Antimicrobial Peptide C18G binds to Lipid Bilayers in a Lipid Composition Dependent Manner

Abstract

Antimicrobial peptides serve as one of the first lines of defense in the immune systems of higher organisms. These peptides specifically target and neutralize infecting bacteria in the host organism while exhibiting little or no toxic effect on host cells. The peptide C18G is a highly cationic, amphiphilic peptide derived from the C-terminal sequence of the human protein platelet factor 4 (involved in blood coagulation and wound repair) exhibited antibacterial activity against both gram positive and gram negative bacteria. Using a modified C18G sequence that did not affect antimicrobial efficacy (Y3 changed to W), the binding affinity of the peptide to model membranes was performed using fluorescence spectroscopy. As anticipated, the binding of C18G to lipid bilayers allowed the Trp side chain to localize to a more hydrophobic environment resulting in a blue shift of Trp emission lambda max and spectral barycenter, concomitant with a narrowing of the emission spectrum and and increase in the overall emission intensity. Binding to lipid vesicles composed of binary and tertiary mixtures of POPC:POPG and POPC:POPG:POPE showed a dramatic lipid dependence on binding affinity, with the tightest binding to the most anionic compositions. Increasing the POPE composition enhanced peptide binding but to a lesser degree compared to the anionic POPG. Fluorescence quenching experiments using the aqueous quencher acrylamide confirmed the decreased exposure of the Trp to the aqueous milieu. Dye release assays were used to monitor lipid composition effects on the ability of C18G to permeabilize lipid vesicles. Circular dichroism spectroscopy indicated a conformational change from a disordered to an alpha-helical secondary structure when the peptide interacts with detergent micelles or anionic lipid vesicles.