GlycolysisBiochemistry, 4th edition, RH Garrett & CM Grisham, Brooks/Cole (Cengage); Boston, MA: 2010

pp 535-562

Instructor: Kirill Popov

1. General metabolism of glucose

2. The reactions of glycolysis

3. Metabolism of hexoses other than glucose

4. Control of metabolic flux

Glycogenolysis

Glycogen Triglyceride Protein

Acetyl-CoA

Amino acidsGlucose Free fatty acidsLipolysis Proteolysis

β-OxidationPyruvate Deamination and

oxidationOxidation

Glycolysis

Fate of major metabolic fuels

Glycogen

Glucose

Ribose 5-phosphate Pyruvate

oxidation viapentose phosphate

pathway

storage

oxidation viaglycolysis

Major metabolic pathways of glucose

Glucose

2 Pyruvate

2 Ethanol + 2 CO22 Lactate

2 Acetyl-CoA

4 CO2 + 4 H2O

glycolysis(10 successivereactions)

Fermentation tolactate in vigorouslycontracting muscle,erythrocytes, someother cells, and insome microorganisms

anaerobicsconditions

anaerobicsconditions

anaerobicsconditions

Fermentation to alcoholin yeast

citricacidcycle

Animal, plant, andmany microbial cellsunder aerobic conditions

2CO2

Catabolic fates of the pyruvate formed in glycolysis

D-Glucose 2 pyruvate 2 acetyl-CoAglycolysis PDH

2CO2 4CO22 L-lactate

No O2 requirementfor glycolysis

O2 requirement for pyruvatedehydrogenase (PDH) plus

TCA cycle activity

TCA

Glycolysis is a preparatory pathway for aerobic metabolism of glucose

Glucose

GlucosetransporterPlasma

membrane

[Glucose] mM

J glu

cose

(mM

îcm

îs-1

î106

)

Mechanism of glucose uptake

Major ways in which glucose is metabolized in different tissues

erythrocytes brain

muscle liver

Glucose

Glucose 6-phosphate

e-

Pentosephosphates

(2)Lactate-

(2) H+

Glucose

Glucose 6-phosphate

e-

Pentosephosphates

(2)Lactate-

(2) H+

(2) CO2

2 Acetyl-CoA(2) CO2

TCA

Glucose

Glucose 6-P

e-

Pentosephosphates

(2) H+

(2) CO2

2 Acetyl-CoA(2) CO2

TCA

(2)Lactate-(2)Pyruvate-

Glycogen?

Glucose

Glucose

e-

Pentosephosphates

(2) H+

(2) CO2

2 Acetyl-CoA(2) CO2

TCA

(2)Lactate-(2)Pyruvate-

Fat

Glucurinides

Glucose 6-P Glycogen

O CH2 C CH2 OH

O

P

O

CH2

OH

OH

OHOH

HO

C CO

O-O

CH3

second ATP-forming reaction(substrate-levelphosphorylation)

Glucose

Glucose 6-phosphate

Fructose 6-phosphate

Fructose 1,6-bisphosphate

Glyceraldehyde 3-phosphate

Dihydroxyacetone phosphate

Glyceraldehyde 3-phosphate (2)

1,3-Bisphosphoglycerate (2)

3-Phosphoglycerate (2)

2-Phosphoglycerate (2)

Phosphoenol pyruvate (2)

Pyruvate (2)

1

2

3

4

5

6

7

8

9

10

ATP

2 ATP

ADP

ATP

2ADP

2ADP

2 ATP

ADP

2 NADH + H+

2Pi2NAD+

firstpriming reaction

secondpriming reaction

cleavageof 6-carbonsugarphosphate totwo 3-carbonsugarphosphates

oxidation andphosphorylation

first ATP-forming reaction(substrate-levelphosphorylation)

Preparatory phasePhosphorylation of glucoseand its conversion toglyceraldehyde 3-phosphate

Payoff phaseOxidative conversion ofglyceraldehyde 3-phosphate topyruvate and the coupledformation of ATP and NADH

1

2

3

4

5

Hexokinase

Phosphohexoseisomerase

Phospho-fructokinase-1Aldolase

Triosephosphateisomerase

6

7

8

9

10

Glyceraldehyde 3-phosphatedehydrogenase

Phosphoglyceratekinase

Phosphoglyceratemutase

EnolasePyruvatekinase

O CH2 CH CO

HOH

P

O CH2 CH CO

HOH

P

CH2 C CO

O-O

P

OCH2

OH

OH

CH2 OHOP

OCH2

OH

OH

CH2 OO PP

O

CH2

OH

OH

OHOH

OP

O CH2 CH CO

O-OH

P

O CH2 CH CO

OOH P

P

CH2 CH CO

O-OH O

P

O

CH2

OH

OH

OHOH

HO

O

CH2

OH

OH

OHOH

OP-O

O-

O

hexokinase

ΔG'º = -16.7 kJ/mol

ATP ADP

Glucose Glucose 6-phosphate

Mg2+

1

23

4

5

6

Phosphorylation of glucose

O

CH2

OH

OH

OHOH

OP-O

O-

O

Glucose 6-phosphate

1

23

4

5

6

phosphohexoseisomerase

Mg2+

OP-O

O-

O

OCH2

OH

OH

CH2 OH

Fructose 6-phosphate

ΔG'º = 1.7 kJ/mol

1

2

34

5

6

Conversion of glucose 6-phosphate to fructose 6-phosphate

OP-O

O-

O

OCH2

OH

OH

CH2 O P

O

O-

O-

OP-O

O-

O

OCH2

OH

OH

CH2 OH

Fructose 6-phosphate

ΔG'º = -14.2 kJ/mol

1

2

34

5

6ATP ADP

Mg2+

phosphofructokinase-1

Fructose 1,6-bisphosphate

1

2

34

5

6

Phosphorylation of fructose 6-phosphate to fructose 1,6-bisphosphate

OP-O

O-

O

OCH2

OH

OH

CH2 O P

O

O-

O-

ΔG'º = 23.8 kJ/mol

Fructose 1,6-bisphosphate

1

2

34

5

6

aldolase

Glyceraldehyde3-phosphate

Dihydroxyacetonephosphate

CH2

C

CH2OH

O

O

P O-

O-

O

+

C

COH

CH2 O

HO

P O-

O-

OH

Cleavage of fructose 1,6-bisphosphate

ΔG'º = 7.5 kJ/mol

triose phosphateisomerase

Glyceraldehyde3-phosphate

Dihydroxyacetonephosphate

CH2OH

C

CH2 O P O-

O-

OO H

C

COH

CH2 O

HO

P O-

O-

O

Interconversion of the triose phosphates

HO P O-

O-

O P O-

O-

O

C

COH

CH2OPO32-

OO

ΔG'º = 6.3 kJ/mol

glyceraldehyde3-phosphate

dehydrogenase

Glyceraldehyde3-phosphate

Inorganicphosphate 1,3-Bisphosphoglycerate

NAD+ NADH + H+C

COH

CH2OPO32-

HO

H

Oxidation of glyceraldehyde 3-phosphate to 1,3 bisphosphoglycerate

C

C

CH2OPO32-

H

H

OH

O

C

C

CH2OPO32-

H

H

OH

OH

S

C

C

CH2OPO32-

H OH

S

O C

C

CH2OPO32-

H OH

S

O

C

C

CH2OPO32-

H OH

O

P-O

O

O

O-

OHP-O

O-

O

Glyceraldehyde 3-phosphatedehydrogenase

NAD+ NADH + H+

NADHNAD+ NAD+

SH

Thiohemiacetal Thioester

H+

Glyceraldehyde3-phosphate

1,3-Bisphospho-glycerate

Cys Cys Cys

Cys

NAD+

1 2 3 4

The glyceraldehyde 3-phosphate dehydrogenase reaction

ΔG'º = -18.5 kJ/mol

1,3-Bisphosphoglycerate

P O-

O-

O

C

COH

CH2OPO32-

OO C

COH

CH2OPO32-

O-O-O P O

O-

ADP ATP

phosphoglyceratekinase

3-Phosphoglycerate

AdenineRib

P

P

O

AdenineRib

P

P

O

+ +Mg2+

Phosphoryl transfer from 1,3-bisphosphoglycerate to ADP

ΔG'º = 4.4 kJ/mol

3-Phosphoglycerate

phosphoglyceratemutase

Mg2+C

C

CH2

O-O

O P O-

O-

OOHH

2-Phosphoglycerate

O

H O P O-

O-

OC

C

CH2

O-

OH

Conversion of 3-phosphoglycerate to 2-phosphoglycerate

3-Phosphoglycerate 2-Phosphoglyceratephosphoglyceratemutase

2,3-Bisphos-phoglycerate

Phosphoenzyme

3-Phosphoglycerate

3-Phosphoglycerate

2,3-Bisphosphoglycerateinitial phosphorylation

of enzyme

ADP ATP

Enzyme withunphosphorylatedHis residue

-O P O

O-

1 2

kinase

His

His

His

Mechanism of the phosphoglycerate mutase reaction

O P O-

O-

OC

C

CH2

O-O

HO

H O P O-

O-

OC

C

CH2

O-O

ΔG'º = 7.5 kJ/mol

enolase

Phosphoenolpyruvate2-Phosphoglycerate

H2O

Dehydration of 2-phosphoglycerate to phosphoenolpyruvate

O P O-

O-

OC

C

CH2

O-O

O

C

C

CH3

O-O-O P O

O-

ΔG'º = -31.4 kJ/mol

ADP

ATP

Pyruvate

Mg2+, K+

Phosphoenolpyruvate

pyruvatekinase

AdenineRib

P

P

OAdenineRib

P

P

O+ +

Transfer of the phosphoryl group from phosphoenolpyruvate to ADP

Glucose

2 Pyruvate 2 Lactate2 NADH

2 NAD+

Pyruvate is the terminal electron acceptor in lactic acid fermentation

O

C

C

CH3

O-O

H

C

C

CH3

O-O

HO

Pyruvate

Lactate

lactatedehydrogenase

ΔG'º = -25.1 kJ/mol

NADH + H+

NAD+

Lactate dehydrogenase reaction

O

CH2OH

OH

OH

OHOH

O

CH2OH

OH

OH

OH

OH

O

CH2OH

OH OHOHOH

OCH2OH

OH

OH

CH2OH

fructose 1-phosphatealdolase

LactoseTrehalose

Sucrose

Glycogen; starch

Glucose1-phosphate

Glucose6-phosphate

Fructose6-phosphate

Fructose 1,6-bisphosphate

Glyceraldehyde3-phosphate

Glyceraldehyde Dihydroxyacetonephosphate

D-Fructose

Mannose 6-phosphate

D-Mannose

D-Galactose

D-Glucose

Pi

ATP

ATP

ATP

UDP-glucose

UDP-galactose

Fructose 1-phosphate

ATP

hexokinase

hexokinase

sucrase

trehalase lactase

hexokinaseATP

phosphorylase

phosphogluco-mutase

fructokinasePhosphomannose

isomerase

triosekinase

triose phosphateisomerase

+

Entry of dietary hexoses into the preparatory stage of glycolysis

½ Glucose

1,3-Bisphosphoglycerate

3-Phosphoglycerate

2-Phosphoglycerate

Lactate

2,3-BisphosphoglycerateATP

ADP

The 2,3bisphosphoglycerate shunt

Glucose

Glucosetransporter

EndocytosisExocytosis

Stimulationby insulin

Membranousvesicle

Plasma membrane

Insulin stimulates glucose uptake by adipose tissue and muscle

Glucose concentration (mM)

Rel

ativ

e en

zym

e ac

tivity

20

1.0

151050

Kinetic properties of glucokinase and hexokinase

Capillary

Glucose

GLUT2

Glucose

Fructose 6-phosphate

Glucose 6-phosphate

Cytosol Nucleus

Hexokinase IV

Regulatorprotein

Plasmamembrain

Hexokinase IV

Regulation of glucokinase by sequestration in the nucleus

Fructose 6- + ATPphosphate

Fructose 1,6- + ADPphosphate

ATP AMP, ADP

citrate fructose 2,6-bisphosphate

Regulation of phosphofructokinase-1

Relative Changes in [ATP] and [AMP] When ATP Is Consumed

Adenine nucleotide

Concentration before ATP

depletion (mM)

Concentration after ATP depletion (mM)

Relative change

ATP 5.0 4.5 10%ADP 1.0 1.0 0AMP 0.1 0.6 600%

Fructose 6-phosphate

PFK

-1 a

ctiv

ityLow [ATP]

High [ATP]

Regulation of phosphofructokinase-1

O

O

H

OH

OH

O

H

C H2 POP O -

O -

-O

O -

OO

HCH 2OH1

2

34

5

6

Fructose 2,6-bisphosphate

[Fructose 6-phosphate] (mM)

PFK

-1 a

ctiv

ity (%

of V

max

)

-F2,6BP

+F2,6BP

100

80

60

40

20

00 0.05 0.1 0.2 0.4 0.7 1.0 2.0 4.0

Role of fructose 2,6-bisphosphate in regulation of PFK-1

All glycolytic tissues, including liverLiver only

transamination

glucagon

ATPADP

Pyruvatekinase L(inactive)

Pyruvatekinase

L/M

PEP

F16BP

ADP

ATP

Pyruvate

Alanine

ATP,acetyl-CoA,long-chainfatty acids

6 steps

PKA

PP

PiH2O

P

Regulation of pyruvate kinase

Glucose

(2) Pi

Glucose 6-phosphate

Fructose 6-phosphate

Fructose 1,6-bisphosphate

(2) Glyceraldehyde 3-phosphate

(2) 1,3-Bisphosphoglycerate

(2) Phosphoenolpyruvate

(2) Pyruvate

(2) Lactate

Glycogensynthesis

Pentosephosphate pathway

ATP

ATP

(2) ADP(2) ATP

ADP

ADP

AMP, Fructose 2,6-P2ATP, citrate, H+

ATP, alanine

(2) NAD+

(2) NADH + 2H+

(2) NADH + 2H+

(2) NAD+

++

++

−

Important regulatory features of glycolytic pathway

1. Glycolysis is a near universal pathway by which a glucose molecule is oxidized to two molecules of pyruvate, with energy conserved as ATP and NADH

2. All ten glycolytic enzymes are in cytosol, and all the intermediates are phosphorylated compounds of three or six carbons

3. In the preparatory phase of glycolysis, ATP is invested to convert glucose to fructose 1,6-bisphosphate; fructose 1,6-bisphosphate is than broken to yield two molecules of triose phosphate

4. In the payoff phase, each of the two molecules of glyceraldehyde 3-phosphate undergoes oxidation; the energy of this oxidation is conserved in the formation of NADH and ATP

5. Glycolysis is tightly regulated in coordination with other energy-yielding pathways