Enhanced Gram-Negative Membrane Disruption and In Vivo Efficacy via Lysine-Arginine Enrichment of Opis16a

Infections complicate burn wound care, especially with the rise of antimicrobial resistance. Antimicrobial peptides (AMPs) offer the potential for advancing wound care by combating persistent infections. Opis16a, a scorpion venom-derived AMP, exhibits potent antibacterial activity by targeting Gram-negative membranes, causing rapid membrane disruption and bacterial cell death. Here, four novel Opis16a analogues were developed with improved membrane targeting and antibacterial efficacy. One analogue shows particular promise for topical application in Gram-negative burn wound infections. Enhanced peptide-lipid hydrogen bonding increases conformational stability, membrane insertion, and permeabilization rates. Substituting lysine residues in the C-terminal with arginine leads to the most consistent improvement in activity, selectivity for pathogen over HaCat cells, and stability in serum. In an in vivo <i>Galleria mellonella</i> burn wound model, a 5 mg/kg topical dose provides better protection than Opis16a against <i>Enterobacter cloacae</i> NICD 16103. These findings highlight the potential of optimized bactericidal AMPs to improve burn wound care.