1. Production of a polar fish antimicrobial peptide in Escherichia coli using an ELP-intein tag.
- Author
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Sousa DA, Mulder KCL, Nobre KS, Parachin NS, and Franco OL
- Subjects
- Anti-Bacterial Agents chemistry, Anti-Bacterial Agents economics, Anti-Bacterial Agents isolation & purification, Escherichia coli genetics, Escherichia coli metabolism, Fermentation, Genome, Bacterial, Peptides chemistry, Peptides economics, Peptides genetics, Peptides isolation & purification, Recombinant Fusion Proteins biosynthesis, Anti-Bacterial Agents biosynthesis, Elastin genetics, Inteins, Peptide Biosynthesis
- Abstract
An important aspect related to infectious pathogens is their exceptional adaptability in developing resistance, which leads to a perpetual challenge in the discovery of antimicrobial drugs with novel mechanisms of action. Among them, antimicrobial peptides (AMPs) stand out as promising anti-infective molecules. In order to overcome the high costs associated with isolation from natural sources or chemical synthesis of AMPs we propose the expression of Pa-MAP 2, a polyalanine AMP. Pa-MAP 2 was fused to an ELP-intein tag where the ELP (Elastin-like polypeptide) was used to promote aggregation and fast and cost-effective isolation after expression, and the intein was used to stimulate a controlled AMP release. For these, the vector pET21a was used to produce Pa-MAP 2 fused to the N-termini region of a modified Mxe GyrA intein followed by 60 repetitions of ELP. Purified Pa-MAP 2 showed a MIC of 25μM against E. coli ATCC 8739. Batch fermentation demonstrated that Pa-MAP-2 can be produced in both rich and defined media at yields 50-fold higher than reported for other AMPs produced by the ELP-intein system, and in comparable yields to expression systems with protease or chemical cleavage., (Copyright © 2016 Elsevier B.V. All rights reserved.)
- Published
- 2016
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