The proposed studies will define, at the molecular level, factors controlling protein-mediated glycolipid transfer between membrane surfaces. Specifically, these studies are aimed at understanding how composition, packing density, and interactions between lipid molecules regulate glycolipid transfer protein activity. Accomplishing such aims will require the use of model membrane systems that provide distinct, yet complimentary, information about glycosphingolipid organization in membranes and about glycolipid transfer protein interaction with membranes. Accordingly, the four specific aims are: (i) to ascertain the mixing properties of glycosphingolipids and phospholipids or glycosphingolipids and cholesterol in monolayers by measuring changes in surface pressure and surface potential as function of molecular area at different lipid compositions; (ii) to delineate surface structural changes occurring at different glycosphingolipid-phospholipid and glycosphingolipid-cholesterol compositions in bilayer membranes by employing freeze-fracture and freeze-etch electron microscopy; (iii) to determine the role of lipid surface structure in regulating glycolipid transfer protein interaction with monolayer surfaces using kinetic and binding approaches; and (iv) to define the structural features which regulate interactions between glycolipid transfer protein and membrane surfaces using fluorescence techniques. The knowledge gained from the proposed studies will be important not only for advancing our basic understanding of the regulation of lipid transfer proteins and the role of glycolipids transfer protein in the expression of glycosphingolipids, but will also provide information of biomedical relevance. Glycolipid transfer protein is potentially a valuable research tool for inserting and/or removing glycosphingolipids into biological and model membrane systems. Such a tool could be used to study certain disease states where normal glycosphingolipid composition is altered (e.g., transformed cells, Gaucher's and Krabbe's diseases). By manipulating the glycolipid composition of cells, new information about the functional involvement of altered glycolipids in the disease process may be revealed. Such studies will also provide pertinent information about surface structural parameters affecting the interaction of other proteins and macromolecules (i.e., antibodies, bacterial toxins, lectins, viruses) that bind to glycolipids in cell membranes surfaces.
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