Lecture 10: Pyruvate Dehydrogenase Complex

Pyruvate dehydrogenase complex (PDH)

Converts Pyruvate to acetyl CoA

Thiamine pyrophosphate (TPP)

Cofactor for E1, Thiazolium ring is an important functional component for aldehyde transport, derived from thiamine (vitamin B1)

Coenzyme A (Co-A)

Cofactor for E2, derived from pantothenic acid (vitamin B5), active group becomes acetylated, acetate ion is high free energy for hydrolysis to activate it for later reactions, is made up of nucleotides

Lipoic acid

Cofactor for E2, not from a vitamin, covalently bound to lysine, can adopt three different forms (oxidized, reduced, Acetylated)

FAD and NAD+

Cofactors for E3, educing equivalents

Flavin Adenine Dinucleotide (FAD) and Flavin Mononucleotide (FMN): not mobile, can accept 2 electrons total, one at a time in the form of H+, derived from riboflavin (Vitamin B2), reduced to FADH2

Nicotinamide adenine dinucleotide (NAD+): can diffuse and move, can accept 2 electrons at the same time in the form of H-, derived from Niacin (Vitamin B3), reduced to NADH

How does pyruvate enter the mitochondrial matrix

Through facilitated diffusion via a specific Uniport

Step one of the PDH mechanism: oxidative decarboxylation

Occurs on E1 (pyruvate dehydrogenase), pyruvate is decarboxylated, acetyl group is transferred onto TPP to form hydroxyl ethyl TPP

Step two of the PDH mechanism: reduction of lipoic acid

Here’s on E2 (Dihydrolipoyl Transacetylase), the lipoamide arm swings over to E1, hydroxyl group is oxidized to an acetyl group and the disulfide bond of the lipoamide is reduced, E1 catalyzes the transfer of the hydroxyethyl group to the oxidized lipoamide arm of E2

Step three of the PDH mechanism: formation of acetyl CoA

Also on E2, the acetyl group is transferred from the sulfur atom of Lipoamide to the thiol group of coenzyme A, forms acetyl CoA

Step four of the PDH mechanism: reduction of FAD and reoxidation of E2

Occurs on E3 (Dihydrolipoyl dehydrogenase), both CO2 and acetyl-CoA are released from the complex electrons are transferred from the reduced thiol group of lipoic acid to FAD to reduce it to FADH2, regenerates E2 to the oxidized state

Step five of the PDH mechanism: reduction of NAD+ and Reoxidation of FADH2

Also on E3 FADH2 needs to be re-oxidized because it is not mobile so FADH2 passes its electrons to NAD+ to reduce it to NADH because it is a mobile electron carrier and will bring the electrons to the electron transport chain

PDH regulation

Regulated by the energy status and Reaction products

Inhibited: high energy (ATP, Acetyl CoA, NADH)

Activated: low energy (AMP, CoA-SH, NAD+)

Covalent modifications

Activated by pyruvate dehydrogenase phosphate (PDP),

Inactivated by pyruvate dehydrogenase kinase (PDK)

pyruvate dehydrogenase kinase (PDK)

Inactivates E1, associated with E2, phosphorylate regulatory serine residues in E1, does not respond to glucagon

pyruvate dehydrogenase phosphate (PDP)

Activates E1 by Dephosphorylating regulatory serine residues, responds to insulin, is associated with E2

Phosphatase 1

In muscles, regulated by Ca2+

Phosphatase 2

In liver, regulated by hormones/metabolic state

A 1 month old male shows neurologic problems and lactic acidosis. Enzyme assay for pyruvate dehydrogenase (PDH) complex activity on extracts of cultured skin fibroblasts showed 5% normal activity with the low concentration of thiamine phosphate (TPP) but 80% of normal activity when the assay contained 8000 fold higher concentration of TPP. Which of the following statements concerning this patient is correct?

Administration of thiamine is expected to reduce the serum lactate level and improve his clinical symptoms