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- PDB-9asb: Structure of human calcium-sensing receptor in complex with chime... -

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Basic information

Entry
Database: PDB / ID: 9asb
TitleStructure of human calcium-sensing receptor in complex with chimeric Gq (miniGisq) protein in nanodiscs
Components
  • (Chimeric mini guanine nucleotide-binding protein G(i)(s)(q) subunit ...) x 2
  • Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
  • Isoform 1 of Extracellular calcium-sensing receptor
KeywordsMEMBRANE PROTEIN / Calcium-sensing receptor / G-protein-coupled receptor / G protein / signal transduction
Function / homology
Function and homology information


bile acid secretion / chemosensory behavior / cellular response to peptide / response to fibroblast growth factor / cellular response to vitamin D / phosphatidylinositol phospholipase C activity / Class C/3 (Metabotropic glutamate/pheromone receptors) / calcium ion import / positive regulation of positive chemotaxis / fat pad development ...bile acid secretion / chemosensory behavior / cellular response to peptide / response to fibroblast growth factor / cellular response to vitamin D / phosphatidylinositol phospholipase C activity / Class C/3 (Metabotropic glutamate/pheromone receptors) / calcium ion import / positive regulation of positive chemotaxis / fat pad development / amino acid binding / cellular response to hepatocyte growth factor stimulus / branching morphogenesis of an epithelial tube / positive regulation of calcium ion import / regulation of calcium ion transport / cellular response to low-density lipoprotein particle stimulus / Adenylate cyclase inhibitory pathway / detection of calcium ion / positive regulation of protein localization to cell cortex / anatomical structure morphogenesis / regulation of cAMP-mediated signaling / D2 dopamine receptor binding / G protein-coupled serotonin receptor binding / axon terminus / regulation of mitotic spindle organization / cellular response to forskolin / positive regulation of vasoconstriction / JNK cascade / chloride transmembrane transport / adenylate cyclase-inhibiting G protein-coupled receptor signaling pathway / ossification / Regulation of insulin secretion / G protein-coupled receptor binding / G protein-coupled receptor activity / response to ischemia / cellular response to glucose stimulus / 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 / adenylate cyclase-modulating G protein-coupled receptor signaling pathway / 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 / intracellular calcium ion homeostasis / ADP signalling through P2Y purinoceptor 12 / positive regulation of insulin secretion / G beta:gamma signalling through BTK / Sensory perception of sweet, bitter, and umami (glutamate) taste / response to peptide hormone / Synthesis, secretion, and inactivation of Glucagon-like Peptide-1 (GLP-1) / photoreceptor disc membrane / Adrenaline,noradrenaline inhibits insulin secretion / Glucagon-type ligand receptors / Vasopressin regulates renal water homeostasis via Aquaporins / G alpha (z) signalling events / cellular response to catecholamine stimulus / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / ADORA2B mediated anti-inflammatory cytokines production / vasodilation / 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 / GDP binding / G-protein beta-subunit binding / heterotrimeric G-protein complex / Inactivation, recovery and regulation of the phototransduction cascade / extracellular vesicle / G alpha (12/13) signalling events / signaling receptor complex adaptor activity / Thrombin signalling through proteinase activated receptors (PARs) / integrin binding / retina development in camera-type eye / GTPase binding / Ca2+ pathway / phospholipase C-activating G protein-coupled receptor signaling pathway / cell cortex / midbody / G alpha (i) signalling events / fibroblast proliferation / cellular response to hypoxia / G alpha (s) signalling events / G alpha (q) signalling events / basolateral plasma membrane / Ras protein signal transduction / cell population proliferation / transmembrane transporter binding / Extra-nuclear estrogen signaling / positive regulation of ERK1 and ERK2 cascade / cell cycle
Similarity search - Function
GPCR, family 3, extracellular calcium-sensing receptor-related / G-protein coupled receptors family 3 signature 1. / G-protein coupled receptors family 3 signature 2. / GPCR, family 3, nine cysteines domain / GPCR, family 3, nine cysteines domain superfamily / Nine Cysteines Domain of family 3 GPCR / GPCR, family 3, conserved site / G-protein coupled receptors family 3 signature 3. / GPCR, family 3 / GPCR family 3, C-terminal ...GPCR, family 3, extracellular calcium-sensing receptor-related / G-protein coupled receptors family 3 signature 1. / G-protein coupled receptors family 3 signature 2. / GPCR, family 3, nine cysteines domain / GPCR, family 3, nine cysteines domain superfamily / Nine Cysteines Domain of family 3 GPCR / GPCR, family 3, conserved site / G-protein coupled receptors family 3 signature 3. / GPCR, family 3 / GPCR family 3, C-terminal / 7 transmembrane sweet-taste receptor of 3 GCPR / G-protein coupled receptors family 3 profile. / G-protein alpha subunit, group S / G-protein alpha subunit, group I / Receptor, ligand binding region / Receptor family ligand binding region / 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 / Periplasmic binding protein-like I / 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 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
Chem-9IG / : / PHOSPHATE ION / CYCLOMETHYLTRYPTOPHAN / CHOLESTEROL HEMISUCCINATE / GNAS complex locus / Extracellular calcium-sensing 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(i) subunit alpha-1
Similarity search - Component
Biological speciesHomo sapiens (human)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsZuo, H. / Park, J. / Frangaj, A. / Ye, J. / Lu, G. / Manning, J.J. / Asher, W.B. / Lu, Z. / Hu, G. / Wang, L. ...Zuo, H. / Park, J. / Frangaj, A. / Ye, J. / Lu, G. / Manning, J.J. / Asher, W.B. / Lu, Z. / Hu, G. / Wang, L. / Mendez, J. / Eng, E. / Zhang, Z. / Lin, X. / Grasucci, R. / Hendrickson, W.A. / Clarke, O.B. / Javitch, J.A. / Conigrave, A.D. / Fan, Q.R.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35GM141871 United States
CitationJournal: Nature / Year: 2024
Title: Promiscuous G-protein activation by the calcium-sensing receptor
Authors: Zuo, H. / Park, J. / Frangaj, A. / Ye, J. / Lu, G. / Manning, J.J. / Asher, W.B. / Lu, Z. / Hu, G. / Wang, L. / Mendez, J. / Eng, E. / Zhang, Z. / Lin, X. / Grasucci, R. / Hendrickson, W.A. ...Authors: Zuo, H. / Park, J. / Frangaj, A. / Ye, J. / Lu, G. / Manning, J.J. / Asher, W.B. / Lu, Z. / Hu, G. / Wang, L. / Mendez, J. / Eng, E. / Zhang, Z. / Lin, X. / Grasucci, R. / Hendrickson, W.A. / Clarke, O.B. / Javitch, J.A. / Conigrave, A.D. / Fan, Q.R.
History
DepositionFeb 24, 2024Deposition site: RCSB / Processing site: RCSB
Revision 1.0Apr 17, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
Q: Isoform 1 of Extracellular calcium-sensing receptor
R: Isoform 1 of Extracellular calcium-sensing receptor
A: Chimeric mini guanine nucleotide-binding protein G(i)(s)(q) subunit alpha
B: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
G: Chimeric mini guanine nucleotide-binding protein G(i)(s)(q) subunit alpha
hetero molecules


Theoretical massNumber of molelcules
Total (without water)286,04229
Polymers280,3105
Non-polymers5,73124
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Protein , 2 types, 3 molecules QRB

#1: Protein Isoform 1 of Extracellular calcium-sensing receptor / CaR / CaSR / hCasR / Parathyroid cell calcium-sensing receptor 1 / PCaR1


Mass: 102864.617 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Details: The CaSR construct consists of residues 1-903 and a Flag tag inserted after the signal peptide
Source: (gene. exp.) Homo sapiens (human) / Gene: CASR, GPRC2A, PCAR1 / Cell line (production host): HEK293 GnTI- / Production host: Homo sapiens (human) / References: UniProt: P41180
#3: Protein Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 / Transducin beta chain 1


Mass: 38413.895 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: The guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1 construct contains a Flag epitope fused to the N-terminus.
Source: (gene. exp.) Homo sapiens (human) / Gene: GNB1 / Cell line (production host): HEK293 GnTI- / Production host: Homo sapiens (human) / References: UniProt: P62873

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Chimeric mini guanine nucleotide-binding protein G(i)(s)(q) subunit ... , 2 types, 2 molecules AG

#2: Protein Chimeric mini guanine nucleotide-binding protein G(i)(s)(q) subunit alpha / Adenylate cyclase-inhibiting G alpha protein


Mass: 28306.148 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: MiniG(q) alpha is a chimeric protein engineered based on the miniG(s) alpha skeleton [miniG(s)(q) alpha]. Chimeric miniG(i)(s)(q) alpha construct was designed where the N-terminal 25 amino ...Details: MiniG(q) alpha is a chimeric protein engineered based on the miniG(s) alpha skeleton [miniG(s)(q) alpha]. Chimeric miniG(i)(s)(q) alpha construct was designed where the N-terminal 25 amino acids of G(s) alpha was replaced with the initial 18 residues of G(i1) alpha
Source: (gene. exp.) Homo sapiens (human) / Gene: GNAI1, GNAS / Cell line (production host): HEK293 GnTl- / Production host: Homo sapiens (human) / References: UniProt: P63096, UniProt: A0A590UJY2
#4: Protein Chimeric mini guanine nucleotide-binding protein G(i)(s)(q) subunit alpha / 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 / Cell line (production host): HEK293 GnTI- / Production host: Homo sapiens (human) / References: UniProt: P59768

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Sugars , 2 types, 10 molecules

#5: Polysaccharide
2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose


Type: oligosaccharide / Mass: 424.401 Da / Num. of mol.: 4
Source method: isolated from a genetically manipulated source
DescriptorTypeProgram
DGlcpNAcb1-4DGlcpNAcb1-ROHGlycam Condensed SequenceGMML 1.0
WURCS=2.0/1,2,1/[a2122h-1b_1-5_2*NCC/3=O]/1-1/a4-b1WURCSPDB2Glycan 1.1.0
[][D-1-deoxy-GlcpNAc]{[(4+1)][b-D-GlcpNAc]{}}LINUCSPDB-CARE
#6: Sugar
ChemComp-NAG / 2-acetamido-2-deoxy-beta-D-glucopyranose / N-acetyl-beta-D-glucosamine / 2-acetamido-2-deoxy-beta-D-glucose / 2-acetamido-2-deoxy-D-glucose / 2-acetamido-2-deoxy-glucose / N-ACETYL-D-GLUCOSAMINE / N-Acetylglucosamine


Type: D-saccharide, beta linking / Mass: 221.208 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: C8H15NO6
IdentifierTypeProgram
DGlcpNAcbCONDENSED IUPAC CARBOHYDRATE SYMBOLGMML 1.0
N-acetyl-b-D-glucopyranosamineCOMMON NAMEGMML 1.0
b-D-GlcpNAcIUPAC CARBOHYDRATE SYMBOLPDB-CARE 1.0
GlcNAcSNFG CARBOHYDRATE SYMBOLGMML 1.0

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Non-polymers , 6 types, 14 molecules

#7: Chemical ChemComp-TCR / CYCLOMETHYLTRYPTOPHAN


Type: L-peptide linking / Mass: 216.236 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C12H12N2O2
#8: Chemical ChemComp-PO4 / PHOSPHATE ION / Phosphate


Mass: 94.971 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: PO4
#9: Chemical
ChemComp-CA / CALCIUM ION


Mass: 40.078 Da / Num. of mol.: 6 / Source method: obtained synthetically / Formula: Ca
#10: Chemical ChemComp-9IG / 3-(2-chlorophenyl)-N-[(1R)-1-(3-methoxyphenyl)ethyl]propan-1-amine


Mass: 303.826 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: C18H22ClNO
#11: Chemical ChemComp-Y01 / CHOLESTEROL HEMISUCCINATE


Mass: 486.726 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C31H50O4
#12: Chemical ChemComp-A1AF7 / (19R,22S,25R)-22,25,26-trihydroxy-16,22-dioxo-17,21,23-trioxa-22lambda~5~-phosphahexacosan-19-yl (9Z)-octadec-9-enoate


Mass: 749.007 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C40H77O10P

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Details

Has ligand of interestN

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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: Human CaSR in complex with chimeric Gq (miniGisq) protein
Type: COMPLEX / Entity ID: #1-#4 / Source: RECOMBINANT
Molecular weightValue: 0.276 MDa / Experimental value: NO
Source (natural)Organism: Homo sapiens (human)
Source (recombinant)Organism: Homo sapiens (human) / Cell: HEK293 GnTI-
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
2100 mMSodium chlorideNaClSodium chloride1
32 mMMagnesium chlorideMgCl21
410 mMCalcium chlorideCaCl21
520 uMTNCAC12H12N2O21
620 uMR568 HClC18H22ClNOHCl1
SpecimenConc.: 3.6 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R0.6/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K
Details: The sample was blotted for 6s before plunge-frozen.

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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 FIELDBright-field microscopy / Nominal magnification: 105000 X / Nominal defocus max: 1900 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / C2 aperture diameter: 100 µm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 100 K
Image recordingAverage exposure time: 3 sec. / Electron dose: 74.9 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 9336
EM imaging opticsEnergyfilter name: GIF Bioquantum / Energyfilter slit width: 15 eV
Image scansWidth: 11520 / Height: 8184

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Processing

EM software
IDNameVersionCategory
1cryoSPARC3.3.2particle selection
2EPUimage acquisition
4cryoSPARC3.3.2CTF correction
7Coot0.9.8.1model fitting
9cryoSPARC3.3.2initial Euler assignment
10cryoSPARC3.3.2final Euler assignment
12cryoSPARC3.3.23D reconstruction
13PHENIX1.19.2_4158:model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1339334
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 91042 / Symmetry type: POINT
Atomic model buildingProtocol: FLEXIBLE FIT / Space: REAL
Atomic model building

3D fitting-ID: 1 / Source name: PDB / Type: experimental model

IDPDB-IDPdb chain-IDAccession codeChain-IDInitial refinement model-ID
17SIL

7sil

Q7SILQ1
27SIL

7sil

R7SILR1
37P00

7p00

A7P00A2
47P00

7p00

B7P00B2
57P00

7p00

G7P00G2
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00418605
ELECTRON MICROSCOPYf_angle_d0.59525212
ELECTRON MICROSCOPYf_dihedral_angle_d13.4186691
ELECTRON MICROSCOPYf_chiral_restr0.0422831
ELECTRON MICROSCOPYf_plane_restr0.0053188

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