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- EMDB-20158: Ribosome 70S at 2.8 Angstrom -

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Basic information

Entry
Database: EMDB / ID: EMD-20158
TitleRibosome 70S at 2.8 Angstrom
Map dataSharpen map
Sample
  • Complex: 70S
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsMehrani A / Mendez JH / Stagg SM
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Human Genome Research InstituteR01GM108753 United States
CitationJournal: IUCrJ / Year: 2019
Title: Throughput and resolution with a next-generation direct electron detector.
Authors: Joshua H Mendez / Atousa Mehrani / Peter Randolph / Scott Stagg /
Abstract: Direct electron detectors (DEDs) have revolutionized cryo-electron microscopy (cryo-EM) by facilitating the correction of beam-induced motion and radiation damage, and also by providing high- ...Direct electron detectors (DEDs) have revolutionized cryo-electron microscopy (cryo-EM) by facilitating the correction of beam-induced motion and radiation damage, and also by providing high-resolution image capture. A new-generation DED, the DE64, has been developed by Direct Electron that has good performance in both integrating and counting modes. The camera has been characterized in both modes in terms of image quality, throughput and resolution of cryo-EM reconstructions. The modulation transfer function, noise power spectrum and detective quantum efficiency (DQE) were determined for both modes, as well as the number of images per unit time. Although the DQE for counting mode was superior to that for integrating mode, the data-collection throughput for this mode was more than ten times slower. Since throughput and resolution are related in single-particle cryo-EM, data for apoferritin were collected and reconstructed using integrating mode, integrating mode in conjunction with a Volta phase plate (VPP) and counting mode. Only the counting-mode data resulted in a better than 3 Å resolution reconstruction with similar numbers of particles, and this increased performance could not be compensated for by the increased throughput of integrating mode or by the increased low-frequency contrast of integrating mode with the VPP. These data show that the superior image quality provided by counting mode is more important for high-resolution cryo-EM reconstructions than the superior throughput of integrating mode.
History
DepositionApr 25, 2019-
Header (metadata) releaseMay 8, 2019-
Map releaseNov 27, 2019-
UpdateNov 27, 2019-
Current statusNov 27, 2019Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.03
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by height
  • Surface level: 0.03
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_20158.png

Downloads & links

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Map

FileDownload / File: emd_20158.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpen map
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.19 Å/pix.
x 384 pix.
= 457.344 Å		1.19 Å/pix.
x 384 pix.
= 457.344 Å		1.19 Å/pix.
x 384 pix.
= 457.344 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.191 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.03 / Movie #1: 0.03
Minimum - Maximum-0.16843809 - 0.28148133
Average (Standard dev.)0.00036377512 (±0.0066105113)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 457.344 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.1911.1911.191
M x/y/z384384384
origin x/y/z0.0000.0000.000
length x/y/z457.344457.344457.344
α/β/γ90.00090.00090.000
start NX/NY/NZ-420-29
NX/NY/NZ887886
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS384384384
D min/max/mean-0.1680.2810.000

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Supplemental data

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Half map: Even half map

Fileemd_20158_half_map_1.map
AnnotationEven half map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Odd half map

Fileemd_20158_half_map_2.map
AnnotationOdd half map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : 70S

EntireName: 70S
Components
  • Complex: 70S

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Supramolecule #1: 70S

SupramoleculeName: 70S / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 2.1 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

BufferpH: 7.4
GridDetails: unspecified
VitrificationCryogen name: NITROGEN

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Image recordingFilm or detector model: DIRECT ELECTRON DE-64 (8k x 8k) / Detector mode: COUNTING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Digitization - Sampling interval: 13.0 µm / Digitization - Frames/image: 1-78 / Number grids imaged: 1 / Number real images: 1752 / Average exposure time: 19.3 sec. / Average electron dose: 25.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

DetailsCollected in counting mode, with hardware binning of 2 resulting in a sensor size is 4k x 4k.
CTF correctionSoftware: (Name: Gctf (ver. 1.06), CTFFIND (ver. 4))
Final reconstructionNumber classes used: 1 / Resolution.type: BY AUTHOR / Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.0) / Number images used: 348462
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
FSC plot (resolution estimation)

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