Glycolysis

Glycolysis takes place in the cytosol of cells.

Glucose enters the Glycolysis pathway by conversion to glucose-6-phosphate.

Initially there is energy input corresponding to cleavage of two ~P bonds of ATP.

H O

OH

H

OHH

OH

CH2OPO32

H

OH

H

1

6

5

4

3 2

glucose-6-phosphate

H O

O H

H

O HH

O H

CH 2O H

H

O H

H H O

O H

H

O HH

O H

CH 2O PO 32

H

O H

H

23

4

5

6

1 1

6

5

4

3 2

A T P A D P

M g 2+

glucose g lucose -6 -phosphate

H ex ok inase

1. Hexokinase catalyzes:

Glucose + ATP glucose-6-P + ADP

The reaction involves nucleophilic attack of the C6 hydroxyl O of glucose on P of the terminal phosphate of ATP.

ATP binds to the enzyme as a complex with Mg++.

2. Phosphoglucose Isomerase catalyzes:

glucose-6-P (aldose) fructose-6-P (ketose)

The mechanism involves acid/base catalysis, with ring opening, isomerization via an intermediate, and then ring closure.

H O

O H

H

O HH

O H

CH 2 O PO 32

H

O H

H

1

6

5

4

3 2

CH 2 O PO 32

O H

CH 2 O H

H

O H H

H HO

O6

5

4 3

2

1

g lu c o s e - 6 - p h o s p h a te f r u c to s e - 6 - p h o s p h a te P h o s p h o g lu c o s e Is o m e r a s e

3. Phosphofructokinase catalyzes: fructose-6-P + ATP fructose-1,6-bisP + ADP

This highly spontaneous reaction has a mechanism similar to that of Hexokinase.

The Phosphofructo kinase reaction is the rate-limiting step of Glycolysis.

The enzyme is highly regulated.

CH 2 O PO 32

O H

CH 2 O H

H

O H H

H HO

O6

5

4 3

2

1 CH 2 O PO 32

O H

CH 2 O PO 32

H

O H H

H HO

O6

5

4 3

2

1

A T P A D P

M g 2 +

f r u c t o s e - 6 - p h o s p h a t e f r u c t o s e - 1 , 6 - b i s p h o s p h a t e

P h o s p h o f r u c t o k i n a s e

4. Aldolase catalyzes: fructose-1,6-bisphosphate dihydroxy acetone-P + glyceraldehyde-3-P

The reaction is an aldol cleavage, the reverse of an aldol condensation.

6

5

4

3

2

1 CH 2 O PO 32

C

C

C

C

CH 2 O PO 32

O

HO H

H O H

H O H

3

2

1

CH 2 O PO 32

C

CH 2 O H

O

C

C

CH 2 O PO 32

H O

H O H+

1

2

3

f ru c to s e -1 ,6 - b is p h o s p h a te

A ld o la s e

d ih y d ro x y a c e to n e g ly c e ra ld e h y d e -3 - p h o s p h a te p h o s p h a te

T rio s e p h o s p h a te Is o m e ra s e

5. Triose Phosphate isomerase catalyzes:

dihydroxy acetone-P glyceraldehyde-3-P

Glycolysis continues from glyceraldehyde-3-P. Keq favors dihydroxyacetone-P.

6

5

4

3

2

1 CH 2 O PO 32

C

C

C

C

CH 2 O PO 32

O

HO H

H O H

H O H

3

2

1

CH 2 O PO 32

C

CH 2 O H

O

C

C

CH 2 O PO 32

H O

H O H+

1

2

3

f ru c to s e -1 ,6 - b is p h o s p h a te

A ld o la s e

d ih y d ro x y a c e to n e g ly c e ra ld e h y d e -3 - p h o s p h a te p h o s p h a te

T rio s e p h o s p h a te Is o m e ra s e

C

C

CH 2 O PO 32

H O

H O H

C

C

CH 2 O PO 32

O O PO 32

H O H+ P i

+ H +

N A D + N A D H 1

2

3

2

3

1

g l y c e r a l d e h y d e - 1 , 3 - b i s p h o s p h o - 3 - p h o s p h a t e g l y c e r a t e

G l y c e r a l d e h y d e - 3 - p h o s p h a t e D e h y d r o g e n a s e

6. Glyceraldehyde-3-phosphate Dehydrogenase catalyzes:

glyceraldehyde-3-P + NAD+ + Pi

1,3-bisphosphoglycerate + NADH + H+

C

C

CH 2 O PO 32

H O

H O H

C

C

CH 2 O PO 32

O O PO 32

H O H+ P i

+ H +

N A D + N A D H 1

2

3

2

3

1

g l y c e r a l d e h y d e - 1 , 3 - b i s p h o s p h o - 3 - p h o s p h a t e g l y c e r a t e

G l y c e r a l d e h y d e - 3 - p h o s p h a t e D e h y d r o g e n a s e

Exergonic oxidation of the aldehyde in glyceraldehyde- 3-phosphate, to a carboxylic acid, drives formation of an acyl phosphate, a "high energy" bond (~P).

This is the only step in Glycolysis in which NAD+ is reduced to NADH.

C

C

CH 2 O PO 32

O O PO 32

H O H

C

C

CH 2 O PO 32

O O

H O H

A D P A T P

1

22

3 3

1

M g 2+

1 , 3 - b i s p h o s p h o - 3 - p h o s p h o g l y c e r a t e g l y c e r a t e

P h o s p h o g l y c e r a t e K i n a s e

7. Phosphoglycerate Kinase catalyzes: 1,3-bisphosphoglycerate + ADP 3-phosphoglycerate + ATP

This phosphate transfer is reversible (low G), since one ~P bond is cleaved & another synthesized.

The enzyme undergoes substrate-induced conformational change similar to that of Hexokinase.

C

C

CH 2 O H

O O

H O PO 32

2

3

1C

C

CH 2 O PO 32

O O

H O H2

3

1

3 - p h o s p h o g l y c e r a t e 2 - p h o s p h o g l y c e r a t e

P h o s p h o g l y c e r a t e M u t a s e

8. Phosphoglycerate Mutase catalyzes: 3-phosphoglycerate 2-phosphoglycerate

Phosphate is shifted from the OH on C3 to the OH on C2.

9. Enolase catalyzes:

2-phosphoglycerate phosphoenolpyruvate + H2O

This dehydration reaction is Mg++-dependent.

2 Mg++ ions interact with oxygen atoms of the substrate carboxyl group at the active site.

The Mg++ ions help to stabilize the enolate anion intermediate that forms when a Lys extracts H+ from C #2.

C

C

C H 2 O H

O O

H O P O 32

C

C

C H 2 O H

O O

O P O 32

C

C

C H 2

O O

O P O 32

O H

2

3

1

2

3

1

H

2 -p h o s p h o g ly c e r a t e e n o la t e in t e r m e d ia te p h o s p h o e n o lp y r u v a te

E n o la s e

10. Pyruvate Kinase catalyzes:

phosphoenolpyruvate + ADP pyruvate + ATP

C

C

CH 3

O O

O2

3

1A D P A T PC

C

CH 2

O O

O PO 32

2

3

1

p h o s p h o e n o l p y r u v a t e p y r u v a t e

P y r u v a t e K i n a s e

Hexokinase

Phosphofructokinase

glucose Glycolysis

ATP

ADP glucose-6-phosphate

Phosphoglucose Isomerase

fructose-6-phosphate

ATP

ADP fructose-1,6-bisphosphate

Aldolase

glyceraldehyde-3-phosphate + dihydroxyacetone-phosphate

Triosephosphate Isomerase Glycolysis continued

Glyceraldehyde-3-phosphate Dehydrogenase

Phosphoglycerate Kinase

Enolase

Pyruvate Kinase

glyceraldehyde-3-phosphate

NAD+ + Pi

NADH + H+

1,3-bisphosphoglycerate

ADP

ATP

3-phosphoglycerate

Phosphoglycerate Mutase

2-phosphoglycerate H2O

phosphoenolpyruvate

ADP

ATP pyruvate

Glycolysis continued.

Recall that there are 2 GAP per glucose.

Glycolysis

Balance sheet for ~P bonds of ATP:

How many ATP ~P bonds expended? __2______

How many ~P bonds of ATP produced? (Remember there are two 3C fragments from glucose.) ___4_____

Net production of ~P bonds of ATP per glucose: ______2__

Balance sheet for ~P bonds of ATP: 2 ATP expended 4 ATP produced (2 from each of two 3C fragments

from glucose) Net production of 2 ~P bonds of ATP per glucose.

Glycolysis - total pathway, omitting H+: glucose + 2 NAD+ + 2 ADP + 2 Pi 2 pyruvate + 2 NADH + 2 ATP

In aerobic organisms: pyruvate produced in Glycolysis is oxidized to CO2

via Krebs Cycle NADH produced in Glycolysis & Krebs Cycle is

reoxidized via the respiratory chain, with production of much additional ATP. 

THANK YOU

Queries welcome