Biosynthesis of porphyrin precursors. Purification and characterization of mammalian L-alanine:gamma,delta-dioxovaleric acid aminotransferase.

Abstract

Bovine liver mitochondria have been found to contain an enzyme which will catalyze the formation of delta-aminolevulinic acid via a transamination reaction rather than via the condensation of glycine and succinyl coenzyme A. The enzyme, L-alanine: gamma,delta-dioxovaleric acid aminotransferase (gamma,delta-dioxovalerate transaminase) was isolated and purifed to apparent homogeneity. gamma,delta-Dioxovalerate transaminase is quite stable, has optimal activity at pH 6.9, requires pyridoxal phosphate as a cofactor and has an apparent molecular weight of 240,000. The enzyme has high specificity for both substrates. The Km for L-alanine is 3.7 x 10(-3) M and the Km for gamma,delta-dioxovalerate is 2.4 x 10(-4) M. Plots of 1/gamma,delta-dioxovalerate against 1/v at varying alanine concentrations suggested a ping-pong reaction mechanism. Although the enzyme appeared to be a typical transaminase, exhaustive experiments failed to demonstrate reversibility of the reaction. The capacity of gamma,delta-dioxovalerate transaminase to synthesize delta-aminolevulinic acid appears to be far greater than the capacity of delta-aminolevulinic acid synthase from the same source. The possibility that gamma,delta-dioxovalerate transaminase plays a role in the biosynthesis of delta-aminolevulinic acid in vivo must be considered.