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- EMDB-5867: Single-particle reconstruction of conformation VII of ligand-free sGC -

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Basic information

Entry
Database: EMDB / ID: 5867
TitleSingle-particle reconstruction of conformation VII of ligand-free sGC
Map dataSingle-particle reconstruction of conformation VII of ligand-free sGC
SampleSoluble Guanylate Cyclase, ligand-freeSoluble guanylyl cyclase
  • Soluble Guanylate CyclaseSoluble guanylyl cyclase
Keywordssoluble guanylate cyclase / conformational heterogeneity
SourceRattus norvegicus (Norway rat)
Methodsingle particle reconstruction / negative staining / 30 Å resolution
AuthorsCampbell MG / Underbakke ES / Potter CS / Carragher B / Marletta MA
CitationJournal: Proc. Natl. Acad. Sci. U.S.A. / Year: 2014
Title: Single-particle EM reveals the higher-order domain architecture of soluble guanylate cyclase.
Authors: Melody G Campbell / Eric S Underbakke / Clinton S Potter / Bridget Carragher / Michael A Marletta
DateDeposition: Jan 17, 2014 / Header (metadata) release: Mar 19, 2014 / Map release: Mar 19, 2014 / Last update: Mar 19, 2014

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.79
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.79
  • 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_5867.map.gz (map file in CCP4 format, 43905 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
224 pix
1.06 Å/pix.
= 237.44 Å
224 pix
1.06 Å/pix.
= 237.44 Å
224 pix
1.06 Å/pix.
= 237.44 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:0.790 (by author), 0.79 (movie #1):
Minimum - Maximum-1.34823692 - 3.32659340
Average (Standard dev.)-0.00239478 (0.25892913)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions224224224
Origin-112-112-112
Limit111111111
Spacing224224224
CellA=B=C: 237.43999 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/z224224224
origin x/y/z0.0000.0000.000
length x/y/z237.440237.440237.440
α/β/γ90.00090.00090.000
start NX/NY/NZ-95-75153
NX/NY/NZ200200200
MAP C/R/S123
start NC/NR/NS-112-112-112
NC/NR/NS224224224
D min/max/mean-1.3483.327-0.002

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Supplemental data

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Sample components

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Entire Soluble Guanylate Cyclase, ligand-free

EntireName: Soluble Guanylate Cyclase, ligand-free / Number of components: 1 / Oligomeric State: Heterodimer
MassTheoretical: 150 kDa / Experimental: 150 kDa

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Component #1: protein, Soluble Guanylate Cyclase

ProteinName: Soluble Guanylate CyclaseSoluble guanylyl cyclase / a.k.a: sGC / Oligomeric Details: Heterodimer / Recombinant expression: Yes / Number of Copies: 1
MassTheoretical: 150 kDa / Experimental: 150 kDa
SourceSpecies: Rattus norvegicus (Norway rat)
Source (engineered)Expression System: Spodoptera frugiperda (fall armyworm) / Vector: pFastBac1/sGCALPHA1 and pFastBac1/sGCBETA1 / Cell of expression system: Sf9

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: negative staining
Sample solutionBuffer solution: 50 mM TEA, 150 mM NaCl, 5 mM DTT / pH: 7.5
Support filmGlow discharged C-flat grid with 2-micron-diameter holes overlaid by thin 1.5 nm continuous carbon
Staining3 microliters of sample were applied to grid. The specimen was stained twice with 2% uranyl formate, then allowed to air-dry.
VitrificationInstrument: NONE / Cryogen name: NONE

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Electron microscopy imaging

Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company
ImagingMicroscope: FEI TECNAI F20 / Date: Jan 26, 2013
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Electron dose: 35 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 80000 X (nominal) / Cs: 2 mm / Imaging mode: BRIGHT FIELD / Defocus: 1200 - 2200 nm
Specimen HolderModel: SIDE ENTRY, EUCENTRIC / Tilt Angle: -55 - 0 deg. / Temperature: 298 K
CameraDetector: TVIPS TEMCAM-F416 (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 2204

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Image processing

ProcessingMethod: single particle reconstruction / Number of class averages: 1 / Applied symmetry: C1 (asymmetric) / Number of projections: 235 / Details: See publication
3D reconstructionAlgorithm: RCT / Software: SPIDER / CTF correction: Each micrograph / Resolution: 30 Å / Resolution method: FSC 0.5, semi-independent

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