MOLECULAR MECHANISMS OF ANNEXIN/MEMBRANE INTERACTIONS

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The annexins are a group of homologous, calcium-dependent, membrane- binding proteins present in a wide variety of cells and tissues. Although normally soluble, in the presence of calcium these proteins bind to acidic lipids in membranes leading to lipid immobilization and sequestration. Most members of the family will also promote the calcium-dependent aggregation and fusion of either pure lipid vesicles or biological membranes. The annexins may underlie a number of different biological processes both inside and outside of cells, including membrane fusion during exocytosis, interactions between the cytoskeleton and membranes, regulation of lipid organization and metabolism, ion fluxes across membranes, and regulation of blood coagulation. The goals of this study are to determine the structural features of the annexins that underlie their interactions with membranes, and to use this information to design inhibitors of the activities of these proteins that may be applied to cells to determine the functions of the annexins in vivo. The structures of several annexins will be determined by X-ray diffraction analysis. The structural data will be used to design mutations in the annexins that should reveal the relationship between specific structural features of these proteins and certain in vitro activities. Specific annexin inhibitors will be designed, beginning with synthetic peptides corresponding to the N-terminal regions of the proteins that are critical for control of their membrane-aggregating activity. Effective in vitro inhibitors will be introduced into model secretory cells and fibroblasts to determine their effects on exocytosis and endocytosis, cell morphology and motility, and membrane-cytoskeletal organization. The information gained about the structure, mechanism of action, and modes of inhibition of these proteins may permit the development of pharmacological means to regulate the annexins in disorders of hormone release, blood coagulation, or cell structure.