Approximately 35% of all drugs currently in clinical use target a single family of proteins: G-Protein Coupled Receptors (GPCRs). Due to the therapeutic importance of this class of proteins, it should be no surprise that proteins involved in regulating signaling initiated by GPCRs are gathering attention as potential drug targets. One such family of proteins is known as the Regulators of G-protein Signaling (RGS). These proteins function as negative regulators for G-protein signaling by binding to the GTP-bound G-alpha subunit, accelerating the hydrolysis of GTP to GDP. In turn, RGS proteins are themselves regulated by direct interactions with other biomolecules. For instance, RGS10, which is integral for neural and cardiovascular function, is differentially regulated by a sophisticated, poorly understood competitive binding to the membrane lipid phosphatidylinositol (3,4,5)-triphosphate (PIP3) and the ubiquitous calcium-signaling protein, calmodulin (CaM). Our current focus is on the interaction of the RGS10 protein with CaM and how this interaction affects the RGS binding to G-alpha. We have produced recombinant full-length RGS10 and the RGS domain only (RGS10 lacking N- and C-terminal regions) and purified them to homogeneity. By monitoring the intrinsic tryptophan fluorescence of RGS10, we have measured the affinities of the RGS10 proteins for CaM and their dependence on ionic strength. Here, the full-length RGS10 binds only marginally tighter to CaM than the RGS domain alone at low ionic strength and the affinities of both complexes decrease with increasing ionic strength. The results indicate 1:1 stoichiometry of the complexes. NMR studies show RGS10 binds preferentially to the C-domain of CaM. Through monitoring chemical shift perturbations in 1H, 15N-HSQC spectra, we have determined that the C-domain of CaM likely binds to the terminal regions of the RGS domain. Interestingly, CaM binding to RGS10 induces an allosteric effect on a critical loop for the interaction of RGS10 with the G-alpha subunit.
 A. J. Kimple, D. E. Bosch, P. M. Giguere and D. P. Siderovski, Pharmacol Rev 2011, 63, 728-749.
 M. Soundararajan, F. S. Willard, A. J. Kimple, A. P. Turnbull, L. J. Ball, G. A. Schoch, C. Gileadi, O. Y. Fedorov, E. F. Dowler, V. A. Higman, S. Q. Hutsell, M. Sundstrom, D. A. Doyle and D. P. Siderovski, Proc Natl Acad Sci U S A 2008, 105, 6457-6462.
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