Switching Agonist/Antagonist Properties of Opiate Alkaloids at the δ Opioid Receptor Using Mutations Based on the Structure of the Orphanin FQ Receptor

Abstract In an earlier study, we have demonstrated that by mutating five amino acid residues to those conserved in the opioid receptors, the OFQ receptor could be converted to a functional receptor that bound many opioid alkaloids with nanomolar affinities. Surprisingly, when the reciprocal mutations, Lys-214 → Ala (TM5), Ile-277 → Val/His-278 → Gln/Ile-279 → Val (TM6), and Ile-304 → Thr (TM7), are introduced in the δ receptor, neither the individual mutations nor their various combinations significantly reduce the binding affinities of opioid alkaloids tested. However, these mutations cause profound alterations in the functional characteristics of the mutant receptors as measured in guanosine 5′-3-O-(thio)triphosphate binding assays. Some agonists become antagonists at some constructs as they lose their ability to activate them. Some alkaloid antagonists are transformed into agonists at other constructs, but their agonistic effects can still be blocked by the peptide antagonist TIPP. Even the δ inverse agonist 7-benzylidenenaltrexone becomes an agonist at the mutant containing both the Ile-277 → Val/His-278 → Gln/Ile-279 → Val and Ile-304 → Thr mutations. Thus, although the mutated residues are thought to be part of the binding pocket, they are critically involved in the control of the δ receptor activation process. These findings shed light on some of the structural bases of ligand efficacy. They are also compatible with the hypothesis that a ligand may achieve high affinity binding in several different ways, each having different effects on receptor activation.