Computational engineering of the scorpion venom peptide Ttap3 enhances membrane selectivity and antimicrobial potential.

ABSTRACT: Venom-derived antimicrobial peptides (AMPs) are promising scaffolds for next-generation antimicrobial agents because of their structural diversity and membrane-targeting mechanisms. Ttap3, a peptide isolated from scorpion venom, exhibits weak antimicrobial activity despite possessing characteristic amphipathic features associated with membrane-active peptides. This study employed a computational engineering strategy to redesign Ttap3 and investigate how sequence modifications influence membrane selectivity and antimicrobial potential. Structural modelling showed that Ttap3 adopted a predominantly α-helical amphipathic conformation with moderate hydrophobic moment and relatively high hydrophobicity. Molecular dynamics simulations demonstrated preferential interaction with bacterial membrane models relative to mammalian membranes, although measurable interaction with mammalian bilayers was also observed. Rational sequence modifications involving residue substitution, sequence reversal, and modulation of amphipathicity and flexibility generated analogues with improved predicted selectivity profiles. Multi-scale molecular dynamics simulations revealed distinct membrane interaction behaviours among the redesigned analogues, including surface-associated destabilization, localized pore formation, and insertion-driven membrane perturbation. Coordinated membrane disruption involving hydrogen bond loss, lipid disorder, phospholipid deflection, altered pore radii, and increased water influx was observed predominantly in bacterial membrane systems. Partial reduction in peptide helicity during membrane interaction also appeared to contribute to selective membrane destabilization. These findings demonstrate that rational computational redesign can substantially improve the predicted antimicrobial potential and membrane selectivity of weakly active venom-derived peptides. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at https://doi.org/10.1007/s40203-026-00742-0.