Purification and characterization of R-stereospecific amidase from Brevibacterium epidermidis ZJB-07021.

Abstract A R -stereospecific amidase was purified from Brevibacterium epidermidis ZJB-07021 and characterized in detail. The amidase was purified to homogeneity by three chromatographic steps for up to 328.9-fold with specific activity of 31.9 U mg ⿿1 . The enzyme was a homodimer with a molecular mass of 94 kDa. It exhibited maximum activity at 40 °C and pH 7.5. The enzyme was strongly inactivated by serine protease inhibitor PMSF. The values of K m and V max for racemic 2,2-dimethylcyclopropane carboxamide (DMCPCA) were 4.58 mM and 35.03 μmol min ⿿1  mg ⿿1 protein, respectively. The amidase showed a broad substrate spectrum toward aliphatic, aromatic and heterocyclic amides, but could hardly hydrolyze the bulky side-chain-containing amides. Furthermore, kinetic resolution of racemic DMCPCA by the amidase afforded S -DMCPCA in 46.3% yield and 99% ee with an average E- value of 67. These unique properties of the amidase imply that it is a promising biocatalyst for the production of chiral amides and carboxylic acids.