Nucleotide Sequence and Genetic Structure of a Novel Carbaryl Hydrolase Gene ( cehA ) from Rhizobium sp. Strain AC100

Abstract

ABSTRACT Rhizobium sp. strain AC100, which is capable of degrading carbaryl (1-naphthyl- N -methylcarbamate), was isolated from soil treated with carbaryl. This bacterium hydrolyzed carbaryl to 1-naphthol and methylamine. Carbaryl hydrolase from the strain was purified to homogeneity, and its N-terminal sequence, molecular mass (82 kDa), and enzymatic properties were determined. The purified enzyme hydrolyzed 1-naphthyl acetate and 4-nitrophenyl acetate indicating that the enzyme is an esterase. We then cloned the carbaryl hydrolase gene ( cehA ) from the plasmid DNA of the strain and determined the nucleotide sequence of the 10-kb region containing cehA . No homologous sequences were found by a database homology search using the nucleotide and deduced amino acid sequences of the cehA gene. Six open reading frames including the cehA gene were found in the 10-kb region, and sequencing analysis shows that the cehA gene is flanked by two copies of insertion sequence-like sequence, suggesting that it makes part of a composite transposon.