Reactions of human liver peroxisomal alanine:glyoxylate aminotransferase with beta-chloro-L-alanine and L-cysteine: spectroscopic and kinetic analysis.

In addition to the main transaminase reaction, the pyridoxal 5'-phosphate-dependent enzyme human liver peroxisomal alanine:glyoxylate aminotransferase (AGT) is able to catalyze the alpha,beta-elimination of beta-chloro-L-alanine with a catalytic efficiency similar to that of the physiological transaminase reaction with L-alanine. On the other hand, during the reaction of AGT with L-cysteine, changes in the coenzyme forms and analysis of the products reveal the occurrence of both beta-elimination and half-transamination of L-cysteine together with the pyruvate transamination. A mechanism in which a ketimine species is the common intermediate of half-transamination and beta-elimination of L-cysteine is proposed. L-cysteine partitions between these two reactions with a ratio of approximately 2.5. Rapid scanning stopped-flow and quench flow experiments permit the identification of reaction intermediates and the measurements of the kinetic parameters of L-cysteine half-transamination. The k(cat) of this reaction is 200- or 60-fold lower than that of L-alanine and L-serine, respectively. Conversely, L-cysteine binds to AGT with a binding affinity 30- and 200-fold higher than that of L-alanine and L-serine, respectively. This appears to be consistent with the calculated interaction energies of the L-cysteine, L-alanine and L-serine docked at the active site of AGT.