STOICHIOMETRY OF RECEPTOR-GS-ADENYLATE CYCLASE INTERACTIONS

Little is known about the relative stoichiometry of guanine nucleotide-binding (G) proteins relative to the effector systems to which they link. We addressed this question for the stimulatory G protein (G(s)) linked to adenylate cyclase. Forskolin stimulates the catalytic subunit of adenylate cyclase (C), but it has a higher efficacy and potency when C also interacts with the G protein G(s). Accordingly, we measured high-affinity [H-3]forskolin binding to intact cells to assay alpha-s-C complexes. No high-affinity specific binding occurred with unstimulated cells. The beta-adrenergic agonist isoproterenol promoted the binding of [H-3]forskolin to about 3000 sites per cell, suggesting that each receptor on average activates at least several G(s) molecules. Activating G(s) directly with cholera toxin maximally promoted [H-3]forskolin binding to a similar number of sites, suggesting that this is the maximal number of alpha-s-C complexes formed per cell. We conclude that each cell likely contains only a few thousand functional copies of C, and that the availability of C (rather than G(s), which exists in more than 100,000 copies per cell) is likely to be limiting for agonist stimulation of adenylate cyclase activity.