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A host-vector toolbox for improved secretory protein overproduction in Bacillus subtilis
- Target proteins in biotechnological applications are highly diverse. Therefore, versatile flexible expression systems for their functional overproduction are required. In order to find the right heterologous gene expression strategy, suitable host-vector systems, which combine different genetic circuits, are useful. In this study, we designed a novel Bacillus subtilis expression toolbox, which allows the overproduction and secretion of potentially toxic enzymes. This toolbox comprises a set of 60 expression vectors, which combine two promoter variants, four strong secretion signals, a translation-enhancing downstream box, and three plasmid backbones. This B. subtilis toolbox is based on a tailor-made, clean deletion mutant strain, which is protease and sporulation deficient and exhibits reduced autolysis and secondary metabolism. The appropriateness of this alternative expression platform was tested for the overproduction of two difficult-to-produce eukaryotic model proteins. These included the sulfhydryl oxidase Sox from Saccharomyces cerevisiae, which forms reactive hydrogen peroxide and undesired cross-linking of functional proteins, and the human interleukin-1β, a pro-inflammatory cytokine. For the best performing Sox and interleukin, overproducing and secreting variants of these new B. subtilis toolbox fermentation strategies were developed and tested. This study demonstrates the suitability of the prokaryotic B. subtilis host-vector system for the extracellular production of two eukaryotic proteins with biotechnological relevance.
Author: | Anna Krüger, Norma Welsch, Alexandra Dürwald, Henrike Brundiek, Rainer Wardenga, Henning Piaschek, Hendrik G. Mengers, Jana Krabbe, Sandra Beyer, Johannes F. Kabisch, Lutz Popper, Tanno Hübel, Garabed Antranikian, Thomas Schweder |
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URN: | urn:nbn:de:gbv:9-opus-107812 |
DOI: | https://doi.org/10.1007/s00253-022-12062-2 |
ISSN: | 1432-0614 |
Parent Title (English): | Applied Microbiology and Biotechnology |
Publisher: | Springer Nature |
Place of publication: | Berlin |
Document Type: | Article |
Language: | English |
Date of Publication (online): | 2022/07/08 |
Date of first Publication: | 2022/08/01 |
Release Date: | 2024/02/29 |
Tag: | Bacillus subtilis; Expression system; Fed batch; Human interleukin-1β; Overproduction; Protease deficiency; Protein secretion; Yeast sulfhydryl oxidase |
Volume: | 106 |
Issue: | 13-16 |
First Page: | 5137 |
Last Page: | 5151 |
Faculties: | Mathematisch-Naturwissenschaftliche Fakultät / Institut für Pharmazie |
Collections: | weitere DFG-förderfähige Artikel |
Licence (German): | Creative Commons - Namensnennung 4.0 International |