Overexpression of the gene for N-acylamino acid racemase from Amycolatopsis sp TS-1-60 in Escherichia coli and continuous production of optically active methionine by a bioreactor

A DNA sequence encoding N-acylamino acid racemase (AAR) was inserted downstream from the T7 promoter in pET3c. The recombinant plasmid was introduced into Escherichia coli MM294 lysogenized with a bacteriophage lambda having a T7 RNA polymerase gene. The amount of AAR produced by the E. coli transformant was 1100-fold more than that produced by Amycolatopsis sp. TS-1-60, the DNA donor strain. The AAR was purified to homogeneity from the crude extract of the E. coli transformant by two steps: heat treatment and Butyl-Toyopearl column chromatography. Bioreactors for the production of optically active amino acids were constructed with DEAE-Toyopearl-immobilized AAR and D- or L-aminoacylase. D- or L-methionine was continuously produced with a high yield from N-acetyl-DL-methionine by the bioreactor.