Properties of D-hydantoinase from Agrobacterium tumefaciens and its use for the preparation of N-carbamyl D-amino acids

Abstract

Agrobacterium tumefaciens strain 47C expresses an inducible D-hydantoinase that catalyzes the formation of optically pure N-carbamyl D-amino acids from racemic hydantoin precursors. The D-Hydantoinase was shown to be active and stable at elevated temperatures and pH values, thus affording favorable bioreaction conditions that result in the racemization of DL-hydantoins to the utilizable D-isomer. The enzyme demonstrated optimal reaction kinetics at pH 10 and 70 degrees C, was not activated by metal ions, and exhibited a distinctive substrate specificity. A. tumefacins hydantoinase was most active on 5,6-dihydrouracil and DL-5-methylhydantoin with only slight activity on DL-benzylhydantoin. Extracts or whole cells of A. tumefaciens were used as biocatalyst to mediate the stereospecific conversion of DL-phenylhydantoin or DL-5-methylhydantoin to the respective N-carbamyl D-amino acids. In addition, immobilized cell systems were shown to be useful for biocatalyst reuse.