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- EMDB-43845: Structure of Circularly Permuted 50S Ribosomal Subunit Assembly I... -

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Basic information

Entry
Database: EMDB / ID: EMD-43845
TitleStructure of Circularly Permuted 50S Ribosomal Subunit Assembly Intermediate - CP45 Class C1
Map dataClass C1 CP45 50S Ribosomal Intermediate
Sample
  • Complex: CP45 iSAT 50S ribosomal subunit assembly intermediate
Keywords50S subunit / assembly intermediate / RNA-protein complex / ribosome
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.4 Å
AuthorsDong X / Sheng K / Williamson JR
Funding support United States, 3 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM136412 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM053757 United States
National Institutes of Health/National Institute Of Allergy and Infectious Diseases (NIH/NIAID)AI170855 United States
CitationJournal: To Be Published
Title: Assembly of Bacterial Ribosome with Circularly Permuted rRNA
Authors: Dong X / Sheng K / Williamson JR / Gebert L / Aiyer S / Lyumkis D
History
DepositionFeb 28, 2024-
Header (metadata) releaseApr 24, 2024-
Map releaseApr 24, 2024-
UpdateApr 24, 2024-
Current statusApr 24, 2024Processing site: RCSB / Status: Released

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Structure visualization

Supplemental images

emd_43845.png

Downloads & links

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Map

FileDownload / File: emd_43845.map.gz / Format: CCP4 / Size: 187 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationClass C1 CP45 50S Ribosomal Intermediate
Voxel sizeX=Y=Z: 1.15 Å
Density
Contour LevelBy AUTHOR: 0.25
Minimum - Maximum-0.16339302 - 0.9089645
Average (Standard dev.)0.0019127954 (±0.04751022)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions366366366
Spacing366366366
CellA=B=C: 420.9 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Class C1 CP45 50S Ribosomal Intermediate - Half Map 2

Fileemd_43845_half_map_1.map
AnnotationClass C1 CP45 50S Ribosomal Intermediate - Half Map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Class C1 CP45 50S Ribosomal Intermediate - Half Map 1

Fileemd_43845_half_map_2.map
AnnotationClass C1 CP45 50S Ribosomal Intermediate - Half Map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : CP45 iSAT 50S ribosomal subunit assembly intermediate

EntireName: CP45 iSAT 50S ribosomal subunit assembly intermediate
Components
  • Complex: CP45 iSAT 50S ribosomal subunit assembly intermediate

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Supramolecule #1: CP45 iSAT 50S ribosomal subunit assembly intermediate

SupramoleculeName: CP45 iSAT 50S ribosomal subunit assembly intermediate / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Escherichia coli (E. coli) / Strain: MRE600

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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

BufferpH: 7.5
Component:
ConcentrationFormulaName
50.0 mMC4H11NO3Tris Hydrochloride
100.0 mMNH4ClAmmonium Chloride
10.0 mMMgCl2Magnesium Chloride
0.5 mMC10H16N2O8Ethylenediaminetetraacetic Acid
6.0 mMC2H6OSBeta-mercaptoethanol2-Mercaptoethanol

Details: Most of the sucrose was removed by spin concentration.
GridModel: Quantifoil R1.2/1.3 / Material: COPPER / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 2
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 298 K / Instrument: FEI VITROBOT MARK IV / Details: 3 microliter of the sample was added..
DetailsThe in vitro assembled large ribosomal subunit was purified by sucrose gradient and was spin-concentrated in a 100 kDa MW-cutoff filter.

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Electron microscopy

MicroscopeFEI TALOS ARCTICA
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 36000
Sample stageCooling holder cryogen: NITROGEN
DetailsIn order to account for highly preferred orientation of the specimen, data were acquired using tilts ranging at -20 degrees.
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Number grids imaged: 1 / Number real images: 1599 / Average exposure time: 5.0 sec. / Average electron dose: 50.0 e/Å2
Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 487886 / Details: Particles were selected by cryoSPARC.
Startup modelType of model: NONE / Details: Ab initio reconstruction in CryoSPARC was used.
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final 3D classificationNumber classes: 10 / Avg.num./class: 6000 / Details: 3D classification of cryoSPARC was used.
Final angle assignmentType: MAXIMUM LIKELIHOOD / Details: Non-uniform Refinement of cryoSPARC was used.
Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC / Number images used: 10990
FSC plot (resolution estimation)

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