By organizing only a portion of its plasma membrane surface a cell can concentrate proteins into smaller areas where they are more likely to associate. To investigate the formation of short range order arising from lipid-lipid and protein-protein associations on the bilayer surface, techniques having a lateral resolution on the length scale of 100 nm are necessary. This proposal outlines a research plan designed to probe two fundamental aspects of membrane dynamics: the mechanism by which lipids associate with other lipids to form regions of short range order (Aim 1) and the further reorganization of these small ordered regions into functional centers composed of multiple proteins (Aim 2). These vital phenomena can be quantitatively investigated within model fluid lipid bilayers patterned on solid supports using secondary-ion mass spectrometry (SIMS) implemented on the NanoSIMS 50. Supported bilayers capture the lateral fluidity of true cellular membranes in a versatile, controlled system. SIMS on the NanoSIMS 50 instrument will be used to image lipid and protein lateral associations at a spatial resolution of 100 nm (in flash-frozen samples), thereby quantitatively identifying the precise composition and organization of functionally relevant, protein-enriched areas on the bilayer. ? ? ?
