Self-assembled nanonetworks of highly stable gemini surfactant-like peptides: antibacterial mechanisms, self-assembly characteristics, and in vivo anti-infection potential.

The escalating prevalence of bacterial resistance underscores the critical demand for developing non-antibiotic therapeutic candidates. Antimicrobial peptides (AMPs) with potent efficacy and low toxicity have emerged as a promising alternative strategy; however, their suboptimal proteolytic stability and bioavailability remain major obstacles to clinical application. Designing self-assembled nanosystems that coordinate the rational arrangement of amino acids to enhance the protease degradation resistance of AMPs provides a breakthrough solution for overcoming their stability bottlenecks. Drawing inspiration from the structure and self-assembly properties of gemini surfactants, we have developed a series of structural templates for gemini surfactant-like peptides that self-assemble through intermolecular noncovalent forces. Among these peptides, IPr exhibits potent antibacterial activity against all ten tested strains (including Gram-negative and Gram-positive bacteria), while demonstrating remarkable protease resistance and tolerance to physiological salt ions. Integrating molecular dynamics simulations with structural characterization, we confirm that IPr self-assembles into short nanoribbons and cross-links into nanonetworks exclusively through noncovalent interactions (including π-π stacking, hydrophobic interactions, and hydrogen bonding). Mechanism studies reveal that IPr exerts its effects predominantly through membrane disruption, which triggers a cascade of cellular events, including reactive oxygen species (ROS) accumulation and ATP leakage, thereby achieving multidimensional synergistic bactericidal effects. IPr exhibits excellent biocompatibility in vivo and significantly reduces the severity of systemic bacterial infection in a mouse peritonitis model. This design paradigm of self-assembled gemini surfactant-like peptides offers a viable strategy for developing highly stable peptide-based biomaterials.