Journal of Bioscience and Bioengineering 2009-01-01

Gene cloning and characterization of arylamineN-acetyltransferase fromBacillus cereusstrain 10-L-2

Shinji Takenaka, Minyi Cheng, Mulyono, Atsushi Koshiya, Shuichiro Murakami, Kenji Aoki, Shinji Takenaka, Minyi Cheng, Mulyono, Atsushi Koshiya, Shuichiro Murakami, Kenji Aoki

Index: J. Biosci. Bioeng. 107(1) , 27-32, (2009)

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Abstract

Bacillus cereus strain 10-L-2 synthesizes two arylamine N-acetyltransferases (Nat-a and Nat-b) with broad substrate specificities toward aniline and its derivatives. In southern blot analysis using probes encoding the NH 2-terminus of Nat-b and a conserved region of N-acetyltransferases, digested total DNA of strain 10-L-2 showed one positive band. We cloned and sequenced the gene encoding Nat-b. The NH 2-terminal amino acid sequence predicted from the open reading frame (768 base pairs) corresponded to that of purified Nat-b. The cloned Nat-b gene was expressed in Escherichia coli. The expressed enzyme (BcNAT) from the recombinant strain was partially purified and characterized. Nat-b from strain 10-L-2 and BcNAT from the recombinant strain were slightly different from each others in substrate specificity and thermo-stability. We examined the biotransformations of 2-aminophenols and phenylenediamines by the whole cells of the recombinant strain. The cells converted these compounds into their corresponding acetanilides. Only one amino group of phenylenediamines was acetylated. The cells utilized 4-nitroacetanilide as an acetyl donor instead of acetyl-CoA. 4-Aminoacetanilide was produced and 4-nitroaniline was released almost stoichiometrically.