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- EMDB-8623: Volta phase plate cryo-electron microscopy structure of a calcito... -

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Entry
Database: EMDB / ID: 8623
TitleVolta phase plate cryo-electron microscopy structure of a calcitonin receptor-heterotrimeric Gs protein complex
Map data
SampleComplex of a full-length, active-state calcitonin receptor with peptide ligand, heterotrimeric Gs protein and nano body 35.
  • (Guanine nucleotide-binding protein ...) x 3
  • NANOBODY 35Single-domain antibody
  • Calcitonin receptor
Function / homologyADP signalling through P2Y purinoceptor 12 / WD40/YVTN repeat-like-containing domain superfamily / G alpha (s) signalling events / 7 transmembrane receptor (Secretin family) / WD40-repeat-containing domain superfamily / G-protein gamma-like domain superfamily / P-loop containing nucleoside triphosphate hydrolase / G beta:gamma signalling through PLC beta / G-protein beta WD-40 repeat / WD40 repeat, conserved site ...ADP signalling through P2Y purinoceptor 12 / WD40/YVTN repeat-like-containing domain superfamily / G alpha (s) signalling events / 7 transmembrane receptor (Secretin family) / WD40-repeat-containing domain superfamily / G-protein gamma-like domain superfamily / P-loop containing nucleoside triphosphate hydrolase / G beta:gamma signalling through PLC beta / G-protein beta WD-40 repeat / WD40 repeat, conserved site / WD40-repeat-containing domain / GPCR, family 2, secretin-like, conserved site / G alpha (12/13) signalling events / G alpha (q) signalling events / GPCR, family 2-like / Guanine nucleotide-binding protein, beta subunit / G-protein gamma-like domain / G alpha (i) signalling events / G protein alpha subunit, helical insertion / GPCR, family 2, calcitonin receptor family / GPCR, family 2, extracellular hormone receptor domain / Ca2+ pathway / G-protein, gamma subunit / GPCR, family 2, calcitonin receptor / G beta:gamma signalling through PI3Kgamma / Adrenaline,noradrenaline inhibits insulin secretion / WD40 repeat / Prostacyclin signalling through prostacyclin receptor / G-protein, beta subunit / Guanine nucleotide binding protein (G-protein), alpha subunit / GPCR, family 2, secretin-like / G-protein alpha subunit, group S / ADP signalling through P2Y purinoceptor 1 / GPCR family 2, extracellular hormone receptor domain superfamily / WD domain, G-beta repeat / G-protein activation / G alpha (z) signalling events / Vasopressin regulates renal water homeostasis via Aquaporins / Thrombin signalling through proteinase activated receptors (PARs) / PKA activation in glucagon signalling / Presynaptic function of Kainate receptors / Hedgehog 'off' state / Cooperation of PDCL (PhLP1) and TRiC/CCT in G-protein beta folding / Activation of G protein gated Potassium channels / Activation of the phototransduction cascade / G beta:gamma signalling through BTK / Inactivation, recovery and regulation of the phototransduction cascade / G beta:gamma signalling through CDC42 / Inhibition of voltage gated Ca2+ channels via Gbeta/gamma subunits / Glucagon-like Peptide-1 (GLP1) regulates insulin secretion / Olfactory Signaling Pathway / Glucagon signaling in metabolic regulation / Thromboxane signalling through TP receptor / Calcitonin-like ligand receptors / G-protein coupled receptors family 2 signature 1. / G-protein alpha subunit / G-protein coupled receptors family 2 profile 2. / G-protein coupled receptors family 2 profile 1. / Trp-Asp (WD) repeats profile. / G-protein gamma subunit domain profile. / Trp-Asp (WD) repeats signature. / GGL domain / G-protein coupled receptors family 2 signature 2. / Hormone receptor domain / Glucagon-type ligand receptors / Trp-Asp (WD) repeats circular profile. / calcitonin binding / amylin receptor complex 3 / cross-receptor inhibition within G protein-coupled receptor heterodimer / amylin receptor complex 2 / amylin receptor complex 1 / amylin receptor activity / calcitonin receptor activity / calcitonin gene-related peptide receptor activity / positive regulation of calcium ion import across plasma membrane / amylin receptor signaling pathway / positive regulation of adenylate cyclase activity / intracellular transport / hair follicle placode formation / intrinsic component of membrane / sensory perception of taste / cellular response to catecholamine stimulus / cardiac muscle cell apoptotic process / G protein-coupled acetylcholine receptor signaling pathway / adenylate cyclase-activating adrenergic receptor signaling pathway / G-protein beta-subunit binding / protein heterotrimerization / alkylglycerophosphoethanolamine phosphodiesterase activity / adenylate cyclase-activating dopamine receptor signaling pathway / rhodopsin mediated signaling pathway / photoreceptor outer segment membrane / positive regulation of cAMP-mediated signaling / sensory perception of smell / developmental growth / bone development / G-protein beta/gamma-subunit complex binding / spectrin binding / photoreceptor disc membrane / response to amyloid-beta / heterotrimeric G-protein complex
Function and homology information
SourceHomo sapiens (human) / Lama Glama (llama)
Methodsingle particle reconstruction / cryo EM / 4.1 Å resolution
AuthorsLiang YL / Khoshouei M / Radjainia M / Zhang Y / Glukhova A / Tarrasch J / Thal DM / Furness SGB / Christopoulos G / Coudrat T / Danev R / Baumeister W / Miller LJ / Christopoulos A / Kobilka BK / Wootten D / Skiniotis G / Sexton PM
CitationJournal: Nature / Year: 2017
Title: Phase-plate cryo-EM structure of a class B GPCR-G-protein complex.
Authors: Yi-Lynn Liang / Maryam Khoshouei / Mazdak Radjainia / Yan Zhang / Alisa Glukhova / Jeffrey Tarrasch / David M Thal / Sebastian G B Furness / George Christopoulos / Thomas Coudrat / Radostin Danev / Wolfgang Baumeister / Laurence J Miller / Arthur Christopoulos / Brian K Kobilka / Denise Wootten / Georgios Skiniotis / Patrick M Sexton
Validation ReportPDB-ID: 5uz7

SummaryFull reportAbout validation report
DateDeposition: Feb 24, 2017 / Header (metadata) release: Mar 29, 2017 / Map release: May 3, 2017 / Last update: Jul 18, 2018

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.04
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-5uz7
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_8623.map.gz (map file in CCP4 format, 32001 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
200 pix
1.06 Å/pix.
= 212. Å
200 pix
1.06 Å/pix.
= 212. Å
200 pix
1.06 Å/pix.
= 212. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.06 Å
Density
Contour Level:0.05 (by author), 0.04 (movie #1):
Minimum - Maximum-0.14220421 - 0.2546117
Average (Standard dev.)0.00021038976 (0.008024996)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions200200200
Origin0.0.0.
Limit199.199.199.
Spacing200200200
CellA=B=C: 211.99998 Å
α=β=γ: 90.0 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.061.061.06
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z212.000212.000212.000
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.1420.2550.000

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Supplemental data

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Sample components

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Entire Complex of a full-length, active-state calcitonin receptor with p...

EntireName: Complex of a full-length, active-state calcitonin receptor with peptide ligand, heterotrimeric Gs protein and nano body 35.
Number of components: 6

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Component #1: protein, Complex of a full-length, active-state calcitonin recept...

ProteinName: Complex of a full-length, active-state calcitonin receptor with peptide ligand, heterotrimeric Gs protein and nano body 35.
Recombinant expression: No
MassTheoretical: 150 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #2: protein, Guanine nucleotide-binding protein G(s) subunit alpha is...

ProteinName: Guanine nucleotide-binding protein G(s) subunit alpha isoforms short
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 44.32616 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #3: protein, Guanine nucleotide-binding protein G(I)/G(S)/G(T) subuni...

ProteinName: Guanine nucleotide-binding protein G(I)/G(S)/G(T) subunit beta-1
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 38.744371 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #4: protein, Guanine nucleotide-binding protein G(I)/G(S)/G(O) subuni...

ProteinName: Guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 7.56375 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Component #5: protein, NANOBODY 35

ProteinName: NANOBODY 35Single-domain antibody / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 15.140742 kDa
SourceSpecies: Lama Glama (llama)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #6: protein, Calcitonin receptor

ProteinName: Calcitonin receptor / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 58.469594 kDa
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Trichoplusia ni (cabbage looper)

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.3 mg/ml / pH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 5 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 106838
3D reconstructionResolution: 4.1 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Output model

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