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Recombinant production of hyperthermostable CelB from Pyrococcus furiosus in Lactobacillus sp.

  • Biotechnological products and process engineering
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Abstract

Lactic acid bacteria (LAB) are used widespread in the food industry as traditional starters for various fermented foods. For recombinant protein production, LAB would be superior with view from the food safety demands since most of them are Generally Recognized As Safe organisms. We investigated the two pSIP expression systems, pSIP403 and pSIP409 (Sørvig et al. 2005), to produce a hyper-thermophilic β-glycosidase (CelB) from Pyrococcus furiosus in Lactobacillus plantarum NC8 and Lactobacillus casei as hosts, respectively. Both lactobacilli harboring the pSIP409-celB vector produced active CelB in batch bioreactor cultivations (MRS medium) while the specific CelB activity of the cell free extract was about 44 % higher with L. plantarum (1,590 ± 90 nkat/mgprotein) than with L. casei (1,070 ± 66 nkat/mgprotein) using p-nitrophenyl-β-galactoside (pNPGal) as the substrate. A fed-batch bioreactor cultivation of L. plantarum NC8 pSIP409-celB resulted in a specific CelB activity of 2,500 ± 120 nkat pNPGal/mgprotein after 28 h. A repeated dosage of the inducer spp-IP did not increase the enzyme expression further. As alternative for the cost intensive MRS medium, a basal whey medium with supplements (yeast extract, Tween 80, NH4-citrate) was developed. In bioreactor cultivations using this medium, about 556 ± 29 nkat pNPGal/mgprotein of CelB activity was achieved. It was shown that both LAB were potential expression hosts for recombinant enzyme production. The pSIP expression system can be applied in L. casei.

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Acknowledgements

The authors wish to thank Dr. Lars Axelsson for providing the plasmid pSIP403, plasmid pSIP409, and L. plantarum NC8. Further thanks the group of Prof. de Vos and Dr. Kengen, University of Wageningen (NL), for the abandoning of CelB gene. Parts of the research were financed by the German Federal Ministry of Economics and Technology (AiF/FEI Project No. 15801 N) which is greatly acknowledged.

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  1. Institute of Food Science and Biotechnology, Department of Biotechnology, University of Hohenheim, Garbenstr. 25, 70599, Stuttgart, Germany

    N. Böhmer, S. Lutz-Wahl & L. Fischer

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  1. N. Böhmer
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  2. S. Lutz-Wahl
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  3. L. Fischer
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Correspondence to L. Fischer.

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Böhmer, N., Lutz-Wahl, S. & Fischer, L. Recombinant production of hyperthermostable CelB from Pyrococcus furiosus in Lactobacillus sp.. Appl Microbiol Biotechnol 96, 903–912 (2012). https://doi.org/10.1007/s00253-012-4212-z

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  • DOI: https://doi.org/10.1007/s00253-012-4212-z

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