Decoding the Membrane Interactions of Antimicrobial Peptoids via Quartz Crystal Microbalance Nano-Viscometry

The rise in antimicrobial resistance underscores the urgent need for novel therapeutics. Membrane-disrupting agents offer a possible solution. Natural antimicrobial peptides are membrane-active but suffer from proteolytic instability and poor bioavailability. Sequence-defined peptoids have emerged as promising antimicrobial peptide mimetics, offering enhanced stability, tunable side-chain chemistry, and potent activity against drug-resistant pathogens. Here, we employed quartz crystal microbalance (QCM)-based viscoelastic fingerprinting, complemented by dye leakage assays, to characterize the membrane-disrupting behavior of three representative peptoids (B26, B14, and B43) across defined lipid bilayer models. Dye release assays confirmed potent membrane permeabilizing activity for all three peptoids, only attenuated by the presence of cholesterol in the membrane. QCM fingerprinting further revealed distinct lipid-composition-dependent interaction profiles that closely aligned with previously reported antimicrobial susceptibility data and predicted membrane-insertion states. The most potent analogue, B26, exhibited rapid, multistage destabilization of saturated bilayers, consistent with deep membrane insertion and pore-forming disruption. In contrast, B14, which features a redistributed charged domain, remained largely confined to the membrane surface, facilitating progressive membrane modulation without complete bilayer collapse. B43 displayed an intermediate activity profile, with reduced membrane disruption, likely attributable to weaker hydrophobic anchoring. Across all peptoids, lipid-specific signatures were observed, characterized by distinct interaction kinetics and outcomes. Together, these findings provide direct biophysical validation of peptoid structure-mechanism relationships and demonstrate the potential of peptoids as next-generation membrane-active therapeutics.