Cholesterol attenuates the interaction of the antimicrobial peptide gramicidin S with phospholipid bilayer membranes

We have investigated the effect of the presence of 25 mol percent cholester ol on the interactions of the antimicrobial peptide gramicidin S (GS) with phosphatidylcholine and phosphatidylethanolamine model membrane systems usi ng a variety of methods. Our circular dichroism spectroscopic measurements indicate that the incorporation of cholesterol into egg phosphatidylcholine vesicles has no significant effect on the conformation of the GS molecule but that this peptide resides in a range of intermediate polarity as compar ed to aqueous solution or an organic solvent. Our Fourier transform infrare d spectroscopic measurements confirm these findings and demonstrate that in both cholesterol-containing and cholesterol-free dimyristoylphosphatidylch oline liquid-crystalline bilayers, GS is located in a region of intermediat e polarity at the polar-nonpolar interfacial region of the lipid bilayer. H owever, GS appears to be located in a more polar environment nearer the bil ayer surface when cholesterol is present. Our P-31-nuclear magnetic resonan ce studies demonstrate that the presence of cholesterol markedly reduces th e tendency of GS to induce the formation of inverted nonlamellar phases in model membranes composed of an unsaturated phosphatidylethanolamine. Finall y. fluorescence dye leakage experiments indicate that cholesterol inhibits the GS-induced permeabilization of phosphatidylcholine vesicles. Thus in al l respects the presence of cholesterol attenuates but does not abolish the interactions of GS with, and the characteristic effects of GS on, phospholi pid bilayers. These findings may explain why it is more potent at disruptin g cholesterol-free bacterial than cholesterol-containing eukaryotic membran es while nevertheless disrupting the integrity of the latter at higher pept ide concentrations. This additional example of the lipid specificity of GS may aid in the rational design of GS analogs with increased antibacterial b ut reduced hemolytic activities. (C) 2001 Elsevier Science B.V. All rights reserved.