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- PDB-7bb7: AVP-V2R-Galphas-beta1-gamma2-Nb35(T state) -

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Basic information

Entry
Database: PDB / ID: 7bb7
TitleAVP-V2R-Galphas-beta1-gamma2-Nb35(T state)
Components
  • (Guanine nucleotide-binding protein ...) x 3
  • Nanobody 35
  • Vasopressin
  • Vasopressin V2 receptor
KeywordsMEMBRANE PROTEIN / Signaling protein / G protein-coupled receptor / Hormrne / Vasopressin / V2R
Function / homology
Function and homology information


renal water retention / Defective AVP does not bind AVPR2 and causes neurohypophyseal diabetes insipidus (NDI) / Vasopressin-like receptors / regulation of systemic arterial blood pressure by vasopressin / vasopressin receptor activity / renal water absorption / hemostasis / telencephalon development / positive regulation of systemic arterial blood pressure / adenylate cyclase-activating G protein-coupled bile acid receptor signaling pathway ...renal water retention / Defective AVP does not bind AVPR2 and causes neurohypophyseal diabetes insipidus (NDI) / Vasopressin-like receptors / regulation of systemic arterial blood pressure by vasopressin / vasopressin receptor activity / renal water absorption / hemostasis / telencephalon development / positive regulation of systemic arterial blood pressure / adenylate cyclase-activating G protein-coupled bile acid receptor signaling pathway / adenylate cyclase-activating serotonin receptor signaling pathway / regulation of skeletal muscle contraction / positive regulation of intracellular signal transduction / PKA activation in glucagon signalling / positive regulation of vasoconstriction / hair follicle placode formation / developmental growth / intracellular transport / endocytic vesicle / D1 dopamine receptor binding / vascular endothelial cell response to laminar fluid shear stress / renal water homeostasis / Hedgehog 'off' state / activation of adenylate cyclase activity / cellular response to hormone stimulus / cellular response to acidic pH / adenylate cyclase-activating adrenergic receptor signaling pathway / response to cytokine / cellular response to glucagon stimulus / intracellular glucose homeostasis / adenylate cyclase activator activity / positive regulation of insulin secretion involved in cellular response to glucose stimulus / trans-Golgi network membrane / negative regulation of inflammatory response to antigenic stimulus / response to prostaglandin E / clathrin-coated endocytic vesicle membrane / bone development / platelet aggregation / cognition / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / G-protein beta/gamma-subunit complex binding / Olfactory Signaling Pathway / Activation of the phototransduction cascade / G beta:gamma signalling through PLC beta / Presynaptic function of Kainate receptors / Thromboxane signalling through TP receptor / G protein-coupled acetylcholine receptor signaling pathway / Activation of G protein gated Potassium channels / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / G-protein activation / positive regulation of insulin secretion / G beta:gamma signalling through CDC42 / Prostacyclin signalling through prostacyclin receptor / Glucagon signaling in metabolic regulation / G beta:gamma signalling through BTK / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / ADP signalling through P2Y purinoceptor 12 / photoreceptor disc membrane / sensory perception of smell / Glucagon-type ligand receptors / Sensory perception of sweet, bitter, and umami (glutamate) taste / Adrenaline,noradrenaline inhibits insulin secretion / Vasopressin regulates renal water homeostasis via Aquaporins / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / G alpha (z) signalling events / ADP signalling through P2Y purinoceptor 1 / ADORA2B mediated anti-inflammatory cytokines production / cellular response to catecholamine stimulus / G beta:gamma signalling through PI3Kgamma / adenylate cyclase-activating dopamine receptor signaling pathway / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / Cargo recognition for clathrin-mediated endocytosis / GPER1 signaling / cellular response to prostaglandin E stimulus / G-protein beta-subunit binding / heterotrimeric G-protein complex / G alpha (12/13) signalling events / Inactivation, recovery and regulation of the phototransduction cascade / positive regulation of cold-induced thermogenesis / extracellular vesicle / sensory perception of taste / Clathrin-mediated endocytosis / adenylate cyclase-activating G protein-coupled receptor signaling pathway / Thrombin signalling through proteinase activated receptors (PARs) / signaling receptor complex adaptor activity / retina development in camera-type eye / G protein activity / GTPase binding / fibroblast proliferation / Ca2+ pathway / High laminar flow shear stress activates signaling by PIEZO1 and PECAM1:CDH5:KDR in endothelial cells / G alpha (i) signalling events / G alpha (s) signalling events / phospholipase C-activating G protein-coupled receptor signaling pathway / G alpha (q) signalling events / Hydrolases; Acting on acid anhydrides; Acting on GTP to facilitate cellular and subcellular movement / Ras protein signal transduction / cell population proliferation / Extra-nuclear estrogen signaling / endosome
Similarity search - Function
Vasopressin V2 receptor / Vasopressin receptor / G-protein alpha subunit, group S / 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. ...Vasopressin V2 receptor / Vasopressin receptor / G-protein alpha subunit, group S / 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 / G protein beta WD-40 repeat protein / Guanine nucleotide-binding protein, beta subunit / G-protein, beta subunit / G-protein coupled receptors family 1 signature. / 7 transmembrane receptor (rhodopsin family) / G protein-coupled receptor, rhodopsin-like / GPCR, rhodopsin-like, 7TM / G-protein coupled receptors family 1 profile. / 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
Vasopressin V2 receptor / 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
Similarity search - Component
Biological speciesHomo sapiens (human)
Lama glama (llama)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.4 Å
AuthorsBous, J. / Mouillac, B. / Bron, P. / Granier, S. / Floquet, N. / Leyrat, C.
Funding support France, 2items
OrganizationGrant numberCountry
Foundation for Medical Research (France)DEQ20150331736 France
Agence Nationale de la Recherche (ANR)ANR-19-CE11-0014 France
CitationJournal: Sci Adv / Year: 2021
Title: Cryo-electron microscopy structure of the antidiuretic hormone arginine-vasopressin V2 receptor signaling complex.
Authors: Julien Bous / Hélène Orcel / Nicolas Floquet / Cédric Leyrat / Joséphine Lai-Kee-Him / Gérald Gaibelet / Aurélie Ancelin / Julie Saint-Paul / Stefano Trapani / Maxime Louet / Rémy ...Authors: Julien Bous / Hélène Orcel / Nicolas Floquet / Cédric Leyrat / Joséphine Lai-Kee-Him / Gérald Gaibelet / Aurélie Ancelin / Julie Saint-Paul / Stefano Trapani / Maxime Louet / Rémy Sounier / Hélène Déméné / Sébastien Granier / Patrick Bron / Bernard Mouillac /
Abstract: The antidiuretic hormone arginine-vasopressin (AVP) forms a signaling complex with the V2 receptor (V2R) and the G protein, promoting kidney water reabsorption. Molecular mechanisms underlying ...The antidiuretic hormone arginine-vasopressin (AVP) forms a signaling complex with the V2 receptor (V2R) and the G protein, promoting kidney water reabsorption. Molecular mechanisms underlying activation of this critical G protein-coupled receptor (GPCR) signaling system are still unknown. To fill this gap of knowledge, we report here the cryo-electron microscopy structure of the AVP-V2R-G complex. Single-particle analysis revealed the presence of three different states. The two best maps were combined with computational and nuclear magnetic resonance spectroscopy constraints to reconstruct two structures of the ternary complex. These structures differ in AVP and G binding modes. They reveal an original receptor-G interface in which the Gα subunit penetrates deep into the active V2R. The structures help to explain how V2R R137H or R137L/C variants can lead to two severe genetic diseases. Our study provides important structural insights into the function of this clinically relevant GPCR signaling complex.
History
DepositionDec 17, 2020Deposition site: PDBE / Processing site: PDBE
Revision 1.0Jun 2, 2021Provider: repository / Type: Initial release
Revision 1.1Jun 30, 2021Group: Structure summary / Category: audit_author
Revision 1.2Nov 13, 2024Group: Data collection / Database references / Structure summary
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / em_admin / pdbx_entry_details / pdbx_modification_feature
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession ..._database_2.pdbx_DOI / _database_2.pdbx_database_accession / _em_admin.last_update / _pdbx_entry_details.has_protein_modification

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Structure visualization

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Assembly

Deposited unit
A: Vasopressin V2 receptor
C: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
E: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
F: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
G: Nanobody 35
H: Vasopressin


Theoretical massNumber of molelcules
Total (without water)158,2076
Polymers158,2076
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area15080 Å2
ΔGint-78 kcal/mol
Surface area45620 Å2
MethodPISA

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Components

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Guanine nucleotide-binding protein ... , 3 types, 3 molecules CEF

#2: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Transducin beta chain 1


Mass: 40507.102 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNB1 / Plasmid: bacmid / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P62873
#3: Protein Guanine nucleotide-binding protein G(s) subunit alpha isoforms short / Adenylate cyclase-stimulating G alpha protein


Mass: 45725.520 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: GNAS, GNAS1, GSP / Plasmid: bacmid / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P63092
#4: 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 / Plasmid: bacmid / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P59768

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Protein / Antibody / Protein/peptide , 3 types, 3 molecules AGH

#1: Protein Vasopressin V2 receptor / V2R / AVPR V2 / Antidiuretic hormone receptor / Renal-type arginine vasopressin receptor


Mass: 47886.191 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Homo sapiens (human) / Gene: AVPR2, ADHR, DIR, DIR3, V2R / Plasmid: bacmid / Cell line (production host): Sf9 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P30518
#5: Antibody Nanobody 35


Mass: 15140.742 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Lama glama (llama) / Production host: Escherichia coli BL21(DE3) (bacteria)
#6: Protein/peptide Vasopressin


Mass: 1086.248 Da / Num. of mol.: 1 / Source method: obtained synthetically / Details: glycinamide c-terminal / Source: (synth.) Homo sapiens (human)

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Details

Has ligand of interestN
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

Component
IDNameTypeEntity IDParent-IDSource
1AVP-V2R-Galphas-beta1-gamma2-Nb35(T state)COMPLEXall0MULTIPLE SOURCES
2V2R and Guanine nucleotide-binding protein G(I)/G(S)/G(T)COMPLEX#1-#41RECOMBINANT
3Nanobody 35COMPLEX#51RECOMBINANT
4VasopressinCOMPLEX#61RECOMBINANT
Molecular weightValue: 0.156 MDa
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Homo sapiens (human)9606
23Lama glama (llama)9844
34Homo sapiens (human)9606
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
12Spodoptera frugiperda (fall armyworm)7108
23Escherichia coli BL21(DE3) (bacteria)469008
34synthetic construct (others)32630
Buffer solutionpH: 7.5
Buffer component
IDConc.UnitsFormulaBuffer-ID
1100 mMNaCl1
220 mMHepes1
30.0011 %LMNG1
40.001 %GDN1
5%CHS1
610 microMAVP1
70.001 %A8-351
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK III / 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 / Cs: 2.7 mm
Image recording
IDImaging-IDElectron dose (e/Å2)Detector modeFilm or detector model
1141.19COUNTINGGATAN K2 SUMMIT (4k x 4k)
2141.19COUNTINGGATAN K2 SUMMIT (4k x 4k)

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Processing

Software
NameVersionClassificationNB
phenix.real_space_refine1.18.2_3874+SVNrefinement
PHENIX1.18.2_3874+SVNrefinement
EM software
IDNameVersionCategory
1RELION3particle selection
4GctfCTF correction
10RELION3.1initial Euler assignment
11RELION3.1final Euler assignment
12RELION3.1classification
13RELION3.13D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 3566007
3D reconstructionResolution: 4.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 420953 / Symmetry type: POINT
RefinementCross valid method: NONE
Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2
Displacement parametersBiso mean: 72.14 Å2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00688206
ELECTRON MICROSCOPYf_angle_d1.010311125
ELECTRON MICROSCOPYf_chiral_restr0.05561246
ELECTRON MICROSCOPYf_plane_restr0.00651430
ELECTRON MICROSCOPYf_dihedral_angle_d24.82821119

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