Selective Membrane Remodeling and Fluidity Reduction by the Cationic Helical Peptide L₁₀-MMBen

Antimicrobial peptide (AMP)-based treatments have exhibited significant therapeutic potential with anti-inflammatory, anticancer, and immunomodulatory activities. While most AMPs exert effects by permeabilizing bacterial cell membranes through poration and hydrolysis, recent findings have revealed a distinct membrane interaction mechanism employed by L₁₀-MMBen, a cationic amphiphilic helical peptide with potent antimicrobial efficacy and moderate cytotoxicity. Herein, we elucidate the ability of L₁₀-MMBen to remodel bacterial cell membranes into double-layered structures rather than inducing permeabilization. Quantitative real-time giant unilamellar vesicle assays, atomic force microscopy characterizations, and simulations demonstrate that L₁₀-MMBen selectively adsorbs onto phosphatidylglycerol-containing membranes with shallow insertion at approximately 0.6 nm; once reaching a threshold local concentration (peptide-to-lipid ratio of 1:20), it extracts lipids from the bilayer and facilitates the formation of double-layered peptide-membrane composite structures. Single-molecule tracking analysis indicates that peptide-induced molecular reorganization significantly reduces membrane fluidity, impeding lipid lateral diffusion from 2.8 μm²/s to an immobile state. These findings may contribute to the design of innovative membrane-active agents for biomedical applications.