Cloning and expression of a novel esterase gene cpoA from Burkholderia cepacia.
OBJECTIVE
To screen and clone a novel enzyme with specific activity for the resolution of (R)-beta-acetylmercaptoisobutyrate (RAM) from (R,S)-beta-acetylmercaptoisobutyrate [(R,S)-ester].


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.


CONCLUSIONS
The cpoA cloned from B. cepacia encodes a novel esterase which is highly related to the nonhaeme chloroperoxidases.


CONCLUSIONS
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.
Kim
C H
CH
Laboratory of Metabolic Engineering, Korea Research Institute of Bioscience and Biotechnology, Oun-dong, Yusong, Daejeon, Korea.
Lee
J H
JH
Heo
J H
JH
Kwon
O S
OS
Kang
H A
HA
Rhee
S K
SK
D000595:Amino Acid Sequence; D001483:Base Sequence; D019117:Burkholderia / Q000145:classification; D016956:Burkholderia cepacia / Q000201:enzymology / Q000235:genetics*; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D004950:Esterases / Q000235:genetics*; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D016366:Open Reading Frames / Q000235:genetics; D010802:Phylogeny; D012329:RNA, Bacterial / Q000235:genetics; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D016415:Sequence Alignment
D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; D004950:Esterases; C022635:serine esterase