Structural aspects of urokinase receptor in vascular biology

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DESCRIPTION (provided by applicant): Urokinase plasminogen activator (uPA) together with its cell surface receptor (uPAR) mediates surface-bound plasminogen activation and has been recognized to play essential roles in several biological processes important in vascular biology, including clot lysis, cell adhesion, cell migration, inflammation, signal transduction, angiogenesis, and chemotaxis. Therefore, the study of this system will be critical to understand the causes, prevention, and treatment of vascular disorders and diseases including tumor invasion, tumor metastasis, neointima formation, infection with human immunodeficiency virus-1 (HIV-1), Alzheimer disease, and multiple sclerosis. The molecular basis for these broad physiological and pathophysiological roles of uPAR/uPA system stems from uPAR's capability to interact with many diverse ligands, e.g., uPA, vitronectin, integrins (31, 32, 33), LRP, GPCR, and the dynamic conformational changes caused by these protein-protein interactions. The principal investigator's recent results (Huai, et al. "Structure of Human Urokinase Plasminogen Activator in Complex", Science, in press, 2006) suggest conformational flexibility of uPAR. The goal of this proposal is to systematically study the structural basis and the dynamic nature of the interactions between uPAR and its ligands. The principal investigator proposes to study the structures of soluble uPAR (suPAR) (aim 1), suPAR in complex with vitronectin (aim 2), suPAR in complex with full- length uPA (aim 3), and to compare and analyze the structures of suPAR and suPAR-ligand complexes (aim 4). Protein X-ray crystallography will be the main tool to study these protein-protein interactions, but other common molecular and cell biology techniques will also be used. The principal investigator has established methods to generate needed reagents and has made several preliminary suPAR-ligand crystals, providing a solid basis for these proposed studies. These systematic structural studies of uPAR and its complexes with physiologically important uPA forms and vitronectin will understand the dynamic interactions between uPAR and its ligands, identify unanimously the binding determinants on the receptor and the ligands that confer therapeutic accessibility, and provide a framework for future design and optimization of small molecular antagonists to regulate these protein-protein interactions and to intervene the pathologic consequences resulting from these interactions.