Location of Regions of the Opioid Receptor Involved in Selective Agonist Binding

Abstract

To elucidate which portions of the opioid receptor molecules are involved in the ligand selectivity, we have expressed chimeric receptors between the rat δ- and μ-opioid receptors from cDNAs and analyzed their ligand binding properties. We demonstrate that the major binding determinant for the δ-selective enkephalin-related peptide, [D-Pen2,D-Pen5]enkephalin, resides within the region comprising the transmembrane segments V-VII and the intervening loop regions. On the other hand, the region spanning from the intracellular loop I to the amino-terminal half of the transmembrane segment III is shown to be involved in determining high-affinity binding of the μ-selective enkephalin-related peptides, [D-Ala2,MePhe4,Gly-ol5]enkephalin and [D-Ala2,MePhe4,Met-ol5]enkephalin, whereas the major determinant for binding of the μ-selective alkaloids, morphine and codeine, is demonstrated to exist in the region spanning the transmembrane segments V-VII. These results indicate that distinct regions of the opioid receptor determine the selectivity for the δ- and the μ-selective enkephalin-related peptides and that the binding determinant for the μ-selective alkaloids is distinct from that for the μ-selective enkephalin-related peptides. To elucidate which portions of the opioid receptor molecules are involved in the ligand selectivity, we have expressed chimeric receptors between the rat δ- and μ-opioid receptors from cDNAs and analyzed their ligand binding properties. We demonstrate that the major binding determinant for the δ-selective enkephalin-related peptide, [D-Pen2,D-Pen5]enkephalin, resides within the region comprising the transmembrane segments V-VII and the intervening loop regions. On the other hand, the region spanning from the intracellular loop I to the amino-terminal half of the transmembrane segment III is shown to be involved in determining high-affinity binding of the μ-selective enkephalin-related peptides, [D-Ala2,MePhe4,Gly-ol5]enkephalin and [D-Ala2,MePhe4,Met-ol5]enkephalin, whereas the major determinant for binding of the μ-selective alkaloids, morphine and codeine, is demonstrated to exist in the region spanning the transmembrane segments V-VII. These results indicate that distinct regions of the opioid receptor determine the selectivity for the δ- and the μ-selective enkephalin-related peptides and that the binding determinant for the μ-selective alkaloids is distinct from that for the μ-selective enkephalin-related peptides.