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Cloning, Overexpression, and Characterization of a High Enantioselective Nitrilase from Sphingomonas wittichii RW1 for Asymmetric Synthesis of (R)-Phenylglycine

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Abstract

In this study, a high (R)-enantioselective nitrilase gene from Sphingomonas wittichii RW1 was cloned and overexpressed in Escherichia coli BL21 (DE3). The recombinant nitrilase was purified to homogeneity with a molecular weight of 40 kDa. The pH and temperature optima were shown to be pH 8.0 and 40 °C, respectively. The purified nitrilase was most active toward succinonitrile, approximately 30-fold higher than that for phenylglycinonitrile. Using the E. coli BL21/ReSWRW1 whole cells as biocatalysts, the kinetic resolution for asymmetric synthesis of (R)-phenylglycine was investigated at pH 6.0. A yield of 46 % was obtained with 95 % enantiomeric excess (ee), which made it a promising biocatalyst for synthesis of (R)-phenylglycine.

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Acknowledgments

This work was supported by the National Basic Research Program of China (No. 2012CB721003) and the Fundamental Research Funds for the Central Universities.

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Authors and Affiliations

  1. State Key Laboratory of Bioreactor Engineering, New World Institute of Biotechnology, East China University of Science and Technology, Shanghai, 200237, People’s Republic of China

    Jian Qiu, Hua-Lei Wang, Wen-Wen Cai, Wei Wang & Dong-Zhi Wei

  2. Enzyme and Fermentation Technology Laboratory, College of Light Industry Science and Engineering, Nanjing Forestry University, Nanjing, 210037, People’s Republic of China

    Er-Zheng Su

Authors
  1. Jian Qiu
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  2. Er-Zheng Su
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  3. Hua-Lei Wang
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  4. Wen-Wen Cai
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  5. Wei Wang
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  6. Dong-Zhi Wei
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Correspondence to Er-Zheng Su or Dong-Zhi Wei.

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Qiu, J., Su, EZ., Wang, HL. et al. Cloning, Overexpression, and Characterization of a High Enantioselective Nitrilase from Sphingomonas wittichii RW1 for Asymmetric Synthesis of (R)-Phenylglycine. Appl Biochem Biotechnol 173, 365–377 (2014). https://doi.org/10.1007/s12010-014-0845-y

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  • DOI: https://doi.org/10.1007/s12010-014-0845-y

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