Structural Studies of Supported Bilayers as Cell Membrane Model Systems
The cell membrane is thought to have a much more active role in cellular function than previously known. As such, there is considerable interest in measuring the physical properties of model cell membranes to determine the biological relevance of these systems; specifically, the differences between the conventional model of a free bilayer in the form of a vesicle and a solid-supported lipid bilayer are of concern. Solid-supported lipid bilayers have been shown, using atomic force and fluorescence microscopy, to display different bulk properties than vesicles. The molecular information offered by nuclear magnetic resonance spectroscopy complements the bulk information provided by surface based techniques. We are developing NMR as a tool to probe the molecular aspects of model cell membrane structure.
A reliable method for creating supported single lipid bilayers on spherical supports has been developed by our lab. The resultant spherically supported bilayers (SSBs) have been studied using 2H NMR spectroscopy to elucidate differences in structure by extracting the order parameter profile of the alkyl chains from the 2H quadrupolar splitting, and by 1H NMR diffusion measurements, where the diffusion coefficient is used as a spectroscopic probe of molecular behavior.
Atomic force and fluorescence microscopy studies were also done on planar supported bilayers, as a confirmation of the bilayer properties as measured by the conventional means. Furthermore, a method was developed using AFM to indicate definitively whether or not a supported bilayer existed on the supporting mica surface.
Results indicate that there is a difference in bilayer structure between supported bilayers and the conventional model of a free bilayer. A method for extracting the alkyl chain order parameter profile is being developed, with promising initial results. Additionally, an NMR diffusion pulse sequence using the WATERGATE water suppression technique has been developed and employed on both free and supported lipid bilayers.
Advisor:David W. Pratt; Sunil K. Saxena; Megan M. Spence
School:University of Pittsburgh
School Location:USA - Pennsylvania
Source Type:Master's Thesis
Date of Publication:06/04/2008