W0104

Position and Orientation of an Amphipathic (-Helix in Oriented Lipid Bilayers Determined Using a Novel X-ray Method. Stephen H. White and Kalina Hristova, Department of Physiology and Biophysics, University of California, Irvine, CA 92697-4560

We present a novel method for determining the position and orientation of proteins and peptides in oriented lipid bilayers by x-ray diffraction that is based upon determination of one-dimensional transbilayer scattering-density profiles on an absolute scale, molecular modeling, and oriented circular dichroism measurements. We demonstrate the method by using it to determine the transbilayer position and orientation of the class A amphipathic (-helical peptide¹ Ac-18A-NH₂ (Ac-DWLKAFYDKVAEKLKEAF-NH₂) in dioleoylphosphatidylcholine (DOPC) bilayers at 66% RH. The membrane scattering-density profiles of bilayers with and without Ac-18A-NH₂ were determined on an absolute scale using difference methods which also yield the transbilayer distribution of the bromine-labeled double-bond of the DOPC sn-2 chain² used in bilayer modeling. An accurate and robust model for the structure of the peptide/bilayer complex was obtained by modeling (1) the peptide-induced bilayer structural changes using the fully resolved structure of the DOPC bilayers at 66% RH³ as a starting point and (2) the structure of thermally disordered (-helical Ac-18A-NH₂ using molecular dynamics simulations. The fitting of the models to the experimental data shows that the Ac-18A-NH₂ helix axis is parallel to the bilayer surface and located at 16.5 ± 0.2 Å from the bilayer center, close to the glycerol moiety.

Supported by NIH grant GM-46823. ¹G.M. Anantharamaiah et al., J. Biol. Chem. 260:10248 (1985). ²M.C. Wiener and S.H. White, Biochemistry 30:6997 (1991). ³M.C. Wiener and S.H. White, Biophys. J. 61:434 (1992).