Detection of intermediates in reactions catalyzed by PLP-dependent enzymes: O-acetylserine sulfhydrylase and serine-glyoxalate aminotransferase

Abstract

All pyridoxal 5'-phosphate (PLP)-dependent enzymes catalyze their reaction via the initial formation of an external Schiff base intermediate. The internal and external Schiff bases exist in several different tautomeric forms. The chapter illustrates two predominant forms, the enolimine and ketoenamine tautomers, along with the ranges of their respective λ max values. It shows ketoenamine form in the case of the external Schiff base. The external Schiff base is formed via the intermediacy of geminal-diamine intermediates. Once the external Schiff base is formed, the reaction pathway is determined by the kind of reaction catalyzed, and this is determined by the identity of the functional group on the a-carbon that is orthogonal to the PLP ring. O-acetylserine sulfhydrylase (OASS) catalyzes the final step in the de novo synthesis of L-cysteine in enteric bacteria, that is, the conversion of O-acetyl-L-serine (OAS) and bisulfide to L-cysteine and acetate. The cysK gene, encoding for OASS-A, has been cloned and sequenced from Salmonella typhimurium , and the enzyme overexpressed using the wild-type cys K promoter.