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Isolation, design, and biological properties of fish-derived cationic antimicrobial peptides

University of British Columbia

Cationic antimicrobial peptides are components of the host innate immune response against infections in almost all species. Although innate defenses in fish are particularly important due to a frequently late onset and short duration of the secondary immune response in these animals, few fish peptides have been studied to date. This research identifies a novel Hl-histone derived cationic peptide (HSDF-1) from coho salmon (Oncorhynchus kisutch), and describes the design, as well as the antibacterial, membrane, and intracellular activities of several other fish-based synthetic peptides of potential importance in aquaculture. HSDF-1, a 26 residue species identical to the N-terminal segment of trout HI histone, was isolated from the serum and mucus of disease-challenged coho salmon. The purified fractions inhibited the growth of antibiotic-supersusceptible S.typhimurium, as well as A. salmonicida and V. anguillarum. Synthetic HSDF-1 and its derivative HSDF-2 were inactive in antimicrobial assays, but they potentiated the antimicrobial activities of the flounder peptide pleurocidin, lysozyme, and crude lysozyme-containing extracts from coho salmon. Although the peptides inserted specifically into anionic lipid monolayers, synergy with pleurocidin did not appear to occur at the cytoplasmic membrane level. The effects of pleurocidin and several of its derivatives on bacterial membranes and intracellular functions were further studied to gain insight into the mode of action of these fish-derived peptides. All peptides tested at their minimal inhibitory concentrations (MICs) inhibited macromolecular synthesis in bacteria, causing a decrease in the incorporation of [3H]thymidine, [3H]uridine, or [3H]histidine into DNA, RNA, and proteins, respectively. However, many of the peptides applied at their MICs showed limited or negligible ability to damage bacterial cytoplasmic membranes, as observed using a fluorescent dye (diSC35) assay. The inhibition of macromolecular synthesis may thus be a result of the peptides entering bacterial cells and acting intracellularly in a direct fashion, as suggested by the fact that when two peptides were tested, they were able to translocate into liposomes without lysing them. A combined approach of isolating antimicrobial peptides from nature and designing them from existing templates, should eventually provide the medical and veterinary clinicians with a useful arsenal of weapons in the face of developing resistance to conventional antibiotics.

Thesis/Dissertation

application/pdf

2009-09-25

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http://hdl.handle.net/2429/13214

Microbiology and Immunology

University of British Columbia

UBCV

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