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Cloning, sequencing, and expression of a novel epoxide hydrolase gene from Rhodococcus opacus in Escherichia coli and characterization of enzyme

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Abstract

An epoxide hydrolase gene of about 0.8 kb was cloned from Rhodococcus opacus ML-0004, and the open reading frame (ORF) sequence predicted a protein of 253 amino acids with a molecular mass of about 28 kDa. An expression plasmid carrying the gene under the control of the tac promotor was introduced into Escherichia coli, and the epoxide hydrolase gene was successfully expressed in the recombinant strains. Some characteristics of purified recombinant epoxide hydrolase were also studied. Epoxide hydrolase showed a high stereospecificity for l(+)-tartaric acid, but not for d(+)-tartaric acid. The epoxide hydrolase activity could be assayed at the pH ranging from 3.5 to 10.0, and its maximum activity was obtained between pH 7.0 and 7.5. The enzyme was sensitive to heat, decreasing slowly between 30°C and 40°C, and significantly at 45°C. The enzyme activity was activated by Ca2+ and Fe2+, while strongly inhibited by Ag+ and Hg+, and slightly inhibited by Cu2+, Zn2+, Ba2+, Ni+, EDTA–Na2 and fumarate.

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Acknowledgement

We wish to thank Dr. Fengjie Cui, School of Food Engineering and Biotechnology, Jiangsu University for his thoughtful suggestions and correction of the English version on an earlier manuscript.

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Authors and Affiliations

  1. Institute of Bioengineering, Zhejiang University of Technology, Hangzhou, 310014, People’s Republic of China

    Zhiqiang Liu & Yuguo Zheng

  2. Department of Plant Science, North Dakota State University, Fargo, ND, 58105, USA

    Yin Li & Yingying Xu

  3. Institute of Quality and Standard for Agricultural Products, Zhejiang Academy of Agricultural Sciences, Hangzhou, 310021, People’s Republic of China

    Lifeng Ping

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  1. Zhiqiang Liu
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  2. Yin Li
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Correspondence to Yuguo Zheng.

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Liu, Z., Li, Y., Xu, Y. et al. Cloning, sequencing, and expression of a novel epoxide hydrolase gene from Rhodococcus opacus in Escherichia coli and characterization of enzyme. Appl Microbiol Biotechnol 74, 99–106 (2007). https://doi.org/10.1007/s00253-006-0635-8

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  • DOI: https://doi.org/10.1007/s00253-006-0635-8

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