Abstract
Cationic antimicrobial peptides are able to kill a broad variety of Gram-negative and Gram positive bacteria and thus are good candidates for a new generation of antibiotics to treat multidrug-resistant bacteria. Here we describe a high-throughput method to screen large numbers of peptides for improved antimicrobial activity. The method relies on peptide synthesis on a cellulose support and a Pseudomonas aeruginosa strain that constitutively expresses bacterial luciferase. A complete substitution library of 12-amino-acid peptides based on a linearized variant (RLARIVVIRVAR-NH2) of the bovine peptide bactenecin was screened and used to determine which substitutions at each position of the peptide chain improved activity. By combining the most favorable substitutions, we designed optimized 12-mer peptides showing broad spectrum activities with minimal inhibitory concentrations (MIC) as low as 0.5 μg/ml against Escherichia coli. Similarly, we generated an 8-mer substituted peptide that showed broad spectrum activity, with an MIC of 2 μg/ml, against E. coli and Staphylococcus aureus.
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Acknowledgements
R.E.W.H. holds a Canada Research Chair. We thank the Applied Food and Materials Network of Centers of Excellence for funding and Oreola Donini for her technical support in demonstrating how to determine the IC50 values.
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The peptide sequences and research results described in this paper are part of a US provisional patent application by the University of British Columbia.
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Hilpert, K., Volkmer-Engert, R., Walter, T. et al. High-throughput generation of small antibacterial peptides with improved activity. Nat Biotechnol 23, 1008–1012 (2005). https://doi.org/10.1038/nbt1113
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DOI: https://doi.org/10.1038/nbt1113
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