Cloning and Characterization of an Enantioselective l-Menthyl Benzoate Hydrolase from Acinetobacter sp ECU2040

A new esterase gene abmbh, encoding a benzoate hydrolase which can enantioselectively hydrolyze l-menthyl benzoate to l-menthol, was recently identified from the genomic library of a soil isolate Acinetobacter sp. ECU2040. The abmbh gene contains a 1080-bp open reading frame encoding a protein of 360 amino acids with a calculated molecular mass of 40.7 kDa. The corresponding enzyme AbMBH was functionally expressed in Escherichia coli BL21 (DE3), purified, and characterized. The AbMBH displayed the maximum activity towards p-nitrophenyl butyrate at 50 degrees C, and an optimum pH of 8.5. A KM of 2.6 mMand a k(cat) of 0.26 s(-1) were observed towards dl-menthyl benzoate. The AbMBH exhibited a moderate enantioselectivity (E=27.5) towards dl-menthyl benzoate. It can also catalyze the enantioselective hydrolysis of a variety of racemic menthyl esters, including dl-menthyl acetate, dl-menthyl chloroacetate, and dl-menthyl butyrate.