[English] 日本語
Yorodumi
- EMDB-15928: Cryo-electron tomogram of VeroE6 cells transfected with HA-GG>AA-... -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-15928
TitleCryo-electron tomogram of VeroE6 cells transfected with HA-GG>AA-nsp4-V5, plunge-frozen at 16 hpt and subjected to cryo-FIB milling.
Map data
Sample
  • Cell: VeroE6 cells transfected with SARS-CoV-2 nsp3-4 truncation: pCDNA3.1-HA-nsp3-deltaUbl1-Mac1-nsp4-V5
Keywordsnsp3-4 protein / VIRAL PROTEIN
Biological speciesSevere acute respiratory syndrome coronavirus 2
Methodelectron tomography / cryo EM
AuthorsChlanda P / Zimmermann L
Funding support Germany, 1 items
OrganizationGrant numberCountry
Other privateChica and Heinz Schaller Foundation Germany
CitationJournal: Nat Commun / Year: 2023
Title: SARS-CoV-2 nsp3 and nsp4 are minimal constituents of a pore spanning replication organelle.
Authors: Liv Zimmermann / Xiaohan Zhao / Jana Makroczyova / Moritz Wachsmuth-Melm / Vibhu Prasad / Zach Hensel / Ralf Bartenschlager / Petr Chlanda /
Abstract: Coronavirus replication is associated with the remodeling of cellular membranes, resulting in the formation of double-membrane vesicles (DMVs). A DMV-spanning pore was identified as a putative portal ...Coronavirus replication is associated with the remodeling of cellular membranes, resulting in the formation of double-membrane vesicles (DMVs). A DMV-spanning pore was identified as a putative portal for viral RNA. However, the exact components and the structure of the SARS-CoV-2 DMV pore remain to be determined. Here, we investigate the structure of the DMV pore by in situ cryo-electron tomography combined with subtomogram averaging. We identify non-structural protein (nsp) 3 and 4 as minimal components required for the formation of a DMV-spanning pore, which is dependent on nsp3-4 proteolytic cleavage. In addition, we show that Mac2-Mac3-DPUP-Ubl2 domains are critical for nsp3 oligomerization and crown integrity which influences membrane curvature required for biogenesis of DMVs. Altogether, SARS-CoV-2 nsp3-4 have a dual role by driving the biogenesis of replication organelles and assembly of DMV-spanning pores which we propose here to term replicopores.
History
DepositionOct 5, 2022-
Header (metadata) releaseOct 18, 2023-
Map releaseOct 18, 2023-
UpdateDec 27, 2023-
Current statusDec 27, 2023Processing site: PDBe / Status: Released

-
Structure visualization

Supplemental images

emd_15928.png

Downloads & links

-
Map

FileDownload / File: emd_15928.map.gz / Format: CCP4 / Size: 821.7 MB / Type: IMAGE STORED AS SIGNED BYTE
Voxel sizeX=Y=Z: 6.468 Å
Density
Minimum - Maximum-128.0 - 127.0
Average (Standard dev.)-0.7005053 (±25.204224)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-344344-204
Dimensions19201364329
Spacing13641920329
CellA: 8822.352 Å / B: 12418.56 Å / C: 2127.972 Å
α=β=γ: 90.0 °

-
Supplemental data

-
Sample components

-
Entire : VeroE6 cells transfected with SARS-CoV-2 nsp3-4 truncation: pCDNA...

EntireName: VeroE6 cells transfected with SARS-CoV-2 nsp3-4 truncation: pCDNA3.1-HA-nsp3-deltaUbl1-Mac1-nsp4-V5
Components
  • Cell: VeroE6 cells transfected with SARS-CoV-2 nsp3-4 truncation: pCDNA3.1-HA-nsp3-deltaUbl1-Mac1-nsp4-V5

-
Supramolecule #1: VeroE6 cells transfected with SARS-CoV-2 nsp3-4 truncation: pCDNA...

SupramoleculeName: VeroE6 cells transfected with SARS-CoV-2 nsp3-4 truncation: pCDNA3.1-HA-nsp3-deltaUbl1-Mac1-nsp4-V5
type: cell / ID: 1 / Parent: 0
Source (natural)Organism: Severe acute respiratory syndrome coronavirus 2

-
Experimental details

-
Structure determination

Methodcryo EM
Processingelectron tomography
Aggregation statecell

-
Sample preparation

BufferpH: 7.4
VitrificationCryogen name: ETHANE
Cryo protectantno
SectioningFocused ion beam - Instrument: OTHER / Focused ion beam - Ion: OTHER / Focused ion beam - Voltage: 30 / Focused ion beam - Current: 0.03 / Focused ion beam - Duration: 200 / Focused ion beam - Temperature: 90 K / Focused ion beam - Initial thickness: 1000 / Focused ion beam - Final thickness: 200
Focused ion beam - Details: The value given for _em_focused_ion_beam.instrument is Aquilos cryo-FIB-SEM (Thermo Fisher Scientific). This is not in a list of allowed values {'DB235', 'OTHER'} so OTHER ...Focused ion beam - Details: The value given for _em_focused_ion_beam.instrument is Aquilos cryo-FIB-SEM (Thermo Fisher Scientific). This is not in a list of allowed values {'DB235', 'OTHER'} so OTHER is written into the XML file.

-
Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 (6k x 4k) / Average electron dose: 3.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: LAB6
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 4.0 µm / Nominal defocus min: 2.5 µm / Nominal magnification: 42000
Sample stageCooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

-
Image processing

Final reconstructionSoftware - Name: SerialEM / Number images used: 41

+
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://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