期刊
JOURNAL OF BIOLOGICAL CHEMISTRY
卷 288, 期 48, 页码 34470-34483出版社
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M113.515668
关键词
G Protein-coupled Receptors (GPCR); Mutagenesis Site-specific; Opiate Opioid; Pharmacology; Receptor Structure-Function
资金
- National Institutes of Health (NIH) [R01 DA017204]
- NIH, NIMH, Psychoactive Drug Screening Program (University of North Carolina at Chapel Hill)
- NIH PSI:Biology Grant [U54 GM094618]
Background: The -opioid receptor can be activated by structurally diverse agonists. Results: Four structurally diverse agonists differentially bound to and activated wild type and mutant -opioid receptors. Conclusion: The structural features of the agonists dictate how they interact with and stabilize G(i)-signaling receptor conformations. Significance: The results provide insights into the structural basis of opioid receptor ligand recognition and activation. The crystal structures of opioid receptors provide a novel platform for inquiry into opioid receptor function. The molecular determinants for activation of the -opioid receptor (KOR) were studied using a combination of agonist docking, functional assays, and site-directed mutagenesis. Eighteen positions in the putative agonist binding site of KOR were selected and evaluated for their effects on receptor binding and activation by ligands representing four distinct chemotypes: the peptide dynorphin A(1-17), the arylacetamide U-69593, and the non-charged ligands salvinorin A and the octahydroisoquinolinone carboxamide 1xx. Minimally biased docking of the tested ligands into the antagonist-bound KOR structure generated distinct binding modes, which were then evaluated biochemically and pharmacologically. Our analysis identified two types of mutations: those that affect receptor function primarily via ligand binding and those that primarily affect function. The shared and differential mechanisms of agonist binding and activation in KOR are further discussed. Usually, mutations affecting function more than binding were located at the periphery of the binding site and did not interact strongly with the various ligands. Analysis of the crystal structure along with the present results provide fundamental insights into the activation mechanism of the KOR and suggest that functional residues, along with water molecules detected in the crystal structure, may be directly involved in transduction of the agonist binding event into structural changes at the conserved rotamer switches, thus leading to receptor activation.
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