The G protein G exhibits basal coupling but not preassembly with G protein-coupled receptors.

The G protein family transduces signals from a diverse group of G protein-coupled receptors (GPCRs). The observed specificity of G-GPCR coupling and the high rate of G signal transduction have been hypothesized to be enabled by the existence of stable associates between G proteins and their cognate GPCRs in the inactive state (G-GPCR preassembly). To test this hypothesis, we applied the recently developed technique of two-photon polarization microscopy (2PPM) to Gα subunits labeled with fluorescent proteins and four GPCRs: the α-adrenergic receptor, GABA, cannabinoid receptor type 1 (CBR), and dopamine receptor type 2. Our experiments with non-dissociating mutants of fluorescently labeled Gα subunits (exhibiting impaired dissociation from activated GPCRs) showed that 2PPM is capable of detecting GPCR-G protein interactions. 2PPM experiments with non-mutated fluorescently labeled Gα subunits and α-adrenergic receptor, GABA, or dopamine receptor type 2 receptors did not reveal any interaction between the G protein and the non-stimulated GPCRs. In contrast, non-stimulated CBR exhibited an interaction with the G protein. Further experiments revealed that this interaction is caused solely by CBR basal activity; no preassembly between CBR and the G protein could be observed. Our results demonstrate that four diverse GPCRs do not preassemble with non-active G However, we also show that basal GPCR activity allows interactions between non-stimulated GPCRs and G (basal coupling). These findings suggest that G interacts only with active GPCRs and that the well known high speed of GPCR signal transduction does not require preassembly between G proteins and GPCRs.