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- EMDB-43653: Kappa opioid receptor:Galphai protein in complex with inverse ago... -

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Basic information

Entry
Database: EMDB / ID: EMD-43653
TitleKappa opioid receptor:Galphai protein in complex with inverse agonist GB18, Original map receptor
Map data
Sample
  • Complex: Kappa opioid receptor in complex with heterotrimeric G protein (Gai/b/g) and inverse agonist JDTic
KeywordsG protein coupled receptor / Opioid receptor / MEMBRANE PROTEIN
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsGati C / Motiwala Z / Tyson AS / Styrpejko D / Han GW / Khan S / Ramos-Gonzalez N / Shenvi R / Majumdar S
Funding support United States, 2 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM144965 United States
National Institutes of Health/National Center for Complementary and Integrative Health (NIH/NCCIH)AT012075 United States
CitationJournal: Nat Chem Biol / Year: 2025
Title: Molecular mechanisms of inverse agonism via κ-opioid receptor-G protein complexes.
Authors: Aaliyah S Tyson / Saif Khan / Zenia Motiwala / Gye Won Han / Zixin Zhang / Mohsen Ranjbar / Daniel Styrpejko / Nokomis Ramos-Gonzalez / Stone Woo / Kelly Villers / Delainey Landaker / Terry ...Authors: Aaliyah S Tyson / Saif Khan / Zenia Motiwala / Gye Won Han / Zixin Zhang / Mohsen Ranjbar / Daniel Styrpejko / Nokomis Ramos-Gonzalez / Stone Woo / Kelly Villers / Delainey Landaker / Terry Kenakin / Ryan Shenvi / Susruta Majumdar / Cornelius Gati /
Abstract: Opioid receptors, a subfamily of G protein-coupled receptors (GPCRs), are key therapeutic targets. In the canonical GPCR activation model, agonist binding is required for receptor-G protein complex ...Opioid receptors, a subfamily of G protein-coupled receptors (GPCRs), are key therapeutic targets. In the canonical GPCR activation model, agonist binding is required for receptor-G protein complex formation, while antagonists prevent G protein coupling. However, many GPCRs exhibit basal activity, allowing G protein association without an agonist. The pharmacological impact of agonist-free receptor-G protein complexes is poorly understood. Here we present biochemical evidence that certain κ-opioid receptor (KOR) inverse agonists can act via KOR-G protein complexes. To investigate this phenomenon, we determined cryo-EM structures of KOR-G protein complexes with three inverse agonists: JDTic, norBNI and GB18, corresponding to structures of inverse agonist-bound GPCR-G protein complexes. Remarkably, the orthosteric binding pocket resembles the G protein-free 'inactive' receptor conformation, while the receptor remains coupled to the G protein. In summary, our work challenges the canonical model of receptor antagonism and offers crucial insights into GPCR pharmacology.
History
DepositionFeb 7, 2024-
Header (metadata) releaseFeb 5, 2025-
Map releaseFeb 5, 2025-
UpdateFeb 5, 2025-
Current statusFeb 5, 2025Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43653.png

Downloads & links

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Map

FileDownload / File: emd_43653.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
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AxesZ (Sec.)Y (Row.)X (Col.)
1.29 Å/pix.
x 256 pix.
= 331.264 Å		1.29 Å/pix.
x 256 pix.
= 331.264 Å		1.29 Å/pix.
x 256 pix.
= 331.264 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.294 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.5
Minimum - Maximum-1.0632378 - 1.678865
Average (Standard dev.)0.00017451198 (±0.034788784)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 331.264 Å
α=β=γ: 90.0 °

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Supplemental data

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Additional map: #1

Fileemd_43653_additional_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

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Half map: #1

Fileemd_43653_half_map_1.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_43653_half_map_2.map
Projections & Slices
AxesZYX

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Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Kappa opioid receptor in complex with heterotrimeric G protein (G...

EntireName: Kappa opioid receptor in complex with heterotrimeric G protein (Gai/b/g) and inverse agonist JDTic
Components
  • Complex: Kappa opioid receptor in complex with heterotrimeric G protein (Gai/b/g) and inverse agonist JDTic

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Supramolecule #1: Kappa opioid receptor in complex with heterotrimeric G protein (G...

SupramoleculeName: Kappa opioid receptor in complex with heterotrimeric G protein (Gai/b/g) and inverse agonist JDTic
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#5
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 130 kDa/nm

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration18 mg/mL
BufferpH: 7.5
Component:
ConcentrationFormulaName
20.0 mMHEPESHEPES
150.0 mMNaClsodium chloride
0.001 %LMNGdetergent
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 200 / Support film - Material: GOLD / Support film - topology: HOLEY ARRAY
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 1 / Average electron dose: 60.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated magnification: 105000 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: -25.0 µm / Nominal defocus min: -15.0 µm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: INSILICO MODEL / In silico model: Ab initio
Final reconstructionResolution.type: BY AUTHOR / Resolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. v4.5.3) / Number images used: 230378
Initial angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. v4.5.3)
Final angle assignmentType: ANGULAR RECONSTITUTION / Software - Name: cryoSPARC (ver. v4.5.3)
Final 3D classificationNumber classes: 2 / Software - Name: cryoSPARC (ver. v4.5.3)

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