Unraveling the antifungal activity of a South American rattlesnake toxin crotamine

• Published in: Biochimie 2013-02, Vol.95 (2), p.231-240
• Main Authors: Yamane, Erica S., Bizerra, Fernando C., Oliveira, Eduardo B., Moreira, Jéssica T., Rajabi, Mohsen, Nunes, Gabriel L.C., de Souza, Ana O., da Silva, Ismael D.C.G., Yamane, Tetsuo, Karpel, Richard L., Silva, Pedro I., Hayashi, Mirian A.F.
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Amphipathic peptide; Animals; Anti-Bacterial Agents - chemical synthesis; Anti-Bacterial Agents - isolation & purification; Anti-Bacterial Agents - pharmacology; Antifungal Agents - chemical synthesis; Antifungal Agents - isolation & purification; Antifungal Agents - pharmacology; Antimicrobial activity; Aspergillus fumigatus; beta-Defensins - chemistry; Candida albicans; Cell Line; Cell Survival - drug effects; Clinical strain; Crotalid Venoms - chemical synthesis; Crotalid Venoms - isolation & purification; Crotalid Venoms - pharmacology; Crotalus - physiology; Crotalus durissus terrificus; Cryptococcus neoformans; Dose-Response Relationship, Drug; Escherichia coli - genetics; Fungi - drug effects; Fungi - growth & development; Fungi - ultrastructure; Gram-Negative Bacteria - drug effects; Gram-Negative Bacteria - growth & development; Gram-Positive Bacteria - drug effects; Gram-Positive Bacteria - growth & development; Humans; Microbial Sensitivity Tests; Microbial Viability - drug effects; Micrococcus luteus; Molecular Sequence Data; Rattlesnake toxin; Recombinant Proteins - chemistry; Recombinant Proteins - genetics; Recombinant Proteins - pharmacology; Trichophyton rubrum; Trichosporon
• ISSN: 0300-9084; 1638-6183
• DOI: 10.1016/j.biochi.2012.09.019

Crotamine is a highly basic peptide from the venom of Crotalus durissus terrificus rattlesnake. Its common gene ancestry and structural similarity with the β-defensins, mainly due to an identical disulfide bond pattern, stimulated us to assess the antimicrobial properties of native, recombinant, and chemically synthesized crotamine. Antimicrobial activities against standard strains and clinical isolates were analyzed by the colorimetric microdilution method showing a weak antibacterial activity against both Gram-positive and Gram-negative bacteria [MIC (Minimum Inhibitory Concentration) of 50–>200 μg/mL], with the exception of Micrococcus luteus [MIC ranging from 1 to 2 μg/mL]. No detectable activity was observed for the filamentous fungus Aspergillus fumigatus and Trichophyton rubrum at concentrations up to 125 μg/mL. However, a pronounced antifungal activity against Candida spp., Trichosporon spp., and Cryptococcus neoformans [12.5–50.0 μg/mL] was observed. Chemically produced synthetic crotamine in general displayed MIC values similar to those observed for native crotamine, whereas recombinant crotamine was overridingly more potent in most assays. On the other hand, derived short linear peptides were not very effective apart from a few exceptions. Pronounced ultrastructure alteration in Candida albicans elicited by crotamine was observed by electron microscopy analyses. The peculiar specificity for highly proliferating cells was confirmed here showing potential low cytotoxic effect of crotamine against nontumoral mammal cell lines (HEK293, PC12, and primary culture astrocyte cells) compared to tumoral B16F10 cells, and no hemolytic activity was observed. Taken together these results suggest that, at low concentration, crotamine is a potentially valuable anti-yeast or candicidal agent, with low harmful effects on normal mammal cells, justifying further studies on its mechanisms of action aiming medical and industrial applications. [Display omitted] Transmission electron micrographs of (A) untreated C. albicans exhibiting a normal morphology, and (B) pronounced alterations of these cells after treatment with crotamine. (C) Kinetics of C. albicans killing by crotamine showing observed viable colony counts (CFU/mL). ► We analyzed the antimicrobial activities against standard and clinical isolates. ► Crotamine is a potentially valuable anti-yeast or candicidal agent. ► Crotamine has no hemolytic activity & low harmful effects on normal mammal cells.