ANALYZE THE COMPLEX PROTEIN ASSEMBLY USING SAXS

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This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Our lab focuses on the molecular basis of protein-protein interaction crucial for cell signal transduction in the normal and pathologic responses of humans. Currently, we work on two major areas. First is the interaction of human protease, insulin-degrading enzyme (IDE) with its substrates. IDE, a 110 kDa metalloprotease, is involved in the clearance of insulin and amyloid ¿, peptides crucial for the progression of diabetes and Alzheimers' disease. We have used SAXS to probe the open-closed conformational switch regulated by substrate binding as well as the allosteric regulation by the oligomerization of IDE. We recently identify macrophage inflammatory protein (MIP)-1¿ and MIP-1¿ as the novel substrate of IDE. MIP-1¿ and MIP-1¿ form high molecular weight aggregates. We used SAXS to demonstrate that MIP-1¿ and MIP-1¿ form the rod-shaped polydisperse polymers in solution. I propose to continue to use SAXS to understand the structure features of IDE and its substrates and how they interact to achieve their biological functions. Second is the interactions of toxins secreted by bacteria that cause anthrax. Anthrax bacteria also secrete >200 secreted factors. This includes three major anthrax toxins, protective antigen (PA), edema factor (EF), and lethal factor (LF) and the protein complex of PA-EF and PA-LF. We use SAXS analysis as a method to probe the structural features of purified anthrax toxins and toxin complex. We have also chosen 10 of such factors including pore-forming toxins, proteases, and cell wall modifying enzymes and will apply SAXS to probe the biophysical properties of these virulence factors.