Small angle neutron scattering study of rhodopsin oligomerization and G-protein coupling in a physiologically relevant lipid membrane

Although the oligomeric states of G-protein-coupled receptors (GPCRs) and interactions with cognate G proteins are central to their signal transduction capabilities, they remain poorly defined. In this study, we used small-angle neutron scattering (SANS) and a neutron contrast matching approach to elucidate the oligomeric states of the archetypal GPCR, rhodopsin, and its interaction with the G protein transducin (G_t). At a rhodopsin/lipid molar ratio of 1/360, we found that dark-adapted rhodopsin exists as a monomer, a finding consistent with its high functional activity measured upon photoactivation by spectrophotometry and the rate of catalyzed [³⁵S]-GTP-γ-S exchange. Following light activation, we observed that rhodopsin forms a stable 1:1 stoichiometric complex with G_t, the structure of which is consistent with recent cryo-EM data. In contrast, activated rhodopsin in the absence of G_t showed a propensity to form higher order oligomers. This research underscores the concentration-dependent nature of rhodopsin oligomerization and establishes SANS and the ability to produce appropriately contrast-matched samples, as a robust strategy for characterizing integral membrane protein interactions under biologically relevant conditions.