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Gene Cloning, Overexpression, and Characterization of the Nitrilase from Rhodococcus rhodochrous tg1-A6 in E. coli

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Abstract

A DNA fragment containing the entire coding sequence of nitrilase gene was amplified from Rhodococcus rhodochrous tg1-A6 with high nitrilase activity using PCR and sequenced. The open reading frame of the nitrilase gene contains 1,101 base pairs, which encodes a putative polypeptide of 366 amino acid residues. The nitrilase gene was cloned into an expression vector pET-28a and expressed in an Escherichia coli strain BL21(DE3). The enzymatic activity of nitrilase, which converts various nitriles to the corresponding carboxylic acids, was detected to reach 24.5 U/ml at 9 h in the recombinant bacteria.

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Authors and Affiliations

  1. Department of Biological Science and Technology, University of Science and Technology Beijing, Beijing, 100083, People’s Republic of China

    Hui Luo, Lu Fan & Jinwei Ma

  2. Department of Environmental Engineering, University of Science and Technology Beijing, Beijing, 100083, People’s Republic of China

    Lu Fan, Yanhong Chang & Jinwei Ma

  3. Department of Chemical Engineering, Tsinghua University, Beijing, 100084, People’s Republic of China

    Huimin Yu & Zhongyao Shen

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  1. Hui Luo
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  2. Lu Fan
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  3. Yanhong Chang
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  4. Jinwei Ma
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  5. Huimin Yu
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  6. Zhongyao Shen
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Correspondence to Hui Luo.

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Luo, H., Fan, L., Chang, Y. et al. Gene Cloning, Overexpression, and Characterization of the Nitrilase from Rhodococcus rhodochrous tg1-A6 in E. coli . Appl Biochem Biotechnol 160, 393–400 (2010). https://doi.org/10.1007/s12010-008-8324-y

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  • DOI: https://doi.org/10.1007/s12010-008-8324-y

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