A Solvent-Assisted Supported Bilayer QCM-D Platform for Resolving Composition- and Structure-Dependent Antimicrobial Peptide-Membrane Interactions

Abstract

Antimicrobial peptides (AMPs) disrupt bacterial membranes through mechanisms that differ fundamentally from conventional antibiotics, yet the interfacial transition from peptide adsorption to membrane destabilization remains difficult to resolve experimentally. Here we use quartz crystal microbalance with dissipation monitoring (QCM-D) together with the solvent-assisted lipid bilayer method (SALB) to quantify concentration-dependent interactions of antimicrobial peptides, namely Cecropin A and Magainin 1, two highly studied AMPs, with supported membrane models in real time. Magainin 1 interactions with Gram-negative (E. coli lipid extract) membranes exhibit a clear transition from adsorption-dominated behavior at low concentrations (1–2 μM), characterized by modest frequency decreases and small dissipation increases, to mechanical perturbation at higher concentrations (≥5 μM). In contrast, cholesterol-containing mammalian membrane mimics display small frequency shifts with minimal dissipation changes, consistent with adsorption without mechanical perturbation. Experiments with the antimicrobial peptide Cecropin A show stronger perturbation of bacterial membranes, producing large frequency decreases and substantial dissipation increases across the entire concentration range examined, while remaining similarly suppressed on the mammalian mimic. Control experiments using triglycine confirm that nonspecific peptide adsorption does not substantially affect either membrane type. Comparison with a cholesterol-free mammalian membrane mimic showed that headgroup charge alone did not enhance AMP adsorption, but instead promoted fully reversible binding. These findings suggest that cholesterol is an important determinant of irreversible AMP intercalation into mammalian membranes. In this study, we quantify composition-dependent mechanical perturbation of bacterial membranes and establish QCM-D as a quantitative probe of potential cooperative membrane disruption by antimicrobial peptides and as a tool to study the mechanistic models of AMPs.