[English] 日本語
Yorodumi
- EMDB-18826: In situ sub-tomogram average of the E. coli 70S ribosome obtained... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-18826
TitleIn situ sub-tomogram average of the E. coli 70S ribosome obtained using honeycomb gold supports
Map dataA sub-tomogram average of the 70S ribosome with E-site tRNA bound, obtained from FIB-milled vertically trapped Escherichia coli cells.
Sample
  • Complex: 70S ribosome with E-site tRNA
Keywordsribosome / tomography / sub-tomogram averaging
Biological speciesEscherichia coli (E. coli)
Methodsubtomogram averaging / cryo EM / Resolution: 6.7 Å
AuthorsHale VL / Hooker JA / Russo CJ / Lowe J
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Medical Research Council (MRC, United Kingdom) United Kingdom
CitationJournal: J Struct Biol / Year: 2024
Title: Honeycomb gold specimen supports enabling orthogonal focussed ion beam-milling of elongated cells for cryo-ET.
Authors: Victoria L Hale / James Hooker / Christopher J Russo / Jan Löwe /
Abstract: Cryo-focussed ion beam (FIB)-milling is a powerful technique that opens up thick, cellular specimens to high-resolution structural analysis by electron cryotomography (cryo-ET). FIB-milled lamellae ...Cryo-focussed ion beam (FIB)-milling is a powerful technique that opens up thick, cellular specimens to high-resolution structural analysis by electron cryotomography (cryo-ET). FIB-milled lamellae can be produced from cells on grids, or cut from thicker, high-pressure frozen specimens. However, these approaches can put geometrical constraints on the specimen that may be unhelpful, particularly when imaging structures within the cell that have a very defined orientation. For example, plunge frozen rod-shaped bacteria orient parallel to the plane of the grid, yet the Z-ring, a filamentous structure of the tubulin-like protein FtsZ and the key organiser of bacterial division, runs around the circumference of the cell such that it is perpendicular to the imaging plane. It is therefore difficult or impractical to image many complete rings with current technologies. To circumvent this problem, we have fabricated monolithic gold specimen supports with a regular array of cylindrical wells in a honeycomb geometry, which trap bacteria in a vertical orientation. These supports, which we call "honeycomb gold discs", replace standard EM grids and when combined with FIB-milling enable the production of lamellae containing cross-sections through cells. The resulting lamellae are more stable and resistant to breakage and charging than conventional lamellae. The design of the honeycomb discs can be modified according to need and so will also enable cryo-ET and cryo-EM imaging of other specimens in otherwise difficult to obtain orientations.
History
DepositionNov 2, 2023-
Header (metadata) releaseJun 5, 2024-
Map releaseJun 5, 2024-
UpdateJun 5, 2024-
Current statusJun 5, 2024Processing site: PDBe / Status: Released

-
Structure visualization

Supplemental images

emd_18826.png

Downloads & links

-
Map

FileDownload / File: emd_18826.map.gz / Format: CCP4 / Size: 28.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationA sub-tomogram average of the 70S ribosome with E-site tRNA bound, obtained from FIB-milled vertically trapped Escherichia coli cells.
Voxel sizeX=Y=Z: 2.659 Å
Density
Contour LevelBy AUTHOR: 0.00178
Minimum - Maximum-0.004542594 - 0.0090929065
Average (Standard dev.)-0.000015406655 (±0.00037909532)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions196196196
Spacing196196196
CellA=B=C: 521.164 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Half map: #2

Fileemd_18826_half_map_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Half map: #1

Fileemd_18826_half_map_2.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

-
Sample components

-
Entire : 70S ribosome with E-site tRNA

EntireName: 70S ribosome with E-site tRNA
Components
  • Complex: 70S ribosome with E-site tRNA

-
Supramolecule #1: 70S ribosome with E-site tRNA

SupramoleculeName: 70S ribosome with E-site tRNA / type: complex / ID: 1 / Parent: 0 / Details: Native ribosomes
Source (natural)Organism: Escherichia coli (E. coli) / Strain: C41(DE3) / Location in cell: cytoplasm
Molecular weightTheoretical: 2.5 MDa

-
Experimental details

-
Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation statecell

-
Sample preparation

BufferpH: 7.7
GridMaterial: GOLD / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 240 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 0.025 kPa / Details: 30 mA, 2 minutes per side
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER / Details: Manually back-blotted at room temperature.

-
Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4088 pixel / Number real images: 5 / Average exposure time: 0.3 sec. / Average electron dose: 3.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 6.0 µm / Nominal defocus min: 2.0 µm / Nominal magnification: 33000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

-
Image processing

Final reconstructionResolution.type: BY AUTHOR / Resolution: 6.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: Warp (ver. 1.0.9) / Software - details: M / Number subtomograms used: 15500
ExtractionNumber tomograms: 23 / Number images used: 83000 / Software: (Name: crYOLO (ver. 1.8.3), Warp (ver. 1.0.9))
Final angle assignmentType: PROJECTION MATCHING / Software - Name: Warp (ver. 1.0.9) / Software - details: M
FSC plot (resolution estimation)

+
About Yorodumi

-
News

-
Feb 9, 2022. New format data for meta-information of EMDB entries

New format data for meta-information of EMDB entries

  • Version 3 of the EMDB header file is now the official format.
  • The previous official version 1.9 will be removed from the archive.

Related info.:EMDB header

External links:wwPDB to switch to version 3 of the EMDB data model

-
Aug 12, 2020. Covid-19 info

Covid-19 info

URL: https://pdbjlvh1.pdbj.org/emnavi/covid19.php

New page: Covid-19 featured information page in EM Navigator.

Related info.:Covid-19 info / Mar 5, 2020. Novel coronavirus structure data

+
Mar 5, 2020. Novel coronavirus structure data

Novel coronavirus structure data

Related info.:Yorodumi Speices / Aug 12, 2020. Covid-19 info

External links:COVID-19 featured content - PDBj / Molecule of the Month (242)：Coronavirus Proteases

+
Jan 31, 2019. EMDB accession codes are about to change! (news from PDBe EMDB page)

EMDB accession codes are about to change! (news from PDBe EMDB page)

  • The allocation of 4 digits for EMDB accession codes will soon come to an end. Whilst these codes will remain in use, new EMDB accession codes will include an additional digit and will expand incrementally as the available range of codes is exhausted. The current 4-digit format prefixed with “EMD-” (i.e. EMD-XXXX) will advance to a 5-digit format (i.e. EMD-XXXXX), and so on. It is currently estimated that the 4-digit codes will be depleted around Spring 2019, at which point the 5-digit format will come into force.
  • The EM Navigator/Yorodumi systems omit the EMD- prefix.

Related info.:Q: What is EMD? / ID/Accession-code notation in Yorodumi/EM Navigator

External links:EMDB Accession Codes are Changing Soon! / Contact to PDBj

+
Jul 12, 2017. Major update of PDB

Major update of PDB

  • wwPDB released updated PDB data conforming to the new PDBx/mmCIF dictionary.
  • This is a major update changing the version number from 4 to 5, and with Remediation, in which all the entries are updated.
  • In this update, many items about electron microscopy experimental information are reorganized (e.g. em_software).
  • Now, EM Navigator and Yorodumi are based on the updated data.

External links:wwPDB Remediation / Enriched Model Files Conforming to OneDep Data Standards Now Available in the PDB FTP Archive

-
Yorodumi

Thousand views of thousand structures

  • Yorodumi is a browser for structure data from EMDB, PDB, SASBDB, etc.
  • This page is also the successor to EM Navigator detail page, and also detail information page/front-end page for Omokage search.
  • The word "yorodu" (or yorozu) is an old Japanese word meaning "ten thousand". "mi" (miru) is to see.

Related info.:EMDB / PDB / SASBDB / Comparison of 3 databanks / Yorodumi Search / Aug 31, 2016. New EM Navigator & Yorodumi / Yorodumi Papers / Jmol/JSmol / Function and homology information / Changes in new EM Navigator and Yorodumi

Read more