title: Retrocyclins neutralize bacterial toxins by potentiating their unfolding.
abstract: Defensins are a class of immune peptides with a broad range of activities against bacterial, fungal and viral pathogens. Besides exerting direct anti-microbial activity via dis-organization of bacterial membranes, defensins are also able to neutralize various unrelated bacterial toxins. Recently, we have demonstrated that in the case of human α- and β-defensins, this later ability is achieved through exploiting toxins' marginal thermodynamic stability, i.e. defensins act as molecular anti-chaperones unfolding toxin molecules and exposing their hydrophobic regions and thus promoting toxin precipitation and inactivation [Kudryashova et al. (2014) Immunity 41, 709-721]. Retrocyclins (RCs) are humanized synthetic θ-defensin peptides that possess unique cyclic structure, differentiating them from α- and β-defensins. Importantly, RCs are more potent against some bacterial and viral pathogens and more stable than their linear counterparts. However, the mechanism of bacterial toxin inactivation by RCs is not known. In the present study, we demonstrate that RCs facilitate unfolding of bacterial toxins. Using differential scanning fluorimetry (DSF), limited proteolysis and collisional quenching of internal tryptophan fluorescence, we show that hydrophobic regions of toxins normally buried in the molecule interior become more exposed to solvents and accessible to proteolytic cleavage in the presence of RCs. The RC-induced unfolding of toxins led to their precipitation and abrogated activity. Toxin inactivation by RCs was strongly diminished under reducing conditions, but preserved at physiological salt and serum concentrations. Therefore, despite significant structural diversity, α-, β- and θ-defensins employ similar mechanisms of toxin inactivation, which may be shared by anti-microbial peptides from other families.
authors: *Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, U.S.A.
authors: Elena
authors: E
authors: Kudryashova
authors: †Department of Microbiology, The Ohio State University, Columbus, OH 43210, U.S.A.
authors: Stephanie
authors: S
authors: Seveau
authors: §Institute of Human Virology and Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, Baltimore, MD 21201, U.S.A.
authors: Wuyuan
authors: W
authors: Lu
authors: *Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, U.S.A.
authors: Dmitri S
authors: DS
authors: Kudryashov
mesh_terms: D001427:Bacterial Toxins / Q000737:chemistry* / Q000378:metabolism; D002460:Cell Line; D006801:Humans; D058767:Protein Unfolding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D023084:alpha-Defensins / Q000737:chemistry* / Q000378:metabolism; D023083:beta-Defensins / Q000737:chemistry* / Q000235:genetics
chemical_list: D001427:Bacterial Toxins; D011993:Recombinant Fusion Proteins; D023084:alpha-Defensins; D023083:beta-Defensins