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Yorodumi- PDB-6niy: A high-resolution cryo-electron microscopy structure of a calcito... -
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-Basic information
Entry | Database: PDB / ID: 6niy | |||||||||||||||
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Title | A high-resolution cryo-electron microscopy structure of a calcitonin receptor-heterotrimeric Gs protein complex | |||||||||||||||
Components |
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Keywords | MEMBRANE PROTEIN / GPCR / transmembrane / receptor / calcitonin | |||||||||||||||
Function / homology | Function and homology information calcitonin binding / amylin receptor complex 1 / amylin receptor complex 2 / cross-receptor inhibition within G protein-coupled receptor heterodimer / amylin receptor complex 3 / amylin receptor activity / calcitonin receptor activity / calcitonin gene-related peptide receptor activity / amylin receptor signaling pathway / positive regulation of adenylate cyclase activity ...calcitonin binding / amylin receptor complex 1 / amylin receptor complex 2 / cross-receptor inhibition within G protein-coupled receptor heterodimer / amylin receptor complex 3 / amylin receptor activity / calcitonin receptor activity / calcitonin gene-related peptide receptor activity / amylin receptor signaling pathway / positive regulation of adenylate cyclase activity / Calcitonin-like ligand receptors / negative regulation of ossification / positive regulation of protein kinase A signaling / PKA activation in glucagon signalling / response to amyloid-beta / hair follicle placode formation / intracellular transport / D1 dopamine receptor binding / developmental growth / Hedgehog 'off' state / positive regulation of cAMP-mediated signaling / positive regulation of calcium-mediated signaling / adenylate cyclase-activating adrenergic receptor signaling pathway / response to glucocorticoid / regulation of mRNA stability / activation of adenylate cyclase activity / adenylate cyclase activator activity / ossification / osteoclast differentiation / acrosomal vesicle / trans-Golgi network membrane / 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 / 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 / cilium / 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 / hormone activity / 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 / platelet aggregation / cognition / Glucagon-type ligand receptors / 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 / extracellular vesicle / G alpha (12/13) signalling events / 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 / amyloid-beta binding / positive regulation of cold-induced thermogenesis / positive regulation of cytosolic calcium ion concentration / 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 / positive regulation of ERK1 and ERK2 cascade / cell surface receptor signaling pathway / positive regulation of phosphatidylinositol 3-kinase/protein kinase B signal transduction / G protein-coupled receptor signaling pathway / lysosomal membrane / axon / GTPase activity / synapse Similarity search - Function | |||||||||||||||
Biological species | Homo sapiens (human) Lama glama (llama) Oncorhynchus sp. (fish) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.34 Å | |||||||||||||||
Authors | dal Maso, E. / Glukhova, A. / Zhu, Y. / Garcia-Nafria, J. / Tate, C.G. / Atanasio, S. / Reynolds, C.A. / Ramirez-Aportela, E. / Carazo, J.-M. / Hick, C.A. ...dal Maso, E. / Glukhova, A. / Zhu, Y. / Garcia-Nafria, J. / Tate, C.G. / Atanasio, S. / Reynolds, C.A. / Ramirez-Aportela, E. / Carazo, J.-M. / Hick, C.A. / Furness, S.G.B. / Hay, D.L. / Liang, Y.-L. / Miller, L.J. / Christopoulos, A. / Wang, M.-W. / Wootten, D. / Sexton, P.M. | |||||||||||||||
Funding support | Australia, 4items
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Citation | Journal: ACS Pharmacol Transl Sci / Year: 2019 Title: The Molecular Control of Calcitonin Receptor Signaling. Authors: Emma Dal Maso / Alisa Glukhova / Yue Zhu / Javier Garcia-Nafria / Christopher G Tate / Silvia Atanasio / Christopher A Reynolds / Erney Ramírez-Aportela / Jose-Maria Carazo / Caroline A ...Authors: Emma Dal Maso / Alisa Glukhova / Yue Zhu / Javier Garcia-Nafria / Christopher G Tate / Silvia Atanasio / Christopher A Reynolds / Erney Ramírez-Aportela / Jose-Maria Carazo / Caroline A Hick / Sebastian G B Furness / Debbie L Hay / Yi-Lynn Liang / Laurence J Miller / Arthur Christopoulos / Ming-Wei Wang / Denise Wootten / Patrick M Sexton / Abstract: The calcitonin receptor (CTR) is a class B G protein-coupled receptor (GPCR) that responds to the peptide hormone calcitonin (CT). CTs are clinically approved for the treatment of bone diseases. We ...The calcitonin receptor (CTR) is a class B G protein-coupled receptor (GPCR) that responds to the peptide hormone calcitonin (CT). CTs are clinically approved for the treatment of bone diseases. We previously reported a 4.1 Å structure of the activated CTR bound to salmon CT (sCT) and heterotrimeric Gs protein by cryo-electron microscopy (Liang, Y.-L., . Phase-plate cryo- EM structure of a class B GPCR-G protein complex. , , 118-123). In the current study, we have reprocessed the electron micrographs to yield a 3.3 Å map of the complex. This has allowed us to model extracellular loops (ECLs) 2 and 3, and the peptide N-terminus that previously could not be resolved. We have also performed alanine scanning mutagenesis of ECL1 and the upper segment of transmembrane helix 1 (TM1) and its extension into the receptor extracellular domain (TM1 stalk), with effects on peptide binding and function assessed by cAMP accumulation and ERK1/2 phosphorylation. These data were combined with previously published alanine scanning mutagenesis of ECL2 and ECL3 and the new structural information to provide a comprehensive 3D map of the molecular surface of the CTR that controls binding and signaling of distinct CT and related peptides. The work highlights distinctions in how different, related, class B receptors may be activated. The new mutational data on the TM1 stalk and ECL1 have also provided critical insights into the divergent control of cAMP versus pERK signaling and, collectively with previous mutagenesis data, offer evidence that the conformations linked to these different signaling pathways are, in many ways, mutually exclusive. This study furthers our understanding of the complex nature of signaling elicited by GPCRs and, in particular, that of the therapeutically important class B subfamily. | |||||||||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 6niy.cif.gz | 192.4 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6niy.ent.gz | 147.3 KB | Display | PDB format |
PDBx/mmJSON format | 6niy.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/ni/6niy ftp://data.pdbj.org/pub/pdb/validation_reports/ni/6niy | HTTPS FTP |
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-Related structure data
Related structure data | 9382MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-Guanine nucleotide-binding protein ... , 3 types, 3 molecules ABG
#1: 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 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P63092 |
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#4: Protein | Mass: 37416.930 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: GNB1 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P62873 |
#5: Protein | 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: Trichoplusia ni (cabbage looper) / References: UniProt: P59768 |
-Antibody / Protein / Protein/peptide , 3 types, 3 molecules NRP
#2: Antibody | 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 (E. coli) |
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#3: Protein | Mass: 55387.316 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Homo sapiens (human) / Gene: CALCR / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P30988 |
#6: Protein/peptide | Mass: 3438.886 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Oncorhynchus sp. (fish) / References: UniProt: Q92163 |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Complex of a full-length, active-state calcitonin receptor with peptide ligand, heterotrimeric Gs protein and nanobody 35 Type: COMPLEX / Entity ID: all / Source: RECOMBINANT |
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Molecular weight | Value: 0.15 MDa / Experimental value: NO |
Source (natural) | Organism: Homo sapiens (human) |
Source (recombinant) | Organism: Trichoplusia ni (cabbage looper) |
Buffer solution | pH: 7.5 |
Specimen | Conc.: 0.3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Calibrated magnification: 47170 X / Cs: 2.7 mm / C2 aperture diameter: 50 µm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 50 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) |
EM imaging optics | Phase plate: OTHER |
-Processing
Software |
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EM software |
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CTF correction | Details: Phase Plate CTF correction / Type: NONE | ||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 1445614 | ||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.34 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 417949 / Symmetry type: POINT | ||||||||||||||||||||||||||||||||
Atomic model building | Space: REAL | ||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 5UZ7 | ||||||||||||||||||||||||||||||||
Refinement | Stereochemistry target values: CDL v1.2 | ||||||||||||||||||||||||||||||||
Refine LS restraints |
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