Nano-viscosimetry analysis of membrane disrupting peptide magainin2 interactions with model membranes

The rapid spread of antibiotic-resistant strains of bacteria has created an urgent need for new alternative antibiotic agents. Membrane disrupting antimicrobial peptides (AMPs): short amino acid sequences with bactericidal and fungicidal activity that kill pathogens by permeabilizing their plasma membrane may offer a solution for this global health crisis. Magainin 2 is an AMP secreted by the African clawed frog (Xenopus laevis) that is described as a toroidal pore former membrane disrupting AMP. Magainin 2 is one of the most thoroughly studied AMPs, yet its mechanism of action is still largely hypothetical: visual evidence of the pore formation is lacking, and the molecular mechanism leading to pore formation is still debated. In the present study, quartz crystal microbalance (QCM) based viscoelastic fingerprinting analysis supported by dye leakage experiments and atomic force microscopy (AFM) imaging was used to glean deeper insights into the mechanism of action. The effect of membrane charge, acyl chain unsaturation and cholesterol concentration were also investigated. The results show lipid specific disruptive mechanism of magainin 2. QCM nano-viscometry measurements revealed the presence of distinct stages in the mechanism of magainin 2 action that, with dye leakage data, confirm the existence of an initial transient pore stage that may result in peptide flip-flop between the outer and inner membrane leaflets. There is evidence of a further mechanistic stage at high peptide concentrations that is consistent with membrane collapse into a peptide-lipid mixed phase that is distinct from the transient pore formation. The results confirm some of the earliest hypotheses about magainin 2 action, while also highlighting the membrane modulating effect of this peptide.