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Cloning and expression of a novel esterase gene cpoA from Burkholderia cepacia
Author(s) -
Kim C.H.,
Lee J.H.,
Heo J.H.,
Kwon O.S.,
Kang H.A.,
Rhee S.K.
Publication year - 2004
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1111/j.1365-2672.2004.02262.x
Subject(s) - esterase , catalytic triad , biology , gene , burkholderia , cloning (programming) , peptide sequence , serine , biochemistry , sequence analysis , molecular cloning , microbiology and biotechnology , homology (biology) , genetics , enzyme , bacteria , computer science , programming language
Aims: To screen and clone a novel enzyme with specific activity for the resolution of (R)‐ β ‐acetylmercaptoisobutyrate (RAM) from (R,S)‐ β ‐acetylmercaptoisobutyrate [(R,S)‐ester]. Methods and Results: A micro‐organism that produces a novel esterase was isolated and identified as the bacterium Burkholderia cepacia by using the analysis of cellular fatty acids, Biolog automated microbial identification/characterization system, and 16S rRNA gene sequence analysis. A novel esterase gene was cloned from the chromosomal DNA of B. cepacia and was designated as cpoA . The cpoA encodes a polypeptide of 273 amino acids which shows a strong sequence homology with many bacterial nonhaeme chloroperoxidases. In addition, a typical serine‐hydrolase motif, Gly‐X‐Ser‐X‐Gly, and the highly conserved catalytic triad, Ser95, Asp224, and His253, were identified in the deduced amino acid sequence of cpoA by multiple sequence alignment. Conclusion: The cpoA cloned from B. cepacia encodes a novel esterase which is highly related to the nonhaeme chloroperoxidases. Significance and Impact of the Study: This is the first report that describes the isolation and cloning of a serine esterase gene from B. cepacia , which is useful in the chiral resolution of (R,S)‐ester. The cloned gene will allow additional research on the bifunctionality of the enzyme with esterase and chloroperoxidase activity at the structural and functional levels.