Theoretical Studies on Pyridoxal 50-Phosphate-Dependent Transamination of a-Amino Acids

Abstract: Density functional methods have been applied to investigate the irreversible transamination between gly-oxylic acid and pyridoxamine analog and the catalytic mechanism for the critical [1,3] proton transfer step in aspar-tate aminotransferase (AATase). The results indicate that the catalytic effect of pyridoxal 50-phosphate (PLP) may be attributed to its ability to stabilize related transition states through structural resonance. Additionally, the PLP hydroxyl group and the carboxylic group of the amino acid can shuttle proton, thereby lowering the barrier. The rate-limiting step is the tautomeric conversion of the aldimine to ketimine by [1,3] proton transfer, with a barrier of 36.3 kcal/mol in water solvent. A quantum chemical model consisting 142 atoms was constructed based on the crys-tal structure of the native AATase complex with the product L-glutamate. The electron-withdrawing stabilization by various residues, involving Arg386, Tyr225, Asp222, Asn194, and peptide backbone, enhances the carbon acidity of 40-C of PLP and Ca of amino acid. The calculations support the proposed proton transfer mechanism in which Lys258 acts as a base to shuttle a proton from the 40-C of PLP to Ca of amino acid. The first step (proton transfer from 40-C to lysine) is shown to be the rate-limiting step. Furthermore, we provided an explanation for the revers-ibility and specificity of the transamination in AATase