Binding of GTPγ[35S] is regulated by GDP and receptor activation. Studies with the nociceptin/orphanin FQ receptor

摘要: Background and purpose:  We have examined the effects of ligand efficacy receptor density on binding guanosine 5′-[γ-thio]triphosphate (GTPγS) GDP to nociceptin/orphanin FQ (N/OFQ) peptide (NOP)-coupled G-proteins. Experimental approach:  In GTPγ[35S] experiments, using stable (CHOhNOP) inducible (CHOINDhNOP) recombinant human rat NOP we measured: (i) ligand-specific requirements; (ii) guanine nucleotide affinity/capacity; (iii) effect GTPγS association kinetics. Key results:  competition curves were shallow modelled by high- low-affinity components that relatively consistent between cell types tissue preparations. presence 1 µM N/OFQ a high-affinity site was also present, but fraction total reduced. an efficacy-dependent manner, partial agonists [F/G]N/OFQ(1-13)NH2 ([Phe1ψ(CH2-NH)Gly2]-nociceptin(1-13)NH2) naloxone benzoylhydrazone both reduced sites for (relative basal). While pIC50 did not decrease in N/OFQ, produced significant reduction site. Agonist-mediated affinity efficacy-dependent. displayed three affinities: high, conserved absence ligand; intermediate, present as low under basal conditions; (efficacy-dependent), during activation representing majority binding. Conclusions implications:  The regulated caused increased through affinity.