Effect of aminoisobutyric acid (Aib) substitutions on the antimicrobial and cytolytic activities of the frog skin peptide, temporin-1DRa

Temporin-1DRa (HFLGTLVNLAKKIL.NH₂), first isolated from the skin of the California red-legged frog Rana draytonii, shows broad-spectrum antimicrobial activity but its therapeutic potential is limited by its toxicity against mammalian cells. The cytolytic properties of cationic α-helical peptides are determined by a complex interaction between cationicity, hydrophobicity, conformation, and amphipathicity. This study has investigated the cytolytic properties of conformationally constrained analogs of temporin-1DRa containing α-aminoisobutyric acid (Aib) substitutions. Cytolytic activity was determined against the bacteria Escherichia coli and Staphylococcus aureus, the opportunistic yeast pathogen, Candida albicans, human erythrocytes, HepG2 hepatoma-derived cells, and L929 fibroblasts. Aib substitutions at Gly⁴, Asn⁸, and Ala¹⁰ increased both % helicity, determined in methanol solution, and hydrophobicity resulting in increases in both antimicrobial potencies and toxicities against the mammalian cells. Substitution at Leu⁶ resulted in an appreciable decrease in cytolytic activity against all cells whereas the substitutions at His¹, Phe², Leu³, Thr⁵, and Val⁷ had only minor effects on activity. Substitutions at Leu⁹, Ile¹³, Leu¹⁴ produced analogs with decreased helicity and hydrophobicity that retained activity against microorganisms but showed appreciably lower cytolytic activities against mammalian cells. In particular, the fourfold increase in therapeutic index [ratio of LC₅₀ against erythrocytes to minimum inhibitory concentration (MIC) against microorganisms] of [Aib¹³]temporin-1DRa identifies it as a compound with potential for development as a therapeutically valuable anti-infective agent.