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Expression and Characterization of Recombinant Serratia liquefaciens Nucleases Produced with Baculovirus-mediated Silkworm Expression System

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Abstract

Baculovirus-Bombyx mori protein expression system has mainly been used for translation of eukaryotic proteins. In contrast, information pertaining to bacterial protein expression using this system is not sufficient. Therefore, recombinant nucleases from Serratia liquefaciens (rSlNucAs) were expressed in a Baculovirus-B. mori protein expression system. rSlNucAs containing the native signal peptide (rSlNucA-NSP) or silkworm 30-K signal peptide (rSlNucA-30K) at the NH2-terminus were constructed to enable secretion into the extracellular fraction. Both rSlNucA-30K and rSlNucA-NSP were successfully secreted into hemolymph of B. mori larvae. Affinity-purified rSlNucAs showed high nuclease activity. Optimum pH was 7.5 and half of maximum activity was maintained between pH 7.0 and 9.5. Optimum temperature was 35 °C. rSlNucAs showed sufficient activity in twofold-diluted radioimmunoprecipitation assay buffer and undiluted, mild lysis buffer. Genomic DNA of Escherichia coli was efficiently digested by rSlNucAs in the bacterial lysate. The results in this study suggest that rSlNucAs expressed by the Baculovirus-B. mori protein expression system will be a useful tool in molecular biology. Functional recombinant protein of bacteria was produced by Baculovirus-B. mori protein expression system. This system may be highly suitable for bacterial extracellular protein secreted via Sec pathway.

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Acknowledgments

The cost of publication was supported in part by a Research Grant for Young Investigators from the Faculty of Agriculture, Kyushu University. We would like to thank Editage (www.editage.jp) for English language editing.

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    Authors and Affiliations

    1. Laboratory of Insect Pathology and Microbial Control, Faculty of Agriculture, Graduate School, Institute of Biological Control, Kyushu University, Fukuoka, Japan

      Kazuhiro Iiyama

    2. Laboratory of Insect Genome Science, Faculty of Agriculture, Graduate School, Kyushu University, Fukuoka, Japan

      Jae Man Lee, Tuneyuki Tatsuke, Hiroaki Mon & Takahiro Kusakabe

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    1. Kazuhiro Iiyama
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    Correspondence to Kazuhiro Iiyama.

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    Kazuhiro Iiyama and Jae Man Lee contributed equally to this manuscript.

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    Iiyama, K., Lee, J.M., Tatsuke, T. et al. Expression and Characterization of Recombinant Serratia liquefaciens Nucleases Produced with Baculovirus-mediated Silkworm Expression System. Mol Biotechnol 58, 393–403 (2016). https://doi.org/10.1007/s12033-016-9937-y

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