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Cloning and expression of gene, and activation of an organic solvent-stable lipase from Pseudomonas aeruginosa LST-03

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Abstract

Organic solvent-tolerant Pseudomonas aeruginosa LST-03 secretes an organic solvent-stable lipase, LST-03 lipase. The gene of the LST-03 lipase (Lip9) and the gene of the lipase-specific foldase (Lif9) were cloned and expressed in Escherichia coli. In the cloned 2.6 kbps DNA fragment, two open reading frames, Lip9 consisting of 933 nucleotides which encoded 311 amino acids and Lif9 consisting of 1,020 nucleotides which encoded 340 amino acids, were found. The overexpression of the lipase gene (lip9) was achieved when T7 promoter was used and the signal peptide of the lipase was deleted. The expressed amount of the lipase was greatly increased and overexpressed lipase formed inclusion body in E. coli cell. The collected inclusion body of the lipase from the cell was easily solubilized by urea and activated by using lipase-specific foldase of which 52 or 58 amino acids of N-terminal were deleted. Especially, the N-terminal methionine of the lipase of which the signal peptide was deleted was released in E. coli and the amino acid sequence was in agreement with that of the originally-produced lipase by P. aeruginosa LST-03. Furthermore, the overexpressed and solubilized lipase of which the signal peptide was deleted was more effectively activated by lipase-specific foldase.

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Acknowledgments

This work was supported by Industrial Technology Research Grant Program in ‘05 from New Energy and Industrial Technology Development Organization (NEDO) of Japan and a Grant-in-Aid for Scientific Research (B) (17310131) from the Japan Society for the Promotion of Science. The authors would like to express their appreciation for these supports

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Authors and Affiliations

  1. Department of Chemical Engineering, Osaka Prefecture University, 1-1 Gakuen-cho, Naka-ku, Sakai, Osaka, 599-8531, Japan

    Hiroyasu Ogino, Yoshikazu Katou, Rieko Akagi, Takashi Mimitsuka, Shinichi Hiroshima, Yuichi Gemba, Masahiro Yasuda, Kosaku Ishimi & Haruo Ishikawa

  2. Department of Life Sciences, Toyo University, Oura-gun, Gunma, 374-0193, Japan

    Noriyuki Doukyu

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  1. Hiroyasu Ogino
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  2. Yoshikazu Katou
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  3. Rieko Akagi
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  4. Takashi Mimitsuka
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  5. Shinichi Hiroshima
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  6. Yuichi Gemba
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  7. Noriyuki Doukyu
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  8. Masahiro Yasuda
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  9. Kosaku Ishimi
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  10. Haruo Ishikawa
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Correspondence to Hiroyasu Ogino.

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Communicated by K. Horikoshi.

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Ogino, H., Katou, Y., Akagi, R. et al. Cloning and expression of gene, and activation of an organic solvent-stable lipase from Pseudomonas aeruginosa LST-03. Extremophiles 11, 809–817 (2007). https://doi.org/10.1007/s00792-007-0101-2

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  • DOI: https://doi.org/10.1007/s00792-007-0101-2

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