Antibacterial activities and conformations of synthetic α-defensin HNP-1 and analogs with one, two and three disulfide bridges

Abstract

Structure and biological activities of synthetic peptides corresponding to human α-defensin HNP-1, AC¹YC²RIPAC³IAGERRYGTC⁴IYQGRLWAFC⁵C⁶ with the S-S connectivities: C¹-C⁶, C²-C⁴, C³-C⁵, and its variants with one, two and three disulfide bridges were investigated. Oxidation of synthetic, reduced HNP-1 yielded a peptide with S-S connectivities C¹-C³, C²-C⁴ and C⁵-C⁶, and not with the S-S linkages as in naturally occurring HNP-1. Selective protection of cysteine sulfhydryls was necessary for the formation of S-S bridges as in native HNP-1. Likewise, oxidation of peptide encompassing the segment from C² to C⁵, resulted in the S-S linkages C²-C³ and C⁴-C⁵ instead of the expected linkage C²-C⁴ and C³-C⁵. Antibacterial activities were observed for all peptides, irrespective of how the S-S bridges were linked. Linear peptides without S-S bridges were inactive. Circular dichroism (CD) spectra suggest that peptides constrained by one and two S-S bridges do not form rigid β-sheet structures in an aqueous environment. The spectrum of HNP-1 in an aqueous environment suggests the presence of a β-hairpin conformation. In the presence of lipid vesicles, the S-S constrained peptides tend to adopt a β-structure. Although the S-S connectivities observed in HNP-1 may be necessary for other physiological activities, such as chemotaxis, they are clearly not essential for antibacterial activity.