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- EMDB-37224: Cryo-EM structure of the GPR61-Gs complex -

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Entry
Database: EMDB / ID: EMD-37224
TitleCryo-EM structure of the GPR61-Gs complex
Map data
Sample
  • Complex: Cryo-EM structure of the GPR61 and mini-Gs complex with Nb35
    • Complex: GPR61 and mini-Gs
      • Protein or peptide: G-protein coupled receptor 61G protein-coupled receptor
      • Protein or peptide: Guanine nucleotide-binding protein G(olf) subunit alpha,Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
      • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
      • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
    • Complex: Nb35
      • Protein or peptide: Nanobody 35Single-domain antibody
KeywordsGPCR / Gs / MEMBRANE PROTEIN
Function / homology
Function and homology information


ligand-independent adenylate cyclase-activating G protein-coupled receptor signaling pathway / Adenylate cyclase activating pathway / arrestin family protein binding / PKA activation in glucagon signalling / hair follicle placode formation / Adenylate cyclase inhibitory pathway / developmental growth / D1 dopamine receptor binding / intracellular transport / Hedgehog 'off' state ...ligand-independent adenylate cyclase-activating G protein-coupled receptor signaling pathway / Adenylate cyclase activating pathway / arrestin family protein binding / PKA activation in glucagon signalling / hair follicle placode formation / Adenylate cyclase inhibitory pathway / developmental growth / D1 dopamine receptor binding / intracellular transport / Hedgehog 'off' state / positive regulation of cAMP-mediated signaling / adenylate cyclase-activating adrenergic receptor signaling pathway / activation of adenylate cyclase activity / adenylate cyclase activator activity / trans-Golgi network membrane / G protein-coupled receptor binding / G protein-coupled receptor activity / Olfactory Signaling Pathway / G-protein beta/gamma-subunit complex binding / Activation of the phototransduction cascade / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / bone development / G-protein activation / G protein-coupled acetylcholine receptor signaling pathway / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Prostacyclin signalling through prostacyclin receptor / Glucagon signaling in metabolic regulation / G beta:gamma signalling through CDC42 / adenylate cyclase-activating G protein-coupled receptor signaling pathway / ADP signalling through P2Y purinoceptor 12 / G beta:gamma signalling through BTK / Sensory perception of sweet, bitter, and umami (glutamate) taste / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / photoreceptor disc membrane / Adrenaline,noradrenaline inhibits insulin secretion / Glucagon-type ligand receptors / platelet aggregation / cognition / Vasopressin regulates renal water homeostasis via Aquaporins / positive regulation of GTPase activity / G alpha (z) signalling events / cellular response to catecholamine stimulus / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / ADORA2B mediated anti-inflammatory cytokines production / adenylate cyclase-activating dopamine receptor signaling pathway / ADP signalling through P2Y purinoceptor 1 / G beta:gamma signalling through PI3Kgamma / cellular response to prostaglandin E stimulus / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / sensory perception of taste / GPER1 signaling / G-protein beta-subunit binding / heterotrimeric G-protein complex / Inactivation, recovery and regulation of the phototransduction cascade / G alpha (12/13) signalling events / extracellular vesicle / signaling receptor complex adaptor activity / sensory perception of smell / Thrombin signalling through proteinase activated receptors (PARs) / retina development in camera-type eye / GTPase binding / Ca2+ pathway / phospholipase C-activating G protein-coupled receptor signaling pathway / positive regulation of cold-induced thermogenesis / G alpha (i) signalling events / fibroblast proliferation / G alpha (s) signalling events / G alpha (q) signalling events / Ras protein signal transduction / cell population proliferation / Extra-nuclear estrogen signaling / receptor complex / endosome membrane / endosome / G protein-coupled receptor signaling pathway / lysosomal membrane / GTPase activity / synapse / protein-containing complex binding / GTP binding / signal transduction / extracellular exosome / membrane / metal ion binding / plasma membrane / cytosol / cytoplasm
Similarity search - Function
G-protein alpha subunit, group S / G-alpha domain profile. / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / GGL domain / G-protein gamma-like domain superfamily ...G-protein alpha subunit, group S / G-alpha domain profile. / Guanine nucleotide binding protein (G-protein), alpha subunit / G protein alpha subunit, helical insertion / G-protein alpha subunit / G protein alpha subunit / G-protein, gamma subunit / G-protein gamma subunit domain profile. / GGL domain / G-protein gamma-like domain superfamily / G-protein gamma-like domain / GGL domain / G protein gamma subunit-like motifs / 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. / WD domain, G-beta repeat / WD40 repeats / WD40 repeat / Trp-Asp (WD) repeats profile. / Trp-Asp (WD) repeats circular profile. / WD40-repeat-containing domain superfamily / WD40/YVTN repeat-like-containing domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Guanine nucleotide-binding protein G(olf) subunit alpha / Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 / Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Guanine nucleotide-binding protein G(s) subunit alpha isoforms short / G-protein coupled receptor 61
Similarity search - Component
Biological speciesHomo sapiens (human) / Lama glama (llama)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.16 Å
AuthorsNie Y / Qiu Z / Zheng S
Funding support China, 1 items
OrganizationGrant numberCountry
Ministry of Science and Technology (MoST, China) China
CitationJournal: Nat Commun / Year: 2023
Title: Specific binding of GPR174 by endogenous lysophosphatidylserine leads to high constitutive G signaling.
Authors: Yingying Nie / Zeming Qiu / Sijia Chen / Zhao Chen / Xiaocui Song / Yan Ma / Niu Huang / Jason G Cyster / Sanduo Zheng /
Abstract: Many orphan G protein-coupled receptors (GPCRs) remain understudied because their endogenous ligands are unknown. Here, we show that a group of class A/rhodopsin-like orphan GPCRs including GPR61, ...Many orphan G protein-coupled receptors (GPCRs) remain understudied because their endogenous ligands are unknown. Here, we show that a group of class A/rhodopsin-like orphan GPCRs including GPR61, GPR161 and GPR174 increase the cAMP level similarly to fully activated D1 dopamine receptor (D1R). We report cryo-electron microscopy structures of the GPR61‒G, GPR161‒G and GPR174‒G complexes without any exogenous ligands. The GPR174 structure reveals that endogenous lysophosphatidylserine (lysoPS) is copurified. While GPR174 fails to respond to exogenous lysoPS, likely owing to its maximal activation by the endogenous ligand, GPR174 mutants with lower ligand binding affinities can be specifically activated by lysoPS but not other lipids, in a dose-dependent manner. Moreover, GPR174 adopts a non-canonical G coupling mode. The structures of GPR161 and GPR61 reveal that the second extracellular loop (ECL2) penetrates into the orthosteric pocket, possibly contributing to constitutive activity. Our work definitively confirms lysoPS as an endogenous GPR174 ligand and suggests that high constitutive activity of some orphan GPCRs could be accounted for by their having naturally abundant ligands.
History
DepositionAug 19, 2023-
Header (metadata) releaseOct 11, 2023-
Map releaseOct 11, 2023-
UpdateOct 11, 2023-
Current statusOct 11, 2023Processing site: PDBj / Status: Released

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Structure visualization

Supplemental images

emd_37224.png

Downloads & links

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Map

FileDownload / File: emd_37224.map.gz / Format: CCP4 / Size: 22.2 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.33
Minimum - Maximum-2.0973816 - 3.1335087
Average (Standard dev.)0.00054808 (±0.09814714)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions180180180
Spacing180180180
CellA=B=C: 194.40001 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: #1

Fileemd_37224_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: #2

Fileemd_37224_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Cryo-EM structure of the GPR61 and mini-Gs complex with Nb35

EntireName: Cryo-EM structure of the GPR61 and mini-Gs complex with Nb35
Components
  • Complex: Cryo-EM structure of the GPR61 and mini-Gs complex with Nb35
    • Complex: GPR61 and mini-Gs
      • Protein or peptide: G-protein coupled receptor 61G protein-coupled receptor
      • Protein or peptide: Guanine nucleotide-binding protein G(olf) subunit alpha,Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
      • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
      • Protein or peptide: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
    • Complex: Nb35
      • Protein or peptide: Nanobody 35Single-domain antibody

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Supramolecule #1: Cryo-EM structure of the GPR61 and mini-Gs complex with Nb35

SupramoleculeName: Cryo-EM structure of the GPR61 and mini-Gs complex with Nb35
type: complex / ID: 1 / Parent: 0 / Macromolecule list: all

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Supramolecule #2: GPR61 and mini-Gs

SupramoleculeName: GPR61 and mini-Gs / type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1-#4
Source (natural)Organism: Homo sapiens (human)

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Supramolecule #3: Nb35

SupramoleculeName: Nb35 / type: complex / ID: 3 / Parent: 1 / Macromolecule list: #5
Source (natural)Organism: Lama glama (llama)

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Macromolecule #1: G-protein coupled receptor 61

MacromoleculeName: G-protein coupled receptor 61 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 42.527605 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: DYKDDDDKAM ESSPIPQSSG NSSTLGRVPQ TPGPSTASGV PEVGLRDVAS ESVALFFMLL LDLTAVAGNA AVMAVIAKTP ALRKFVFVF HLCLVDLLAA LTLMPLAMLS SSALFDHALF GEVACRLYLF LSVCFVSLAI LSVSAINVER YYYVVHPMRY E VRMTLGLV ...String:
DYKDDDDKAM ESSPIPQSSG NSSTLGRVPQ TPGPSTASGV PEVGLRDVAS ESVALFFMLL LDLTAVAGNA AVMAVIAKTP ALRKFVFVF HLCLVDLLAA LTLMPLAMLS SSALFDHALF GEVACRLYLF LSVCFVSLAI LSVSAINVER YYYVVHPMRY E VRMTLGLV ASVLVGVWVK ALAMASVPVL GRVSWEEGAP SVPPGCSLQW SHSAYCQLFV VVFAVLYFLL PLLLILVVYC SM FRVARVA AMQHGPLPTW METPRQRSES LSSRSTMVTS SGAPQTTPHR TFGGGKAAVV LLAVGGQFLL CWLPYFSFHL YVA LSAQPI STGQVESVVT WIGYFCFTSN PFFYGCLNRQ IRGELSKQFV CFFKPAPELE VLFQGPLEVL FQGP

UniProtKB: G-protein coupled receptor 61

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Macromolecule #2: Guanine nucleotide-binding protein G(olf) subunit alpha,Guanine n...

MacromoleculeName: Guanine nucleotide-binding protein G(olf) subunit alpha,Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
type: protein_or_peptide / ID: 2 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 29.008785 KDa
Recombinant expressionOrganism: Homo sapiens (human)
SequenceString: NSKTTEDQRN EEKAQREANK KIEKQLQKDK QVYRATHRLL LLGADNSGKS TIVKQMRILH GGSGGSGGTS GIFETKFQVD KVNFHMFDV GGQRDERRKW IQCFNDVTAI IFVVDSSDYN RLQEALNLFK SIWNNRWLRT ISVILFLNKQ DLLAEKVLAG K SKIEDYFP ...String:
NSKTTEDQRN EEKAQREANK KIEKQLQKDK QVYRATHRLL LLGADNSGKS TIVKQMRILH GGSGGSGGTS GIFETKFQVD KVNFHMFDV GGQRDERRKW IQCFNDVTAI IFVVDSSDYN RLQEALNLFK SIWNNRWLRT ISVILFLNKQ DLLAEKVLAG K SKIEDYFP EFARYTTPED ATPEPGEDPR VTRAKYFIRD EFLRISTASG DGRHYCYPHF TCAVDTENAR RIFNDCRDII QR MHLRQYE LL

UniProtKB: Guanine nucleotide-binding protein G(olf) subunit alpha, Guanine nucleotide-binding protein G(s) subunit alpha isoforms short

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Macromolecule #3: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1

MacromoleculeName: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
type: protein_or_peptide / ID: 3 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 39.286891 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString: HHHHHHLEVL FQGPGSSGSE LDQLRQEAEQ LKNQIRDARK ACADATLSQI TNNIDPVGRI QMRTRRTLRG HLAKIYAMHW GTDSRLLVS ASQDGKLIIW DSYTTNKVHA IPLRSSWVMT CAYAPSGNYV ACGGLDNICS IYNLKTREGN VRVSRELAGH T GYLSCCRF ...String:
HHHHHHLEVL FQGPGSSGSE LDQLRQEAEQ LKNQIRDARK ACADATLSQI TNNIDPVGRI QMRTRRTLRG HLAKIYAMHW GTDSRLLVS ASQDGKLIIW DSYTTNKVHA IPLRSSWVMT CAYAPSGNYV ACGGLDNICS IYNLKTREGN VRVSRELAGH T GYLSCCRF LDDNQIVTSS GDTTCALWDI ETGQQTTTFT GHTGDVMSLS LAPDTRLFVS GACDASAKLW DVREGMCRQT FT GHESDIN AICFFPNGNA FATGSDDATC RLFDLRADQE LMTYSHDNII CGITSVSFSK SGRLLLAGYD DFNCNVWDAL KAD RAGVLA GHDNRVSCLG VTDDGMAVAT GSWDSFLKIW N

UniProtKB: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1

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Macromolecule #4: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2

MacromoleculeName: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
type: protein_or_peptide / ID: 4 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 7.845078 KDa
Recombinant expressionOrganism: Spodoptera frugiperda (fall armyworm)
SequenceString:
MASNNTASIA QARKLVEQLK MEANIDRIKV SKAAADLMAY CEAHAKEDPL LTPVPASENP FREKKFFSAI L

UniProtKB: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2

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Macromolecule #5: Nanobody 35

MacromoleculeName: Nanobody 35 / type: protein_or_peptide / ID: 5 / Number of copies: 1 / Enantiomer: LEVO
Source (natural)Organism: Lama glama (llama)
Molecular weightTheoretical: 17.352498 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString:
MKYLLPTAAA GLLLLAAQPA MAQVQLQESG GGLVQPGGSL RLSCAASGFT FSNYKMNWVR QAPGKGLEWV SDISQSGASI SYTGSVKGR FTISRDNAKN TLYLQMNSLK PEDTAVYYCA RCPAPFTRDC FDVTSTTYAY RGQGTQVTVS SHHHHHHEPE A

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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

Concentration7.0 mg/mL
BufferpH: 7.4
GridModel: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 35 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 281 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.48 µm / Nominal defocus min: 0.87 µm / Nominal magnification: 64000
Sample stageCooling holder cryogen: NITROGEN
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number real images: 923 / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 277452
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

7f0t

Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final 3D classificationNumber classes: 3
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionNumber classes used: 1 / Resolution.type: BY AUTHOR / Resolution: 3.16 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 192672

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