Membrane Perturbing Effects of Antimicrobial Peptides: A Systematic Spectroscopic Analysis

Antimicrobial peptides (AMPs) exhibit a strong activity against a wide range of microorganisms, mainly by perturbing the permeability of bacterial membranes through the formation of pores. However, AMPs effects on membrane properties probably extend beyond pore-formation. We performed a systematic spectroscopic analysis of the effects on membrane structure and dynamics of two very different AMPs: the cationic PMAP-23, which creates pores according to the “carpet” model, and alamethicin, which forms “barrel-stave” channels. By using fluorescence anisotropy measurements on liposomes comprising probes localized at different depths in the bilayer, we measured peptide effects on membrane fluidity and order. Laurdan spectral shifts provided indications about water penetration in the bilayer. In the case of PMAP-23, it was possible to focus specifically on the lipids surrounding the peptide by following the membrane-probe fluorescence due to FRET from the peptide Trp residues. Finally, peptide-induced perturbation of lateral mobility and domain formation were determined by several methods. All experiments were compared with liposome-leakage measurements: while for PMAP-23 all membrane-perturbing effects are correlated with the vesicle leakage process, alamethicin does not significantly influence membrane dynamics at the concentrations in which it forms pores. Surprisingly, in all cases the most significant peptide-induced effect is a reduction in membrane fluidity.