Abstract
In recent years, antimicrobial peptides (AMPs) have attracted increasing attention. The microbial cells provide a simple, cost-effective platform to produce AMPs in industrial quantities. While AMP production as fusion proteins in microorganisms is commonly used, the recovery of AMPs necessitates the use of expensive proteases and extra purification steps. Here, we develop a novel fusion protein DAMP4-F-pexiganan comprising a carrier protein DAMP4 linked to the AMP, pexiganan, through a long, flexible linker. We show that this fusion protein can be purified using a non-chromatography approach and exhibits the same antimicrobial activity as the chemically synthesized pexiganan peptide without any cleavage step. Activity of the fusion protein is dependent on a long, flexible linker between the AMP and carrier domains, as well as on the expression conditions of the fusion protein, with low-temperature expression promoting better folding of the AMP domain. The production of DAMP4-F-pexiganan circumvents the time-consuming and costly steps of chromatography-based purification and enzymatic cleavages, therefore shows considerable advantages over traditional microbial production of AMPs. We expect this novel fusion protein, and the studies on the effect of linker and expression conditions on its antimicrobial activity, will broaden the rational design and production of antimicrobial products based on AMPs.
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Acknowledgments
This research was supported by the Australian Research Council (ARC) under Discovery Project (DP150100798). C.-X Zhao acknowledges financial support from the award of the Australian Research Council (ARC) Future Fellowship (FT140100726). We acknowledge the facilities of Australian Genome Research Facility, The University of Queensland for gene sequencing. This work was performed in part at the Queensland node of the Australian National Fabrication Facility (ANFF-Q), a company established under the National Collaborative Research Infrastructure Strategy to provide nano and micro-fabrication facilities for Australia’s researchers.
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Sun, B., Wibowo, D., Sainsbury, F. et al. Design and production of a novel antimicrobial fusion protein in Escherichia coli. Appl Microbiol Biotechnol 102, 8763–8772 (2018). https://doi.org/10.1007/s00253-018-9319-4
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DOI: https://doi.org/10.1007/s00253-018-9319-4