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- EMDB-1750: Structure of E. coli Hibernating Ribosomes in 'f-f' Organization -

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Basic information

Entry
Database: EMDB / ID: EMD-1750
TitleStructure of E. coli Hibernating Ribosomes in 'f-f' Organization
Map dataThis is the density map of an E.coli 100S ribosome with 'f-f' organization obtained in vitro
Sample
  • Sample: 100S ribosomes in a 'f-f' 3D organization solved in vitro
  • Complex: 70S ribosomeRibosome
KeywordsRibosome / dimers / hybernating / 100S / RMF / HPF
Biological speciesEscherichia coli (E. coli)
Methodsubtomogram averaging / cryo EM / negative staining
AuthorsOrtiz JO / Brandt F / Valerio M / Sennels L / Rappsilber J / Scheres SHW / Eibauer M / Hartl FU / Baumeister W
CitationJournal: J Cell Biol / Year: 2010
Title: Structure of hibernating ribosomes studied by cryoelectron tomography in vitro and in situ.
Authors: Julio O Ortiz / Florian Brandt / Valério R F Matias / Lau Sennels / Juri Rappsilber / Sjors H W Scheres / Matthias Eibauer / F Ulrich Hartl / Wolfgang Baumeister /
Abstract: Ribosomes arranged in pairs (100S) have been related with nutritional stress response and are believed to represent a "hibernation state." Several proteins have been identified that are associated ...Ribosomes arranged in pairs (100S) have been related with nutritional stress response and are believed to represent a "hibernation state." Several proteins have been identified that are associated with 100S ribosomes but their spatial organization has hitherto not been characterized. We have used cryoelectron tomography to reveal the three-dimensional configuration of 100S ribosomes isolated from starved Escherichia coli cells and we have described their mode of interaction. In situ studies with intact E. coli cells allowed us to demonstrate that 100S ribosomes do exist in vivo and represent an easily reversible state of quiescence; they readily vanish when the growth medium is replenished.
History
DepositionJun 11, 2010-
Header (metadata) releaseJun 16, 2010-
Map releaseSep 2, 2010-
UpdateDec 11, 2013-
Current statusDec 11, 2013Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 2.9E-8
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 2.9E-8
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_1750.map.gz / Format: CCP4 / Size: 12.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationThis is the density map of an E.coli 100S ribosome with 'f-f' organization obtained in vitro
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
5.6 Å/pix.
x 150 pix.
= 840. Å
5.6 Å/pix.
x 150 pix.
= 840. Å
5.6 Å/pix.
x 150 pix.
= 840. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 5.6 Å
Density
Contour LevelBy AUTHOR: 0.0000000292
Minimum - Maximum-0.000000119888 - 0.00000017079
Average (Standard dev.)0.000000000555724 (±0.00000000881405)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions150150150
Spacing150150150
CellA=B=C: 840 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z5.65.65.6
M x/y/z150150150
origin x/y/z0.0000.0000.000
length x/y/z840.000840.000840.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-100-100-100
NX/NY/NZ200200200
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS150150150
D min/max/mean-0.0000.0000.000

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Supplemental data

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Sample components

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Entire : 100S ribosomes in a 'f-f' 3D organization solved in vitro

EntireName: 100S ribosomes in a 'f-f' 3D organization solved in vitro
Components
  • Sample: 100S ribosomes in a 'f-f' 3D organization solved in vitro
  • Complex: 70S ribosomeRibosome

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Supramolecule #1000: 100S ribosomes in a 'f-f' 3D organization solved in vitro

SupramoleculeName: 100S ribosomes in a 'f-f' 3D organization solved in vitro
type: sample / ID: 1000
Details: Samples are crude fraction of sucrose gradient composed by a mix of ribosomes in different states of aggregation.
Oligomeric state: Dimer / Number unique components: 2
Molecular weightTheoretical: 5.4 MDa

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Supramolecule #1: 70S ribosome

SupramoleculeName: 70S ribosome / type: complex / ID: 1 / Recombinant expression: No / Database: NCBI / Ribosome-details: ribosome-prokaryote: ALL
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 2.7 MDa

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Experimental details

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Structure determination

Methodnegative staining, cryo EM
Processingsubtomogram averaging
Aggregation stateparticle

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Sample preparation

BufferpH: 7.6
Details: 20 mM Tris-HCl, 15.2 mM (CH3COO)2Mg, 0.8 mM EDTA, 100 mM CH3COONH4, 3 mM DTT
StainingType: NEGATIVE / Details: No staining
GridDetails: R2/2 Quantifoil cupper grids
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER
Details: Vitrification instrument: Plunger. Vitrification carried out in air
Method: Blot for 2 seconds before plunging

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Electron microscopy

MicroscopeFEI/PHILIPS CM200FEG
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 53960 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 3.0 µm / Nominal magnification: 27500
Sample stageSpecimen holder: Side entry liquid nitrogen / Specimen holder model: GATAN LIQUID NITROGEN / Tilt series - Axis1 - Min angle: -60 ° / Tilt series - Axis1 - Max angle: 60 °
Alignment procedureLegacy - Astigmatism: Objective astigmatism was corrected using a quadrupole stigmator at 50,000 times magnification
Legacy - Electron beam tilt params: -4
DateJun 1, 2007
Image recordingNumber real images: 205 / Average electron dose: 50 e/Å2

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Image processing

CTF correctionDetails: Each projection
Final reconstructionAlgorithm: OTHER / Software - Name: TOM ToolBox
DetailsAverage number of projections used in the 3D reconstructions: 35.

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Atomic model buiding 1

Initial modelPDB ID:

2aw7
PDB Unreleased entry

SoftwareName: Chimera
DetailsProtocol: Rigid Body
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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Atomic model buiding 2

Initial modelPDB ID:

2awb
PDB Unreleased entry

SoftwareName: Chimera
DetailsProtocol: Rigid Body
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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