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KMID : 0364820210570010046
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Korean Journal of Microbiology
2021 Volume.57 No. 1 p.46 ~ p.51
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Cloning and high-level expression of the esterase gene from Klebsiella pneumoniae in Escherichia coli
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Chung Yong-Joon
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Abstract
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Optically active Dβ-Acetylthioisobutyric acid (DAT) is used as a starting material in the synthesis of captopril as a valuable chiral drug in the treatment of hypertension. For an efficient DAT production by a stereo-selective hydrolysis, Klebsiella pneumoniae CJ-317 producing a thermostable esterase was isolated in previous study. In order to construct an esterase hyper-producing strain, the esterase gene of K. pneumoniae CJ-317 was cloned into Escherichia coli and was found to show a high-level expression, which the esterase activity of the recombinant E. coli was more than 800 times higher than that of parental strain. The nucleotide sequences of 1.6 kb BamHI-EcoRI fragment containing the esterase gene were sequenced and found to include an open reading frame (ORF) of 837 bp coding 278 amino acid residues. The deduced amino acid sequence of the ORF was classified as an α/β hydrolase superfamily and showed the highest identity of 71% with that of an esterase from Pseudomonas putida MR-2068. By using the recombinant E. coli as a catalyst, highly expressing the thermostable esterase gene, an efficient production of DAT by stereo-selective hydrolysis with microbial cells could be useful.
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KEYWORD
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Klebsiella pneumoniae, DβAcetylthioisobutyric acid (DAT), esterase gene, highlevel expression, nucleotide sequence
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