Structural and functional analysis of horse cathelicidin peptides

Cathelicidin-derived antimicrobial peptides are a component of the peptide- based host defense of neutrophils and epithelia, with a widespread distribu tion in mammals. We recently reported the cDNA sequences of three putative horse myeloid cathelicidins, named eCATH-1, -2, and -3, A Western analysis was performed to investigate their presence in neutrophils and processing t o mature peptides, eCATH-2 and eCATH-3, but not eCATH-1, were found to be p resent in uncleaved forms in horse neutrophils, The corresponding mature pe ptides were detected in inflammatory sites, suggesting that processing of t he propeptides takes place upon neutrophil activation. A functional charact erization was then performed with synthetic eCATH peptides, Circular dichro ism measurements indicated an amphipathic alpha -helical conformation of th ese peptides in an anisotropic environment, and in vitro assays revealed a potent activity and a broad spectrum of antimicrobial activity for eCATH-1 and a somewhat more restricted spectrum of activity for eCATH-2, Conversely , a strong dependence on salt concentration was observed when the activity of eCATH-3 was tested. This peptide efficiently killed bacteria and some fu ngal species, i.e., Cryptococcus neoformans and Rhodotorula rubra, in low-i onic-strength media, but the activity was inhibited in the presence of phys iological salt medium. This behavior could be modified by modulating the am phipathicity of the molecule, In fact, the synthetic analogue LLK-eCATH-3, with a slightly modified sequence that increases the hydrophobic moment of the peptide, displayed a potent activity in physiological salt medium again st the strains resistant to eCATH-3 under these conditions.