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Ligand and G-protein selectivity in the κ-opioid receptor.

ABSTRACT: The κ-opioid receptor (KOR) represents a highly desirable therapeutic target for treating not only pain but also addiction and affective disorders¹. However, the development of KOR analgesics has been hindered by the associated hallucinogenic side effects². The initiation of KOR signalling requires the G_i/o-family proteins including the conventional (G_i1, G_i2, G_i3, G_oA and G_oB) and nonconventional (G_z and G_g) subtypes. How hallucinogens exert their actions through KOR and how KOR determines G-protein subtype selectivity are not well understood. Here we determined the active-state structures of KOR in a complex with multiple G-protein heterotrimers-G_i1, G_oA, G_z and G_g-using cryo-electron microscopy. The KOR-G-protein complexes are bound to hallucinogenic salvinorins or highly selective KOR agonists. Comparisons of these structures reveal molecular determinants critical for KOR-G-protein interactions as well as key elements governing G_i/o-family subtype selectivity and KOR ligand selectivity. Furthermore, the four G-protein subtypes display an intrinsically different binding affinity and allosteric activity on agonist binding at KOR. These results provide insights into the actions of opioids and G-protein-coupling specificity at KOR and establish a foundation to examine the therapeutic potential of pathway-selective agonists of KOR.

SUBMITTER: Han J

PROVIDER: S-EPMC10172140 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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Ligand and G-protein selectivity in the κ-opioid receptor.

Han Jianming J Zhang Jingying J Nazarova Antonina L AL Bernhard Sarah M SM Krumm Brian E BE Zhao Lei L Lam Jordy Homing JH Rangari Vipin A VA Majumdar Susruta S Nichols David E DE Katritch Vsevolod V Yuan Peng P Fay Jonathan F JF Che Tao T

Nature 20230503 7960

The κ-opioid receptor (KOR) represents a highly desirable therapeutic target for treating not only pain but also addiction and affective disorders1. However, the development of KOR analgesics has been hindered by the associated hallucinogenic side effects2. The initiation of KOR signalling requires the Gi/o-family proteins including the conventional (Gi1, Gi2, Gi3, GoA and GoB) and nonconventional (Gz</s ...[more]

PMID: 37138078

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Project description:Kappa Opioid receptors（KORs） belong to family A of G-protein–coupled receptors (GPCRs), and the involvement of their ligands in analgesia and sedation has been extensively studied. KORs, which are expressed in the central and peripheral nervous systems are well known to exert potent anti-nociceptive effects by negatively modulating excitatory neurotransmission. The present study reveals that acute activation of KOR weaken SERT function via protein kinase B/Akt (Akt) and calcium calmodulin dependent kinase II (CaMKII) dependent signaling pathways. Therefore, studying phosphorylation will be helpful to elucidate the cellular mechanisms and downstream effector(s). Studies correlating the dysphoric effects of KOR agonists to the G-protein–independent activation of the p38 MAPK pathway suggest that identification of KOR agonists biased to G-protein–mediated signaling could lead to the development of therapeutics with reduced side effects. Therefore, to better understand the cellular mechanisms and downstream effector(s) by which KOR ligands produce these effects, we employed dimethyl labeling quantitative proteomic approach in this study. In this study, three monoclonal HEK293T cell lines with KOR over-expression were successfully constructed as our experimental models. Then, mass spectrometry-based quantitative approach, based on dimethyl labeling and TiO2 beads-coupled phosphopeptides enrichment, was utilized to analyze the KOR-mediated dynamic phosphorylation network in these three KOR-over-expressed monoclonal cell lines disturbed by LPK-26 or U50,488 treatments. Further Substrate-kinase prediction analysis of 212 up-regulated phosphosites indicated that 18 kinases, such as AKT1 and p70S6K, were involved in the up-regulation of KOR phosphorylation. The up-regulated phosphorylation of AKT1 and p70S6K were further confirmed by western blot, which was consistent with our mass spectrometry results. In summary, quantitative phosphoproteomic study of KOR-over-expressed cells upon agonist stimulation paves the way to systematical investigation of KOR-mediated phosphorylation network dynamics, which will be helpful to research about targeted therapy of KOR related diseases and mechanism study of analgesia addiction.

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Ligand and G-protein selectivity in the κ-opioid receptor.

Publications

Ligand and G-protein selectivity in the κ-opioid receptor.

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