Probing Heterotrimeric G-protein Signaling using High-resolution NMR
John Marino, NIST, Center for Advanced Research in Biotechnology
28 Oct 2005
G-protein coupled receptors (GPCR's) represent a diverse group of seven transmembrane helix receptors that require ligand-dependent activation to initiate heterotrimeric (abg) G-protein mediated intracellular signaling cascades. Activation of a G-protein by its agonist stimulated GPCR (R*) requires the propagation of structural signals from the receptor binding interface to the guanine nucleotide-binding pocket. The structural basis for the interaction of a GPCR with its cognate G-protein and the subsequent activation of the G-protein by R* is not well understood. Using light-activated signaling of the G-protein, transducin (Gt), by rhodopsin as a model system, the interaction of G-protein with solubilized native receptor, as well as a soluble mimic of R*, is being probed using high-resolution NMR methods. Selective stable isotope labeling and mutation of the G-protein a-subunit (Ga) has allowed the detection of structural and dynamic changes in Ga that correlate with heterotrimer formation, receptor interaction and guanine nucleotide exchange. Analysis of the conformations of the different functional states of Ga and their implications for how signals may propagate from the receptor to the guanine nucleotide-binding pocket of Ga will be presented.