Usually involved in oxidation of primary
heat-stable factors: alcohol to carbonyl:
–  Inorganic cofactors: Pi, Mg++ R-CH2-OH + NAD+ => R-CH=O + NADH + H+
–  Organic coenzymes: ADP, NAD+ •  Derived from niacin (vitamin b3)

Overview of Glycolysis
•  Glycolysis means “sweet splitting”
–  Glyco: Greek for sweet
–  Lysis: Greek for splitting or loosening
•  Split a 6-carbon molecule (Glucose) to two 3-carbon
molecules (pyruvate).
•  Largest carbon flux in most cells.
•  Best understood metabolic pathway.
•  Central to many other pathways.
•  Also called fermentation:
–  Yeast: Glucose -> 2 CO2 + 2 Ethanol
–  Exercising muscle, RBCs, bacteria: Glucose -> 2 Lactic acid

Figure 11-4

2
10/28/10

Glycolysis Summary
Preparatory
Stage
•  10 step breakdown of one glucose (C6) to
two pyruvates (C3).
•  Balanced equation:
Glc + 2NAD+ + 2ADP + 2Pi --->
2Pyr + 2NADH + 2ATP + 2H2O + 2H+ Payoff
•  Two stages: Stage
–  Preparatory stage (reactions 1-5): invest 2 ATP
–  Payoff stage (reactions 6-10): produce 4 ATP

Figure 15-1

Reaction 1:
Recall that hexokinase catalyzes a coupled
Hexokinase reaction:

First priming reaction Coupling the energetically unfavorable phosphorylation of
glucose to the energetically favorable hydrolysis of ATP.

Kinases: Enzymes ΔGo’

that transfer Pi from
ATP to a substrate

Hexokinase also
phosphorylates
mannose and fructose.

Page 489

3
10/28/10

Glucose binding causes a large conformational shift,


Kinases bind ATP complexed with Mg++ excluding water and bringing the 6’-OH close to the ATP

Mg++ shields negative charges, allowing Catalysis by


nucleophilic attack of 6’-OH. proximity

Page 489 Figure 15-2a

Glucose binding causes a large conformational shift, Reaction 2:


excluding water and bringing the 6’-OH close to the ATP
Phoshoglucose
Catalysis by isomerase
proximity

Catalyzes reversible
isomerization using
general acid-base
catalysis

Figure 15-2b Page 490

4
10/28/10

Reaction 3:
PGI Mechanism
Phosphofructokinase

Committed step, so
key regulatory site.

Second priming
reaction

Coupled reaction, like


that catalyzed by
hexokinase.
Irreversible in cells

Bisphosphate: 2
phosphates not linked
to one another.
Figure 15-3 Page 491

Reaction 4:
Why is phosphofructokinase the committed step in
glycolysis? Aldolase

Glucose Reversible aldol


condensation

Glucose-6P Cleavage reaction

Note change in
Glycogen Pentose-P numbering!!
Fructose-6P

Fructose-1,6-BP

Page 492

5
10/28/10

Aldolase mechanism:
Non-enzymatic, base catalyzed aldol cleavage:

Stabilization of C-
by resonance

Ketone

Aldehyde

Note position of carbonyl relative to bond cleaved.


Provides rationale for the phosphoglucose isomerase
reaction.
Figure 15-4 Figure 15-5

Schiff base = imine.


Protonated Schiff
base serves as an
electron sink,
pulling electrons
towards itself to
initiate cleavage
reaction.

Figure 15-5 part 1 Figure 15-5 part 2

6
10/28/10

Resonance stabilized

Figure 15-5 part 3 Figure 15-5 part 4

Reaction 5:
Triose phosphate isomerase (TIM)
Enediol intermediate, like PGI

Figure 15-5 part 5 Part 494

7
10/28/10

Compounds that resemble the transition state (enediol)


PGI Mechanism bind to the enzyme more tightly than either substrate

Figure 15-3 Part 494

TIM has a flexible loop that closes over the active site, Toxic compound released upon dephosphorylation
preventing loss of the phosphate from the endiol intermediate of enediol intermediate
TIM barrel structure:

α/ß barrel, with a


cylinder of 8 parallel
ß-strands
surrounded by 8 α-
helices.

Figure 15-6 Part 495

8
10/28/10

Fate of C-atoms after Aldolase and Triose- Review of Preparatory


phosphase isomerase reactions
Stage of Glycolysis

Figure 15-7

Reaction 6:

GAP dehydrogenase
Preparatory
Dehydrogenases:
Stage
involved in
oxididation/reduction
reactions.

Couples favorable
oxidation to
Payoff unfavorable
Stage phosphorylation
through covalent
intermediate:
thioester

Figure 15-1 Page 497

9
10/28/10

GAP
dehydrogenase
mechanism δ-
Charge separation

δ+

Figure 15-9 Figure 15-9 part 1

Easier to extract H- from thiohemiacetal


than carbohyl due to dissipation of
charge separation. Energetically
favorable oxidation

δ-
Charge separation
δ+ Energy from
oxidation
conserved in
thioester

Figure 15-9 part 2 Figure 15-9 part 3

10
10/28/10

Phosphorolysis:

Conserves energy of
thioester (originally from
oxidation) in acyl-phosphate

(Acyl-phosphate)

Figure 15-9 part 4 Figure 15-9 part 5

GAP Experiments supporting GAPDH mechanism


dehydrogenase
mechanism (inactive)

GAPDH catalyzes Pi
exchange

Figure 15-9 Figure 15-8

11

You might also like