Question

BIOCHEM: How are the three active sites of PDH linked?

Answer 1

PDHC comprises three principal enzymes (E1, pyruvate dehydrogenase or pyruvate decarboxylase; E2, dihydrolipoyl transacetylase; and E3, dihydrolipoyl dehydrogenase), and five different coenzymes (thiamine pyrophsphate, lipoic acid, coenzyme A, flavin adenine dinucleotide, and nicotinamide adenine dinucleotide). In addition, a specific kinase for inactivation and a phosphatase for activation of the pyruvate dehydrogenase have been demonstrated, as well as protein X . The E1 component is a tetramer containing a 41-kDa and 36-kDa subunit.

The sequence of the PDHC reactions can be simplified as follows :

E1 TPP + CH3 COCOOH → CO2 + E1 TPP-CHOHCH3

E1 TPP-CHOHCH3 + E2 lipoic(ox) → E1 TPP + CH3 COE2 lipoic

CH3 COE2 lipoic + CoASH → CH3 COSCoA + E2 lipoic (red)

E2 lipoic (red) + E3 FAD → E2 lipoic(ox) + E3 FADH2

E3 FADH2 + NAD+ → E3 FAD + NADH + H+

Pyruvate dehydrogenase is composed of four subunits: two E1 alpha and two E1 beta. The alpha subunit is X-linked. Because of the complexity of this enzyme system, it is important to have reliable methods for the enzyme assays to pinpoint the basic defect. PDHC activity is present in cultured skin fibroblasts, and therefore, these cells are a good source for enzyme diagnosis. Caution is needed to avoid mycoplasma contamination because an unusually large amount of pyruvate dehydrogenase activity has been demonstrated in contaminated cultures .

In patients with PDHC deficiency, the levels of pyruvate, lactate, and alanine are increased in blood and urine. In milder cases, random samples of blood or urine may show normal amounts of lactate. Oral glucose loads or high carbohydrate intake may induce elevations in blood lactic acid and aggravate the symptoms. Two brothers with pyruvate dehydrogenase deficiency showed absent to trace amounts of both the E1 alpha and beta subunits in liver, skeletal muscle, and heart . The lymphocytes had enzyme activity about 10% of normal controls, whereas the activity in cultured skin fibroblasts was normal. Both patients had abnormal neuromuscular development and lactic acidosis. Isolated dihydrolipoyl dehydrogenase (lipoamide dehydrogenase) deficiency has been reported in which patients presented lactic acidosis and ketosis with neurologic abnormality. However, this deficiency appeared to be more common among the Ashkenazi Jewish population . A recent study shows how the reactive oxygen species generated by the mutations responsible for lipoamide dehydrogenase deficiency may in fact explain certain disease characteristics as well as proffer the prospect of antioxidant therapy .