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- EMDB-30211: State 4 of Pre50SH ribosome from RrmJ knock out strain -

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Basic information

Entry
Database: EMDB / ID: EMD-30211
TitleState 4 of Pre50SH ribosome from RrmJ knock out strain
Map dataState 4
Sample
  • Complex: State 4 of pre50SH ribosome
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 11.1 Å
AuthorsWang W / Li WQ / Ge XL / Yan KG / Mandava CS / Sanyal S / Gao N
Funding support China, 5 items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)31630087 China
Ministry of Science and Technology (MoST, China)2019YFA0508900 China
National Natural Science Foundation of China (NSFC)31725007 China
Other governmentShenzhen Science and Technology Innovation Committee (JCYJ20180302174213122) China
Other governmentChina Postdoctoral Science Foundation (1131000065) China
CitationJournal: To Be Published
Title: Loss of a single methylation in 23S rRNA delays 50S assembly at multiple late stages and impairs translation initiation and elongation
Authors: Wang W / Li WQ / Ge XL / Yan KG / Mandava CS / Sanyal S / Gao N
History
DepositionApr 9, 2020-
Header (metadata) releaseJul 1, 2020-
Map releaseJul 1, 2020-
UpdateJul 1, 2020-
Current statusJul 1, 2020Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.034
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.034
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_30211.png

Downloads & links

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Map

FileDownload / File: emd_30211.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationState 4
Voxel sizeX=Y=Z: 1.08 Å
Density
Contour LevelBy AUTHOR: 0.034 / Movie #1: 0.034
Minimum - Maximum-0.043240014 - 0.11720092
Average (Standard dev.)0.0028343329 (±0.012187416)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 345.6 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.081.081.08
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z345.600345.600345.600
α/β/γ90.00090.00090.000
start NX/NY/NZ-200-200-200
NX/NY/NZ400400400
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-0.0430.1170.003

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Supplemental data

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Sample components

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Entire : State 4 of pre50SH ribosome

EntireName: State 4 of pre50SH ribosome
Components
  • Complex: State 4 of pre50SH ribosome

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Supramolecule #1: State 4 of pre50SH ribosome

SupramoleculeName: State 4 of pre50SH ribosome / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#32
Details: State 4 of pre50SH ribosome from RrmJ knock out E.coli strain
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 1.5 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.15 mg/mL
BufferpH: 7.5
Details: Solutions were made fresh form concentrated to avoid microbial contamination.
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 7000.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.03 kPa
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: blot for 2 seconds before plunging.
DetailsThis sample was monodisperse.

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated magnification: 75000 / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 75000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
DetailsPreliminary grid screening was performed manually.
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Digitization - Frames/image: 3-28 / Number grids imaged: 1 / Number real images: 1262 / Average electron dose: 45.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 235600
Details: Images were binned and lowpass filtered for autopicking.
CTF correctionSoftware - Name: CTFFIND (ver. 3.0)
Details: CTF amplitude correction was performed following 3D reconstruction.
Startup modelType of model: OTHER
Details: The model was a low resolution 50S ribosome generated from our own data.
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 1.4) / Details: Relion software was used in the reconstruction.
Final 3D classificationNumber classes: 6 / Avg.num./class: 15300 / Software - Name: RELION (ver. 1.4)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 1.4) / Details: Relion software was used in the reconstruction.
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: SIMULTANEOUS ITERATIVE (SIRT) / Resolution.type: BY AUTHOR / Resolution: 11.1 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 1.4) / Number images used: 1372
DetailsThe images were normalized and manually selected.

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Atomic model buiding 1

Initial model(PDB ID:

4ybb

,

5h5u

)

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