Membranolytic selectivity of cystine-stabilized cyclic protegrins

To correlate conformational rigidity with membranolytic selectivity of anti microbial activity and cytotoxicity, we prepared six cyclic analogs of prot egrin-1 (PG-1), an 18-residue cationic peptide with a broad-spectrum antimi crobial activity. These cyclic protegrins bear end-to-end peptide bonds tog ether with varying numbers (zero to three) of cross-strand disulfide constr aints. The most constrained analog is a cyclic tricystine protegrin (ccPG 3 ) containing three evenly spaced, parallel disulfide bonds. Antimicrobial a ssays against 10 organisms in low- and high-salt conditions showed that the se cyclic protegrins were broadly active with different antimicrobial profi les against Gram-positive and Gram-negative bacteria, fungi and one tested virus, HIV-1. Compared to PG-1, the cyclic tricystine ccPG 3 displayed appr oximately a 10-fold decrease in hemolytic activity against human cells and 6- to 30-fold improvement of membranolytic selectivity against six of the 1 0 tested organisms. In contrast, [Delta SS]cPG 8, a cyclic protegrin with n o disulfide bond, and [Delta Cys6,15]cPG 5, a cyclic mimic of PG-1 with one disulfide bond, exhibited activity spectra, potency, and cytotoxicity simi lar to PG-1. Circular dichroism showed that cyclic protegrins containing wi th one to three cystine bonds displayed some degree of beta-strand structur es in water/trifluoroethanol or phosphate-buffered solutions. Collectively, our results indicate that cyclic structures are useful in the design of an timicrobial peptides and that an increase in the conformational rigidity of protegrins may confer membranolytic selectivity that dissociates antimicro bial activity from hemolytic activity.