Engineered salt-insensitive alpha-defensins with end-to-end circularized structures

We designed a retro-isomer and seven circularized "beta-tile" peptide analo gs of a typical rabbit cu-defensin, NP-1. The analogs retained defensin-lik e architecture after the characteristic end-to-end, Cys(3,31) (C I:C VI), a lpha-defensin disulfide bond was replaced by a backbone peptide bond. The r etro-isomer of NP-1 was as active as the parent compound, suggesting that o verall topology and amphipathicity governed its antimicrobial activity. A b eta-tile design with or without a single cross-bracing disulfide bond suffi ced for antimicrobial activity, and some of the analogs retained activity a gainst Escherichia coli and Salmonella typhimurium in NaCl concentrations t hat rendered NP-I inactive. The new molecules had clustered positive charge s resembling those in protegrins and tachyplesins, but were less cytotoxic. Such simplified alpha-defensin analogs minimize problems encountered durin g the oxidative folding of three-disulfide defensins, In addition, they are readily accessible to a novel thia zip cyclization procedure applicable to large unprotected peptide precursors of 31 amino acids in aqueous solution s. Collectively, these findings provide new and improved methodology to cre ate salt-insensitive defensin-like peptides for application against bacteri al diseases.