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- PDB-8k9k: Full agonist-bound mu-type opioid receptor-G protein complex -

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Basic information

Entry
Database: PDB / ID: 8k9k
TitleFull agonist-bound mu-type opioid receptor-G protein complex
Components
  • DAMGO
  • G protein subunit alpha i3
  • Guanine nucleotide binding protein, beta polypeptide 1
  • Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
  • Soluble cytochrome b562,Mu-type opioid receptor
  • scFv16
KeywordsSIGNALING PROTEIN
Function / homology
Function and homology information


positive regulation of cAMP-mediated signaling / Opioid Signalling / beta-endorphin receptor activity / morphine receptor activity / negative regulation of Wnt protein secretion / negative regulation of cAMP-mediated signaling / regulation of cellular response to stress / G protein-coupled opioid receptor signaling pathway / negative regulation of nitric oxide biosynthetic process / behavioral response to ethanol ...positive regulation of cAMP-mediated signaling / Opioid Signalling / beta-endorphin receptor activity / morphine receptor activity / negative regulation of Wnt protein secretion / negative regulation of cAMP-mediated signaling / regulation of cellular response to stress / G protein-coupled opioid receptor signaling pathway / negative regulation of nitric oxide biosynthetic process / behavioral response to ethanol / adenylate cyclase-inhibiting G protein-coupled acetylcholine receptor signaling pathway / sensory perception / neuropeptide binding / regulation of NMDA receptor activity / GTP metabolic process / positive regulation of neurogenesis / negative regulation of cytosolic calcium ion concentration / positive regulation of macroautophagy / G protein-coupled receptor signaling pathway, coupled to cyclic nucleotide second messenger / G-protein alpha-subunit binding / neuropeptide signaling pathway / voltage-gated calcium channel activity / MECP2 regulates neuronal receptors and channels / sensory perception of pain / Peptide ligand-binding receptors / G protein-coupled receptor binding / electron transport chain / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / G protein-coupled receptor activity / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / G-protein beta/gamma-subunit complex binding / centriolar satellite / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / G-protein activation / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Prostacyclin signalling through prostacyclin receptor / G beta:gamma signalling through CDC42 / Glucagon signaling in metabolic regulation / G beta:gamma signalling through BTK / ADP signalling through P2Y purinoceptor 12 / Glucagon-type ligand receptors / Adrenaline,noradrenaline inhibits insulin secretion / Vasopressin regulates renal water homeostasis via Aquaporins / GDP binding / G alpha (z) signalling events / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / cellular response to catecholamine stimulus / positive regulation of nitric oxide biosynthetic process / ADORA2B mediated anti-inflammatory cytokines production / ADP signalling through P2Y purinoceptor 1 / G beta:gamma signalling through PI3Kgamma / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / adenylate cyclase-activating dopamine receptor signaling pathway / GPER1 signaling / cellular response to prostaglandin E stimulus / G-protein beta-subunit binding / heterotrimeric G-protein complex / G alpha (12/13) signalling events / Thrombin signalling through proteinase activated receptors (PARs) / positive regulation of cytosolic calcium ion concentration / Ca2+ pathway / midbody / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / fibroblast proliferation / G alpha (i) signalling events / Interleukin-4 and Interleukin-13 signaling / G alpha (s) signalling events / phospholipase C-activating G protein-coupled receptor signaling pathway / G alpha (q) signalling events / perikaryon / periplasmic space / Extra-nuclear estrogen signaling / electron transfer activity / positive regulation of ERK1 and ERK2 cascade / neuron projection / endosome / ciliary basal body / G protein-coupled receptor signaling pathway / iron ion binding / axon / Golgi membrane / negative regulation of cell population proliferation / cell division / GTPase activity / synapse / heme binding / dendrite / endoplasmic reticulum membrane / GTP binding / nucleolus / endoplasmic reticulum / Golgi apparatus / signal transduction / extracellular exosome / nucleoplasm / metal ion binding / membrane
Similarity search - Function
G protein beta WD-40 repeat protein / Mu opioid receptor / Opioid receptor / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / G-protein alpha subunit, group I / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit ...G protein beta WD-40 repeat protein / Mu opioid receptor / Opioid receptor / Cytochrome b562 / Cytochrome b562 / Cytochrome c/b562 / G-protein alpha subunit, group I / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G-alpha domain profile. / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / G-protein gamma-like domain / G-protein gamma-like domain superfamily / GGL domain / G protein gamma subunit-like motifs / GGL domain / Guanine nucleotide-binding protein, beta subunit / G-protein, beta subunit / G-protein coupled receptors family 1 signature. / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / 7 transmembrane receptor (rhodopsin family) / G-protein beta WD-40 repeat / WD40 repeat, conserved site / Trp-Asp (WD) repeats signature. / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40 repeats / WD40 repeat / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
DAMGO / G protein subunit alpha i3 / Guanine nucleotide binding protein, beta polypeptide 1 / Soluble cytochrome b562 / Mu-type opioid receptor / Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Homo sapiens (human)
Macaca mulatta (Rhesus monkey)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.98 Å
AuthorsHisano, T. / Uchikubo-Kamo, T. / Shirouzu, M. / Imai, S. / Kaneko, S. / Shimada, I.
Funding support Japan, 4items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)JP17H06097 Japan
Japan Society for the Promotion of Science (JSPS)JP21H02619 Japan
Japan Society for the Promotion of Science (JSPS)JP23KJ0574 Japan
Japan Agency for Medical Research and Development (AMED)21ae0121028h0001 Japan
CitationJournal: Nat Commun / Year: 2024
Title: Structural and dynamic insights into the activation of the μ-opioid receptor by an allosteric modulator.
Authors: Shun Kaneko / Shunsuke Imai / Tomomi Uchikubo-Kamo / Tamao Hisano / Nobuaki Asao / Mikako Shirouzu / Ichio Shimada /
Abstract: G-protein-coupled receptors (GPCRs) play pivotal roles in various physiological processes. These receptors are activated to different extents by diverse orthosteric ligands and allosteric modulators. ...G-protein-coupled receptors (GPCRs) play pivotal roles in various physiological processes. These receptors are activated to different extents by diverse orthosteric ligands and allosteric modulators. However, the mechanisms underlying these variations in signaling activity by allosteric modulators remain largely elusive. Here, we determine the three-dimensional structure of the μ-opioid receptor (MOR), a class A GPCR, in complex with the G protein and an allosteric modulator, BMS-986122, using cryogenic electron microscopy. Our results reveal that BMS-986122 binding induces changes in the map densities corresponding to R167 and Y254, key residues in the structural motifs conserved among class A GPCRs. Nuclear magnetic resonance analyses of MOR in the absence of the G protein reveal that BMS-986122 binding enhances the formation of the interaction between R167 and Y254, thus stabilizing the fully-activated conformation, where the intracellular half of TM6 is outward-shifted to allow for interaction with the G protein. These findings illuminate that allosteric modulators like BMS-986122 can potentiate receptor activation through alterations in the conformational dynamics in the core region of GPCRs. Together, our results demonstrate the regulatory mechanisms of GPCRs, providing insights into the rational development of therapeutics targeting GPCRs.
History
DepositionAug 1, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0May 29, 2024Provider: repository / Type: Initial release
Revision 1.1Oct 23, 2024Group: Data collection / Structure summary
Category: em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
R: Soluble cytochrome b562,Mu-type opioid receptor
S: DAMGO
A: G protein subunit alpha i3
B: Guanine nucleotide binding protein, beta polypeptide 1
G: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
H: scFv16


Theoretical massNumber of molelcules
Total (without water)166,2136
Polymers166,2136
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Protein , 4 types, 4 molecules RABG

#1: Protein Soluble cytochrome b562,Mu-type opioid receptor / Cytochrome b-562 / M-OR-1 / MOR-1 / Mu opiate receptor / Mu opioid receptor / MOP / hMOP


Mass: 51321.855 Da / Num. of mol.: 1 / Mutation: M7W,H102I,R106L
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli (E. coli), (gene. exp.) Homo sapiens (human)
Gene: cybC, OPRM1, MOR1 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P0ABE7, UniProt: P35372
#3: Protein G protein subunit alpha i3 / Guanine nucleotide-binding protein G(K) subunit alpha


Mass: 40585.137 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Macaca mulatta (Rhesus monkey) / Gene: GNAI3 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: F6VL43
#4: Protein Guanine nucleotide binding protein, beta polypeptide 1


Mass: 38522.098 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: M1ERZ7
#5: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / G gamma-I


Mass: 7861.143 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNG2 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P59768

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Protein/peptide / Antibody , 2 types, 2 molecules SH

#2: Protein/peptide DAMGO


Type: Peptide-like / Class: Synthetic opioid / Mass: 513.587 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) / References: DAMGO
#6: Antibody scFv16


Mass: 27409.588 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) synthetic construct (others) / Production host: Spodoptera frugiperda (fall armyworm)

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Details

Has ligand of interestY
Has protein modificationY

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Full agonist-bound mu-type opioid receptor-G protein complex
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Molecular weightUnits: MEGADALTONS / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Spodoptera frugiperda (fall armyworm)
Buffer solutionpH: 7.5
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2000 nm / Nominal defocus min: 800 nm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 2.3 sec. / Electron dose: 50.5 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)
EM imaging opticsEnergyfilter name: GIF Quantum LS / Energyfilter slit width: 15 eV

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Processing

EM software
IDNameVersionCategory
2PHENIX1.21rc1_5015:model refinement
5CTFFIND4CTF correction
10RELION3.1.2initial Euler assignment
11RELION3.1.2final Euler assignment
12RELION3.1.2classification
13RELION3.1.23D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.98 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 308249 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0039119
ELECTRON MICROSCOPYf_angle_d0.62412382
ELECTRON MICROSCOPYf_dihedral_angle_d4.9951257
ELECTRON MICROSCOPYf_chiral_restr0.0451424
ELECTRON MICROSCOPYf_plane_restr0.0051557

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