Peptide–liposome association. A critical examination with mastoparan-X

Abstract

Mastoparan-X, a wasp venom factor, is a membrane active peptide whose binding to lipid vesicles is of basic interest towards an analysis of its functions. Titration of aqueous peptide solutions with liposomes allows the determination of the association isotherm, i.e. a plot of bound peptide per lipid versus the free peptide concentration. We have scrutinised the various steps in the evaluation procedure, considering circular dichroism as well as fluorescence intensity as possible signals for the binding process. First of all the measured data had to be corrected for light scattering effects which may otherwise appreciably falsify the final results. Uncertainties due to inherent difficulties regarding the reproducibility of lipid preparations and inevitable titration errors have to be considered. The consequences of these errors for the quantitative analysis of the titration curves were investigated. The plotted curves can be reasonably well fitted by a functional relationship derived from a Gouy–Chapman model approach that assumes a partitioning of monomeric peptide. The two relevant parameters, partition coefficient and effective charge number, and their error ranges have been determined for mastoparan-X and a series of phosphatidylcholine vesicle sizes and various ionic strengths. These findings show that the applied analysis implies a sufficient basis for calculations of the amount of lipid bound peptide in practice. However, the possible existence of peptide aggregates cannot generally be excluded from a formal monomer associated curve fit as indicated by computer simulations.