Lys-258 of aspartate aminotransferase forms a Schiff base with pyridoxal phosphate and is responsible for catalysis of the 1,3-prototropic shift central to the transamination reaction sequence. Substitution of arginine for Lys-258 stabilizes the otherwise elusive quinonoid intermediate, as assessed by the long wavelength absorption bands observed in the reactions of this mutant with several amino acid substrates. The external aldimine intermediate is not detectable during reactions of this mutant with amino acids, although the inhibitor α-methylaspartate does slowly and stably form this species. These results suggest that external aldimine formation is one of the rate-determining steps of the reaction. The pyridoxamine-5′-phosphate-like enzyme form (330-nm absorption maximum) is unreactive toward keto acid substrates, and the coenzyme bound to this species is not dissociable from the protein.
|Original language||English (US)|
|Number of pages||4|
|Journal||Journal of Biological Chemistry|
|State||Published - Dec 15 1991|
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