Development of novel biosensors to study receptor-mediated activation of the G-protein subunits G_s and G_olf

G_s (G_s) and G_olf (G_olf) are highly homologous G-protein subunits that activate adenylate cyclase, thereby serving as crucial mediators of intracellular signaling. Because of their dramatically different brain expression patterns, we studied similarities and differences between their activation processes with the aim of comparing their receptor coupling mechanisms. We engineered novel luciferase- and Venus-fused Gα constructs that can be used in bioluminescence resonance energy transfer assays. In conjunction with molecular simulations, these novel biosensors were used to determine receptor activation–induced changes in conformation. Relative movements in G_s were consistent with the crystal structure of β₂ adrenergic receptor in complex with G_s. Conformational changes in G_olf activation are shown to be similar to those in G_s. Overall the current study reveals general similarities between G_s and G_olf activation at the molecular level and provides a novel set of tools to search for G_s- and G_olf-specific receptor pharmacology. In view of the wide functional and pharmacological roles of G_s- and G_olf-coupled dopamine D₁ receptor and adenosine A_2A receptor in the brain and other organs, elucidating their differential structure–function relationships with G_s and G_olf might provide new approaches for the treatment of a variety of neuropsychiatric disorders. In particular, these novel biosensors can be used to reveal potentially therapeutic dopamine D₁ receptor and adenosine A_2A receptor ligands with functionally selective properties between G_s and G_olf signaling.