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Molecular control of ?-opioid receptor signalling.

ABSTRACT: Opioids represent widely prescribed and abused medications, although their signal transduction mechanisms are not well understood. Here we present the 1.8?Å high-resolution crystal structure of the human ?-opioid receptor (?-OR), revealing the presence and fundamental role of a sodium ion in mediating allosteric control of receptor functional selectivity and constitutive activity. The distinctive ?-OR sodium ion site architecture is centrally located in a polar interaction network in the seven-transmembrane bundle core, with the sodium ion stabilizing a reduced agonist affinity state, and thereby modulating signal transduction. Site-directed mutagenesis and functional studies reveal that changing the allosteric sodium site residue Asn?131 to an alanine or a valine augments constitutive ?-arrestin-mediated signalling. Asp95Ala, Asn310Ala and Asn314Ala mutations transform classical ?-opioid antagonists such as naltrindole into potent ?-arrestin-biased agonists. The data establish the molecular basis for allosteric sodium ion control in opioid signalling, revealing that sodium-coordinating residues act as 'efficacy switches' at a prototypic G-protein-coupled receptor.

SUBMITTER: Fenalti G

PROVIDER: S-EPMC3931418 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

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Molecular control of δ-opioid receptor signalling.

Fenalti Gustavo G Giguere Patrick M PM Katritch Vsevolod V Huang Xi-Ping XP Thompson Aaron A AA Cherezov Vadim V Roth Bryan L BL Stevens Raymond C RC

Nature 20140112 7487

Opioids represent widely prescribed and abused medications, although their signal transduction mechanisms are not well understood. Here we present the 1.8 Å high-resolution crystal structure of the human δ-opioid receptor (δ-OR), revealing the presence and fundamental role of a sodium ion in mediating allosteric control of receptor functional selectivity and constitutive activity. The distinctive δ-OR sodium ion site architecture is centrally located in a polar interaction network in the seven-t ...[more]

PMID: 24413399

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Project description:BackgroundThe misuse of opioids stems, in part, from inadequate knowledge of molecular interactions between opioids and opioid receptors. It is still unclear why some opioids are far more addictive than others. The κ-opioid receptor (KOR) plays a critical role in modulating pain, addiction, and many other physiological and pathological processes. Butorphanol, an opioid analgesic, is a less addictive opioid with unique pharmacological profiles. In this study, we investigated the interaction between butorphanol and KOR to obtain insights into the safe usage of this medication.MethodsWe determined the binding affinity of butorphanol to KOR with a naltrexone competition study. Recombinant KORs expressed in mammalian cell membranes (Chem-1) were used for G-protein activation studies, and a human embryonic kidney-293 (HEK-293) cell line stably transfected with the human KOR was used for β-arrestin study as previously described in the literature. The effects of butorphanol on KOR internalization were investigated using mouse neuroblastoma Neuro2A cells stably transfected with mKOR-tdTomato fusion protein (N2A-mKOR-tdT) cells overexpressing KOR. The active-state KOR crystal structure was used for docking calculation of butorphanol to characterize the ligand binding site. Salvinorin A, a full KOR agonist, was used as a control for comparison.ResultsThe affinity of KOR for butorphanol is characterized by Kd of 0.1 ± 0.02 nM, about 20-fold higher compared with that of the µ-opioid receptor (MOR; 2.4 ± 1.2 nM). Our data indicate that butorphanol is more potent on KOR than on MOR. In addition, butorphanol acts as a partial agonist of KOR in the G-protein activation pathway and is a full agonist on the β-arrestin recruitment pathway, similar to that of salvinorin A. The activation of the β-arrestin pathway is further confirmed by KOR internalization. The in silico docking model indicates that both salvinorin A and butorphanol share the same binding cavity with the KOR full agonist MP1104. This cavity plays an important role in determining either agonist or antagonist effects of the ligand.ConclusionsIn conclusion, butorphanol is a partial KOR agonist in the G-protein activation pathway and a potent KOR full agonist in the β-arrestin recruitment pathway. The structure analysis offers insights into the molecular mechanism of KOR interaction and activation by butorphanol.

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Molecular control of ?-opioid receptor signalling.

Publications

Molecular control of δ-opioid receptor signalling.

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