Selectivity of mu-opioid receptor determined by interfacial residues near third extracellular loop.
We hypothesized that the selectivity profile of the rat mu-opioid receptor for opioid receptor-selective ligands is determined by the nature of the amino acid residues at highly divergent sites in the ligand-binding pocket. To determine which characteristics of these residues contribute to opioid receptor ligand selectivity, we made various mutant receptors that replaced the Lys(303) and Trp(318) residues near the extracellular interface of transmembrane domains VI and VII, respectively. Ligand binding determinations using transiently transfected monkey kidney epithelial (COS-1) cells show that Lys(303) mutations cause little change in the receptor binding profile, whereas the Trp(318) mutant receptors have considerably lower affinity for micro-opioid receptor-selective ligands and greatly increased affinity for delta-opioid receptor-selective ligands. The nature of these mutations show that this effect is not due to sterics or charge alone. [35S]guanosine-5'-O-(3-thio)-triphosphate ([35S]GTPgammaS) activity assays show that these residues may influence functional, as well as binding selection. We conclude that a primary role for Trp(318) is to form a basis for ligand selectivity.
Bonner
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G
Mental Health Research Institute, University of Michigan, Ann Arbor 48109-0720, USA. gbonner@umich.edu
Meng
F
F
Akil
H
H
D019943:Amino Acid Substitution; D000818:Animals; D001549:Benzamides / Q000378:metabolism / Q000494:pharmacology; D001575:Benzomorphans / Q000378:metabolism / Q000494:pharmacology; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive / Q000187:drug effects; D019556:COS Cells; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)- / Q000378:metabolism / Q000494:pharmacology; D020881:Enkephalin, D-Penicillamine (2,5)- / Q000378:metabolism / Q000494:pharmacology; D005283:Fentanyl / Q000378:metabolism / Q000494:pharmacology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D008024:Ligands; D009020:Morphine / Q000378:metabolism / Q000494:pharmacology; D009154:Mutation; D009270:Naloxone / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D009271:Naltrexone / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D009292:Narcotic Antagonists / Q000378:metabolism / Q000494:pharmacology; D010455:Peptides / Q000378:metabolism / Q000494:pharmacology; D010879:Piperazines / Q000378:metabolism / Q000494:pharmacology; D011487:Protein Conformation; D011869:Radioligand Assay; D051381:Rats; D017450:Receptors, Opioid, mu / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D013462:Sulfur Radioisotopes
D001549:Benzamides; D001575:Benzomorphans; D008024:Ligands; D009292:Narcotic Antagonists; D010455:Peptides; D010879:Piperazines; D017450:Receptors, Opioid, mu; D013462:Sulfur Radioisotopes; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; C088464:4-(alpha-(4-allyl-2,5-dimethyl-1-piperazinyl)-3-methoxybenzyl)-N,N-diethylbenzamide; D009270:Naloxone; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D009271:Naltrexone; C009862:connective tissue-activating peptide; D009020:Morphine; C034643:naloxonazine; D020881:Enkephalin, D-Penicillamine (2,5)-; C055382:naltrindole; C027248:bremazocine; D005283:Fentanyl