Orientation and Peptide−Lipid Interactions of Alamethicin Incorporated in Phospholipid Membranes: Polarized Infrared and Spin-Label EPR Spectroscopy

摘要

Alamethicin is a 20-residue peptaibiotic that induces voltage-dependent ion channels in lipidnmembranes. The mode by which alamethicin inserts into membranes was investigated using measurementsnof peptide-lipid interactions by spin-label electron paramagnetic resonance (EPR) and of peptide orientationnby polarized infrared (IR) spectroscopy. In fluid membranes, spin-labeled stearic acid shows no evidencenof a specific motionally restricted population of lipid chains, such as that found at the intramembranousnsurface of integral membrane proteins or oligomeric assemblies of transmembrane R-helices. In agreementnwith recent results from TOAC-substituted alamethicin analogues, native alamethicin is predominantlynmonomeric in fluid lipid membranes and presents an intramembrane surface that integrates well with thenlipid chains but is insufficiently extensive to induce specific motional restriction. Channel formation takesnplace by transient association of transmembrane monomers. In aligned fluid membranes, alamethicin exhibitsna large tilt in short chain-length lipids that decreases first rapidly with increasing chain-length and thennmore gradually for the lipids with longer chains. This macroscopically low order contrasts with the highnlocal order, relative to the local membrane normal, that is found by EPR for alamethicins spin-labelednwith TOAC. The macroscopic behavior is consistent with predictions for the chain-length dependence ofnelastic bending fluctuations of the membrane surface, which was invoked recently to explain the spontaneousninsertion of u0001-barrel proteins in short-chain lipid membranes.