Asymmetric synthesis of L-α-methylcysteine with the amidase from Xanthobacter flavus NR303

Abstract

An amidase acting on D, L-α-methylcystein-amide was purified from Xanthobacter flavus NR303. The enzyme acted L-stereoselectively on D,L-α-methylcysteinamide to yield L-α-methylcysteine. Based on the N-terminal amino acid sequence of the amidase, the gene encoding the enzyme was cloned from the genomic DNA of X. flavus and sequenced. Analysis of 4840 bp of the genomic DNA revealed the presence of an open reading frame (mcaA) which encodes the amidase. This enzyme is composed of 355 amino acid residues (molecular mass, 38555 Da). The intact cells of the Escherichia coli transformant could be used for the L-stereoselective hydrolysis of racemic α-methylcysteinamide. It was activated in the presence of Mn 2+, and had maximal activity at pH 7.0 and 55°C. The E. coli transformant catalyzed the synthesis of L-α-methylcysteine from D, L-α- methylcysteinamide in a yield of 80% with high optical purity (>98% ee). © 2005 Wiley-VCH Verlag GmbH & Co.