Research Project
6647525
DESCRIPTION (provided by applicant): Obtaining atomic resolution structures of membrane proteins such as G-protein coupled receptors (GPCRs) has been hampered by the difficulty to purify sufficient amounts of membrane protein. The only GPCR crystal structure available is bovine rhodopsin, which can be isolated in large amounts from bovine retina. We propose a novel approach to generate a source to purify large amounts of receptor, which could solve the bottleneck for GPCR crystallization. The covalently bound retinal of rhodopsin acts as an inverse agonist that maintains firmly the receptor in a very stable inactive state. Stability and homogeneity of the protein are key factors for the formation of highly ordered crystals that diffract at high resolution. The cannabinoid receptors (CBR) are the closest to rhodopsin in that when bound to the inverse agonist they exhibit a very stable inactive state, in this case associated with the G-protein, making them attractive GPCR targets for crystallization and structural elucidation. In Phase I we will test the feasibility of our novel source for receptor purification for CB1 and CB2 receptors, a prerequisite to a Phase II application for crystallization and structural elucidation of the CB1 and CB2 receptors, alone and complexed with G-protein in the inverse agonist bound state.
