Carbohydrate Metabolism

A closer Look at Glycolysis

Metabolism
The complex of physical and chemical processes occurring within a living cell or organism that are necessary for the maintenance of life. In metabolism some substances are broken down to yield energy for vital processes while other substances, necessary for life, are synthesized.

Overview to carbohydrate Metabolism

The major source of dietary carbohydrate for humans is starch from consumed plant material. This is supplemented with a small amount of glycogen from animal tissue, disaccharides such as sucrose from products containing refined sugar and lactose in milk.

Glucose in the body undergoes one of Three metabolic fates

Glucose is catabolised to produce ATP
Catabolism occurs in all peripheral tissues, particularly in brain, muscle and kidney.

Glucose is stored as glycogen

This storage occurs in liver and muscle.

Glucose is converted to fatty acids.

Once converted to fatty acids, these are stored in adipose tissue as triglycerides.

Glucose catabolism
Glucose will be oxidized by all tissues to synthesis ATP. The first pathway which begins the complete oxidation of glucose is called Glycolysis. This pathway cleaves the six carbon glucose molecule (C6H12O6) into two molecules of the three carbon compound Pyruvate (C3H3O3-). This oxidation is coupled to the net production of two molecules of ATP/glucose.

Carbohydrates
Glucos e

Cytosol - anaerobic
Hexokinase Pentose Phosphate Shunt

Glucose-6-P
glycolysis

Glc-1- phosphate

Glycogen

Pyruv ate

Cytosol

Pyruvate Acetyl CoA

Mitochondria Aerobic)

FATTY ACIDS

AMINO ACIDS

Krebs cycle

Reducing equivalents

Oxidative Phosphorylation (ATP)

Glycolysis
Catabolism of Glucose

Glycolysis: General Functions

Provide ATP energy Generate intermediates pathways for other

Hexose monophosphate pathway Glycogen synthesis Pyruvate dehydrogenase

Fatty acid synthesis Krebs Cycle

Glycerol-phosphate (TG synthesis)

Glycolysis: Specific Tissue Functions

RBCs
Rely exclusively for energy

Skeletal muscle
Source of energy during exercise, particularly high intensity exercise

Adipose tissue
Source of glycerol-P for TG synthesis Source of acetyl-CoA for FA synthesis

Liver
Source of acetyl-CoA for FA synthesis Source of glycerol-P for TG synthesis

glucose ATP ADP glucose-6-phosphate Hexokinase

Glycolysis

Phosphoglucose Isomerase fructose-6-phosphate ATP Phosphofructokinase ADP fructose-1,6-bisphosphate Aldolase glyceraldehyde-3-phosphate + dihydroxyacetone-phosphate Triosephosphate Isomerase Glycolysis continued

glyceraldehyde-3-phosphate NAD+ + Pi Glyceraldehyde-3-phosphate Dehydrogenase NADH + H+ 1,3-bisphosphoglycerate ADP Phosphoglycerate Kinase ATP 3-phosphoglycerate Phosphoglycerate Mutase 2-phosphoglycerate Enolase H2O phosphoenolpyruvate ADP Pyruvate Kinase ATP pyruvate

H
4

6 CH OPO 2 2 3 5 O

H OH
3

H
1

H
2

OH

OH

OH

glucose-6-phosphate

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

6 CH2OH

H
4

O H
2

H OH
3

ATP ADP H H
1

6 CH OPO 2 2 3 5 4

O H
2

OH

OH

Mg2+

H OH
3

H
1

OH

OH

OH

Hexokinase

OH

glucose

glucose-6-phosphate

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++.

Properties of Glucokinase and Hexokinase

6 CH2OH

H
4

O H
2

H OH
3

ATP ADP H H
1

6 CH OPO 2 2 3 5 4

O H
2

OH

OH

Mg2+

H OH
3

H
1

OH

OH

OH

Hexokinase

OH

glucose

glucose-6-phosphate

The reaction catalyzed by Hexokinase is highly spontaneous. A phosphoanhydride bond of ATP (~P) is cleaved. The phosphate ester formed in glucose-6phosphate has a lower G of hydrolysis.

H
4

6 CH OPO 2 2 3 5 O

H OH
3

H
1

6 CH OPO 2 2 3

1CH2OH

H
2

H
4

HO H

OH

OH

3 OH

OH

OH

Phosphoglucose Isomerase glucose-6-phosphate fructose-6-phosphate

2. Phosphoglucose Isomerase catalyzes: glucose-6-P (aldose) fructose-6-P (ketose) The mechanism involves acid/base catalysis, with ring opening, isomerization via an enediolate intermediate, and then ring

Phosphofructokinase
6 CH OPO 2 2 3

1CH2OH

ATP ADP Mg2+

6 CH OPO 2 2 3

1 CH2OPO32

H
4

HO H

H
4

HO H

3 OH

3 OH

OH

OH

fructose-6-phosphate

fructose-1,6-bisphosphate

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 Phosphofructokinase reaction is the rate-limiting step of Glycolysis.

1CH2OPO3 2C

O H OH OH

HO 3C H 4C H
5

Aldolase

CH2OPO32 3
2C

H
1C

1CH2OH

H 2C OH 2 3 CH2OPO3

CH2OPO32 6

fructose-1,6bisphosphate

dihydroxyacetone phosphate

glyceraldehyde-3phosphate

Triosephosphate Isomerase

4. Aldolase catalyzes: fructose-1,6-bisphosphate dihydroxyacetone-P + glyceraldehyde-3-P Note that C atoms are renumbered in products of Aldolase.

lysine
H H3N
+

CH2OPO32 1
COO

C CH2 CH2 CH2 CH2

+

2C

HO H H

CH C C

NH (CH2)4 + OH OH

Enzyme

4 5

CH2OPO32 6

NH3

Schiff base intermediate of Aldolase reaction

A lysine residue at the active site functions in catalysis. The keto group of fructose-1,6-bisphosphate reacts with the -amino group of the active site lysine, to form a protonated Schiff base intermediate. Cleavage of the bond between C3 & C4 follows.

1CH2OPO3 2C

O H OH OH

HO 3C H 4C H
5

Aldolase

CH2OPO32 3
2C

H
1C

1CH2OH

H 2C OH 2 3 CH2OPO3

CH2OPO32 6

fructose-1,6bisphosphate

dihydroxyacetone phosphate

glyceraldehyde-3phosphate

Triosephosphate Isomerase

5. Triose Phosphate Isomerase (TIM) catalyzes: dihydroxyacetone-P glyceraldehyde-3-P

Glyceraldehyde-3-phosphate Dehydrogenase
H
1C

O C OH

NAD+ + Pi

OPO32 + H+ O NADH 1C H CH2OPO32 3

2

CH2OPO32 3

OH

glyceraldehyde3-phosphate

1,3-bisphosphoglycerate

6. Glyceraldehyde-3-phosphate Dehydrogenase catalyzes: glyceraldehyde-3-P + NAD+ + Pi 1,3-bisphosphoglycerate + NADH + H+

NAD+ and FAD Oxidation-reduction reactions use NAD+ (nicotinamide adenine dinucleotide) FADH (flavin adenine dinucleotide). When a metabolite is oxidized, NAD+ accepts two electrons plus a hydrogen ion (H+) and NADH results. - NAD+ is reduced to NADH Conversely, NADH can also reduce a metabolite by giving up electrons. - NADH is oxidized to NAD+

Phosphoglycerate Kinase
O
1C

OPO32 ADP ATP O

1

O C

H 2C OH Mg2+ CH2OPO32 3

H 2C OH CH2OPO32 3

1,3-bisphosphoglycerate

3-phosphoglycerate

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. .

Phosphoglycerate Mutase
O
1

O C

O
1

O C

H 2C OH CH2OPO32 3

H 2C OPO32 3 CH2OH

3-phosphoglycerate

2-phosphoglycerate

8. Phosphoglycerate Mutase catalyzes: 3-phosphoglycerate 2-phosphoglycerate Phosphate is shifted from the OH on C3 to the OH on C2.

Phosphoglycerate Mutase
O
1

histidine
H H3N+
2

O C

O
1

O C

C CH2 C

COO

H 2C OH CH2OPO32 3

H 2C OPO3 3 CH2OH

HN HC
+

CH NH

3-phosphoglycerate

2-phosphoglycerate
O

An active site histidine side-chain participates in Pi transfer, by donating & accepting phosphate. The process involves a 2,3-bisphosphate intermediate.

O
1

H 2C OPO32 2 3 CH2OPO3

2,3-bisphosphoglycerate

Enolase
O
1

O C

H+ O C C

OH

O
1

O C OPO32

H 2 C OPO32 3 CH2OH

OPO32

2C

CH2OH

3 CH2

2-phosphoglycerate enolate intermediate phosphoenolpyruvate

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 ion.

Pyruvate Kinase
O C 1 C 2
3 CH2

ADP ATP

O
1 2

O C C OH

O
1 2

O C C O

OPO32

3 CH2

3 CH3

phosphoenolpyruvate

enolpyruvate

pyruvate

10. Pyruvate Kinase catalyzes: phosphoenolpyruvate + ADP pyruvate + ATP

Pyruvate Kinase
O C 1 C 2
3 CH2

ADP ATP

O
1 2

O C C OH

O
1 2

O C C O

OPO32

3 CH2

3 CH3

phosphoenolpyruvate

enolpyruvate

pyruvate

This phosphate transfer from PEP to ADP is spontaneous. PEP has a larger G of phosphate hydrolysis than ATP. Removal of Pi from PEP yields an unstable enol, which spontaneously converts to the keto form of pyruvate. Required inorganic cations K+ and Mg++ bind to anionic residues at the active site of Pyruvate Kinase.

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.

Glyceraldehyde-3-phosphate Dehydrogenase
H O
1C

Fermentation: Anaerobic organisms lack a respiratory chain.

NAD+ + Pi

OPO32 + H+ O NADH 1C H CH2OPO32 3

2

CH2OPO32 3

OH

OH

glyceraldehyde3-phosphate

1,3-bisphosphoglycerate

They must reoxidize NADH produced in Glycolysis through some other reaction, because NAD+ is needed for the Glyceraldehyde-3-phosphate Dehydrogenase reaction. Usually NADH is reoxidized as pyruvate is converted to a more reduced compound, that may be excreted. The complete pathway, including Glycolysis and the reoxidation of NADH, is called fermentation.

E.g., Lactate Dehydrogenase catalyzes reduction of the keto in pyruvate to a hydroxyl, yielding lactate, as NADH is oxidized to NAD+.

Lactate Dehydrogenase
O C C O O NADH + H+ NAD+ O C HC OH O

CH3

CH3

pyruvate

lactate

Skeletal muscles ferment glucose to lactate during exercise, when aerobic metabolism cannot keep up with energy needs. Lactate released to the blood may be taken up by other tissues, or by muscle after exercise, and converted via the reversible Lactate Dehydrogenase back to pyruvate, e.g., for entry into Krebs Cycle.

Lactate Dehydrogenase
O C C O O NADH + H+ NAD+ O C HC OH O

CH3

CH3

pyruvate

lactate

Lactate is also a significant energy source for neurons in the brain. Astrocytes, which surround and protect neurons in the brain, ferment glucose to lactate and release it. Lactate taken up by adjacent neurons is converted to pyruvate that is oxidized via Krebs Cycle.

O C C

O O

Pyruvate Decarboxylase
CO2 H C CH3 O

Alcohol Dehydrogenase
NADH + H+ NAD+ H H C OH

pyruvate

CH3

CH3

acetaldehyde

ethanol

Some anaerobic organisms metabolize pyruvate to ethanol, which is excreted as a waste product. NADH is converted to NAD+ in the reaction catalyzed by Alcohol Dehydrogenase.