Pyruvate Oxidation

One source of acetyl-CoA molecules for the citric acid cycle is via oxidation of pyruvate in a reaction catalyzed by the pyruvate dehydrogenase complex. The process of converting pyruvate to acetyl-CoA is an oxidative decarboxylation. In the overall reaction (below), the carboxyl group of pyruvate is lost as CO 2, while the remaining two carbons form the acetyl moiety of acetyl-CoA.

> Pyruvate + NAD+ + CoASH <=> Acetyl-CoA + NADH + CO 2 ( = -33.5 kJ/mol)

This reaction involves electron transfer to make NADH, decarboxylation of pyruvate, and formation of actetyl-CoA, an activated two carbon compound. In yeast, acetaldehyde is formed by pyruvate decarboxylase, which is subsequently converted to ethanol by action of the enzyme alcohol dehydrogenase.

Enzymatic activities contained in the pyruvate dehydrogenase complex include:

- Pyruvate decarboxylase (E1)
- Dihydrolipoamide transacetylase (E2)
- Dihydrolipoamide dehydrogenase (E3)

and five coenzymes

- Thiamine Pyrophosphate (TPP)
- Lipoic Acid-Lipoamide
- FAD
- NAD+
- CoASH

All of these entities together make up the pyruvate dehydrogenase complex. An overview of the process is shown in Figure 14.4 and the reaction mechanism is shown in Figure 14.10. In summary, reaction steps are as follows:

> 1. E1 accepts a two carbon aldehyde from the decarboxylation of pyruvate.

> 2. The aldehyde group is transferred to the first lipoamide arm of E2 and is oxidized to an acetyl group.

> 3. The acetyl group is transferred to the second lipoamide arm of E2.

> 4. The acetyl group is linked to CoASH, forming acetyl-CoA.

> 5. E3 oxidizes the reduced lipoamide arm by transferring two hydrogens to FAD, forming FADH 2

> 6. FADH 2 is oxidized by NAD+, forming FAD and NADH + H+.

Decarboxylation of -ketoglutarate by the -ketoglutarate dehydrogenase complex (AKGDH) involves very similar reactions with the same coenzymes in a complex arrangement very much like the pyruvate dehydrogenase complex.

Allosteric Regulation

Pyruvate dehydrogenase is a major regulatory point for entry of materials into the citric acid cycle.. The enzyme is regulated allosterically and by covalent modification.

E2 - inhibited by acetyl-CoA, activated by CoA-SH

E3 - inhibited by NADH, activated by NAD+.

ATP is an allosteric inhibitor of the complex, and AMP is an activator. The activity of this key reaction is coordinated with the energy charge, the [NAD+]/[NADH] ratio, and the ratio of acetylated to free coenzyme A.

Covalent Regulation

Part of the pyruvate dehydrogenase complex, pyruvate dehydrogenase kinase, phosphorylates three specific E1 serine residues, resulting in loss of activity of pyruvate dehydrogenase. NADH and acetyl-CoA both activate the kinase. The serines are dephosphorylated by a specific enzyme called pyruvate dehydrogenase phosphatase that hydrolyzes the phosphates from the E1 subunit of the pyruvate dehydgrogenase complex. This has the effect of activating the complex. The phosphatase is activated by Ca2+ and Mg2+. Because ATP and ADP differ in their affinities for Mg2+, the concentration of free Mg2+ reflects the ATP/ADP ratio within the mitochondrion. Thus, pyruvate dehydrogenase responds to ATP levels by being turned off when ATP is abundant and further energy production is unneeded.

In mammalian tissues at rest, much less than half of the total pyruvate dehydrogenase is in the active, nonphosphorylated form. The complex can be turned on when low ATP levels signal a need to generate more ATP. The kinase protein is an integral part of the pyruvate dehydrogenase complex, whereas the phosphatase is but loosely bound.

See also: Pyruvate Dehydrogenase Kinase, Pyruvate Dehydrogenase Phosphatase

INTERNET LINK: -Keto Acid Complexes - A Review