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Isolation of a Novel Lipase Gene from Serratia liquefaciens S33 DB-1, Functional Expression in Pichia pastoris and its Properties

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Abstract

A new lipase gene designated as SlLipA was isolated from Serratia liquefaciens S33 DB-1 by the genomic-walking method. The cloned gene contained an open reading frame (ORF) of 1,845 bp encoding 615 amino acids with a conserved GXSXG motif. Genome sequence analysis showed that an aldo/keto reductase gene closed to the SlLipA gene. The lipase gene was cloned into the expression vector pPICZαA and successfully integrated into the heterologous host, methylotrophic yeast Pichia pastoris GS115. Five transformants could be expressed as secreted recombinant proteins with the high activity on Triglyceride–Agarose plate and as candidates to produce the recombinant enzyme. A C-terminal His tag was used for its purification. The lipase activity of different transformants against substrate para-nitrophenyl laurate (p-NPL) varied from 14 to 16 U ml−1. For the substrates para-nitrophenyl caprate (p-NPC), p-NPL, para-nitrophenyl myristate (p-NPM), para-nitrophenyl palmitate (p-NPP), and para-nitrophenyl stearate (p-NPS), the specific activity was shown to be preferred to long acyl chain length of p-NPS.

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Acknowledgments

We are grateful to Dr. Wenxin Zou for kindly providing S. liquefaciens S33 DB-1 strain, and we thank China National “863” High-Tech Program, China Ministry of Education and Shanghai Science and Technology Committee for fund support under Project No. 2006AA020202.

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Authors and Affiliations

  1. Plant Biotechnology Research Center, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, School of Agriculture and Biology, School of Life Science and Technology, Shanghai Jiao Tong University, Shanghai, 200030, P.R. China

    Hongyan Yao, Lida Zhang, Kaijing Zuo, Hua Ling, Fei Zhang & Kexuan Tang

  2. Shanghai Agrobiological Gene Center, Shanghai Academy of Agricultural Sciences, Shanghai, 201106, P.R. China

    Shunwu Yu

  3. State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Morgan-Tan International Center for Life Sciences, Fudan University, Shanghai, 200433, P.R. China

    Kexuan Tang

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  1. Hongyan Yao
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  2. Shunwu Yu
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Correspondence to Kexuan Tang.

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Yao, H., Yu, S., Zhang, L. et al. Isolation of a Novel Lipase Gene from Serratia liquefaciens S33 DB-1, Functional Expression in Pichia pastoris and its Properties. Mol Biotechnol 38, 99–107 (2008). https://doi.org/10.1007/s12033-007-9007-6

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  • DOI: https://doi.org/10.1007/s12033-007-9007-6

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