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Regulated expression systems for the development of whole-cell biocatalysts expressing oxidative enzymes in a sequential manner

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Abstract

This work reports the preparation of two recombinant strains each containing two enzymatic activities mutually expressed through regulated systems for production of functionalized epoxides in one-pot reactions. One strain was Pseudomonas putida PaW340, containing the gene coding for styrene monooxygenase (SMO) from Pseudomonas fluorescens ST under the auto-inducing Ptou promoter and the TouR regulator of Pseudomonas sp. OX1 and the gene coding for naphthalene dihydrodiol dehydrogenase (NDDH) from P. fluorescens N3 under the Ptac promoter inducible by IPTG. The second strain was Escherichia coli JM109, in which the expression of SMO was under the control of the Pnah promoter and the NahR regulator of P. fluorescens N3 inducible by salicylate, while the gene expressing NDDH was under the control of the Plac promoter inducible by IPTG. SMO and NDDH activities were tested in bioconversion experiments using cinnamyl alcohol as reference substrate. The application that we selected is one example of the sequential use of the two enzymatic activities which require a temporal control of the expression of both genes.

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Acknowledgments

This work was supported by MIUR-PRIN 2007-2009 (Grant 20077MY8M9 004) and by MIUR-FAR 2011.

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Authors and Affiliations

  1. Department of Biotechnology and Biosciences, University of Milano-Bicocca, Piazza della Scienza 2, 20126, Milan, Italy

    Patrizia Di Gennaro, Linda Veronique Kazandjian & Francesco Mezzetti

  2. Department of Organic and Industrial Chemistry, University of Milano, via Venezian 21, 20133, Milan, Italy

    Guido Sello

Authors
  1. Patrizia Di Gennaro
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  2. Linda Veronique Kazandjian
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  3. Francesco Mezzetti
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  4. Guido Sello
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Correspondence to Patrizia Di Gennaro.

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Communicated by Eriko Takano.

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Di Gennaro, P., Kazandjian, L.V., Mezzetti, F. et al. Regulated expression systems for the development of whole-cell biocatalysts expressing oxidative enzymes in a sequential manner. Arch Microbiol 195, 269–278 (2013). https://doi.org/10.1007/s00203-013-0875-9

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  • DOI: https://doi.org/10.1007/s00203-013-0875-9

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