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The Citric Acid Cycle

The citric acid cycle (Krebs cycle) is a central metabolic pathway that generates energy through the oxidation of acetyl-CoA derived from carbohydrates, fats, and proteins. It takes place in the matrix of mitochondria and involves 8 steps and 10 enzymes that completely oxidize acetyl-CoA, generating carbon dioxide and reducing equivalents in the form of NADH and FADH2 that are used in oxidative phosphorylation. Pyruvate dehydrogenase is a multi-enzyme complex that converts pyruvate into acetyl-CoA, an essential substrate for the citric acid cycle. It consists of multiple copies of three catalytic components and requires five cofactors and coenzymes to carry out the reaction

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Raju Gangadharan
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95 views

The Citric Acid Cycle

The citric acid cycle (Krebs cycle) is a central metabolic pathway that generates energy through the oxidation of acetyl-CoA derived from carbohydrates, fats, and proteins. It takes place in the matrix of mitochondria and involves 8 steps and 10 enzymes that completely oxidize acetyl-CoA, generating carbon dioxide and reducing equivalents in the form of NADH and FADH2 that are used in oxidative phosphorylation. Pyruvate dehydrogenase is a multi-enzyme complex that converts pyruvate into acetyl-CoA, an essential substrate for the citric acid cycle. It consists of multiple copies of three catalytic components and requires five cofactors and coenzymes to carry out the reaction

Uploaded by

Raju Gangadharan
Copyright
© Attribution Non-Commercial (BY-NC)
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Download as PPT, PDF, TXT or read online on Scribd
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The Citric acid cycle

4/16/2003

The Citric acid cycle


It is called the Krebs cycle or the tricarboxylic and is the hub of the metabolic system. It accounts for the majority of carbohydrate, fatty acid and amino acid oxidation. It also accounts for a majority of the generation of these compounds and others as well.
Amphibolic - acts both catabolically and anabolically 3NAD+ + FAD + GDP + Pi + acetyl-CoA 3NADH + FADH + GTP + CoA + 2CO2

History
By 1930 it was established that the addition of lactate, acetate succinate, malate, a-ketoglutaric acid (dicarboxylic acids) and citrate and isocitrate (tricarboxylic acids) when added to muscle mince that they stimulated oxygen consumption and release of CO2

1935Albert Szent-Gyorgyi showed that


Succinate Citrate succinate Fumarate cis-aconitate fumarate Malate isocitrate malate Oxaloacetate a ketoglutarate oxaloacetate Carl Martius and Franz Knoop showed

Martius and Knoop showed that pyruvate and oxaloacetate could form citrate non-enzymatically by the addition of peroxide under basic conditions.
Krebs showed that succinate is formed from fumarate, malate or oxaloacetate. This is interesting since it was shown that the other way worked as well!! Pyruvate can form citrate enzymatically Pyruvate + oxaloacetate citrate + CO2

The interconversion rates of the intermediates was fast enough to support respiration rates.

Overview

The citric acid cycle enzymes are found in the matrix of the mitochondria
Substrates have to flow across the outer and inner parts of the mitochondria

Nathan Kaplan and Fritz Lipmann discovered Coenzyme A and Ochoa and Lynen showed that acetylCoA was intermediate from pyruvate to citrate.

CoA acts as a carrier of acetyl groups

Acetyl-CoA is a high energy compound: The DG' for the hydrolysis of its thioester is -31.5 kJ mol-1 making it greater than the hydrolysis of ATP

Pyruvate dehydrogenase converts pyruvate to acetyl-CoA and CO2

Pyruvate dehydrogenase
A multienzyme complexes are groups of non covalently associated enzymes that catalyze two or more sequential steps in a metabolic pathway. Molecular weight of 4,600,000 Da E. coli Pyruvate dehydrogenase -E1 24 24 12 yeast 60 60 12

dihydrolipoyl transacetylase --E2 dihydrolipoyl dehydrogenase--E3

24 E2 subunits

24 E1 orange 12 E3 Red

a and b together

EM based image of the core E2 from yeast pyruvate dh


60 subunits associated as 20 cone-shaped trimers that are verticies of a dodecahedron

Why such a complex set of enzymes?


1 Enzymatic reactions rates are limited by diffusion, with shorter distance between subunits a enzyme can almost direct the substrate from one subunit (catalytic site) to another.
2. Channeling metabolic intermediates between successive enzymes minimizes side reactions 3. The reactions of a multienzyme complex can be coordinately controlled

Covalent modification of eukaryotic pyruvate dehydrogenase

The five reactions of the pyruvate dehydrogenase multi enzyme complex

The enzyme requires five coenzymes and five reactions


Pyruvate + CoA + NAD+ acetyl-CoA + CO2 + NADH

The Coenzymes and prosthetic groups of pyruvate dehydrogenase


Cofactor
Thiamine pyrophosphate Lipoic acid

Location
Bound to E1 Covalently linked to a Lys on E2 (lipoamide)

Function
Decarboxylates pyruvate Accepts hydroxyethyl carbanion from TPP

CoenzymeA
FAD (flavin) NADH

Substrate for E2
Bound to E3 Substrate for E3

Accepts acetyl group from lipoamide reduced by lipoamide


reduced by FADH2

Domain structure of dihydrolipoyl transacetylase E2

Pyruvate dehydrogenase
1. Pyruvate dh decarboxylates pyruvate using a TPP cofactor forming hydroxyethyl-TPP. 2 The hydroxyethyl group is transferred to the oxidized lipoamide on E2 to form Acetyl dihydrolipoamide-E2 3 E2 catalyzes the transfer of the acetyl groups to CoA yielding acetyl-CoA and reduced dihydrolipoamide-E2 4 Dihydrolipoyl dh E3 reoxidizes dihydrolipoamide-E2 and itself becomes reduced as FADH2 is formed

5 Reduced E3 is reoxidized by NAD+ to form FAD and NADH The enzymes SH groups are reoxidized by the FAD and the electrons are transferred to NADH

H3C N C HO C S S

R1 S CH3

H3C N C HO C S HS

R1 S CH3

H3C N C

R1 S

+
O CH3 S
+

HS

E2
O CH3 S
+

E2
E2

CoA

SH

HS

O
HS

HS

+
E2

CoA

S Ch3

E2

FAD
S S +
HS HS

FAD SH SH +
S S

E2

E2

NAD+

NADH + H+

FAD
SH SH

FADH2
S S

FAD
S S

CH3 S HS

O S S S S SH S

CH3

E2

E2

E2

E2

CH3 S S S

O HS SH S S S

CH3

E2

E2

E2

E2

Arsenite or organic arsenical compound inhibition


S O- As HS R S R

OH OAs OH +

HS

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