INSERTION OF MAGAININ INTO THE LIPID BILAYER DETECTED USING LIPID PHOTOLABELS

We investigated the interaction! of the antimicrobial peptides Ala(19) -magainin 2 amide and magainin 2 amide with lipid using two lipid phot olabels, azidobenzoyl galactosylceramide (GalCer-PL) and azidobenzoyla mido capryloyl galactosylceramide (GalCer-C8-PL), which position their photosensitive groups near the apolar-polar interface and near the ce nter of the bilayer, respectively. Magainins have been postulated to p ermeabilize membranes either by inserting in a transmembrane fashion i nto the bilayer and forming a channel or by binding to the surface of the bilayer and disturbing lipid packing. Evidence for channel formati on has been difficult to obtain, possibly because only a fraction of t he peptide may form a channel at any one time and because the channels may have a short lifetime. Both photolabels significantly labeled the peptides when bound to acidic phospholipid vesicles. The extent of la beling by GalCer-C8-PL was at least 70% of that by GalCer-PL, indicati ng that some of the peptide was inserted deeply into the bilayer at le ast transiently. The extent of labeling of Ala(19)-magainin 2 amide in creased significantly with an increase in the peptide to lipid mole ra tio, indicating cooperativity and supporting the channel model. The ex tent of labeling of this peptide was maximal by 30 a and did not chang e over 30 min, indicating that peptide insertion is rapid and either t hat the peptide remains inserted for at least 30 min or that equilibri um between inserted and noninserted peptide is achieved by 30 s. The l atter is supported by other studies in the literature. Use of this hyd rophobic photolabeling technique has permitted detection of peptide mo nomers which inserted into the bilayer and/or formed a channel at some time during the labeling procedure.