Identification and Characterization of an In Silico Designed Membrane‐Active Peptide with Antiviral Properties

ABSTRACT Broad‐spectrum antivirals are urgently needed to counter emerging viral threats. Targeting the viral envelope, an essential, conserved, and host‐derived structure, offers a promising strategy with a low risk of resistance. Here, we report the in silico design and experimental characterization of P1.6, a 24‐mer peptide generated using an evolutionary molecular dynamics (Evo‐MD) platform and optimized to sense and exploit lipid packing defects in viral membranes. Among nine Evo‐MD–derived candidates, P1.6 showed the strongest membrane‐disruptive activity and inhibited HIV‐1, Zika virus, and herpes simplex viruses with IC 50 values ranging from ∼0.06 to 3.5 µ m . P1.6 efficiently disrupted virus‐like liposomes without causing cytotoxicity or hemolysis at antiviral concentrations. All‐atom MD simulations predicted a predominantly α‐helical solution structure with a central kink and flexible termini. Upon membrane engagement, this kink was largely lost, yielding a more continuous and stabilized helix. ATR‐FTIR spectroscopy confirmed the membrane‐induced increase in helicity. Coarse‐grained MD simulations further demonstrated that P1.6 stabilizes transient membrane pores, while electron microscopy of treated HIV‐1 particles revealed extensive envelope rupture and capsid release. Together, these results establish P1.6 as a potent membrane‐active antiviral lead and highlight the utility of Evo‐MD–guided peptide design to target conserved biophysical vulnerabilities in viral envelopes.