[English] 日本語
Yorodumi
- EMDB-10217: Cryo-EM structure of the RecBCD no Chi negative control complex -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-10217
TitleCryo-EM structure of the RecBCD no Chi negative control complex
Map data
SampleRecBCD bound to a forked DNA substrate that does not contain Chi:
RecBCD / DNA / (RecBCD enzyme subunit ...) x 3 / nucleic-acidNucleic acid
Function / homology
Function and homology information


exodeoxyribonuclease V activity / exodeoxyribonuclease V / exodeoxyribonuclease V complex / clearance of foreign intracellular DNA / DNA translocase activity / single-stranded DNA helicase activity / recombinational repair / 3'-5' DNA helicase activity / ATPase, acting on DNA / endodeoxyribonuclease activity ...exodeoxyribonuclease V activity / exodeoxyribonuclease V / exodeoxyribonuclease V complex / clearance of foreign intracellular DNA / DNA translocase activity / single-stranded DNA helicase activity / recombinational repair / 3'-5' DNA helicase activity / ATPase, acting on DNA / endodeoxyribonuclease activity / double-strand break repair via homologous recombination / helicase activity / response to radiation / DNA helicase activity / DNA recombination / DNA repair / cellular response to DNA damage stimulus / magnesium ion binding / DNA binding / ATP binding / cytosol
AAA+ ATPase domain / DNA helicase, UvrD/REP type / RecBCD enzyme subunit RecB / RecBCD enzyme subunit RecD / RecBCD enzyme subunit RecC / Restriction endonuclease type II-like / Exonuclease, phage-type/RecB, C-terminal / DExx box DNA helicase domain superfamily / UvrD-like helicase, ATP-binding domain / UvrD-like DNA helicase, C-terminal ...AAA+ ATPase domain / DNA helicase, UvrD/REP type / RecBCD enzyme subunit RecB / RecBCD enzyme subunit RecD / RecBCD enzyme subunit RecC / Restriction endonuclease type II-like / Exonuclease, phage-type/RecB, C-terminal / DExx box DNA helicase domain superfamily / UvrD-like helicase, ATP-binding domain / UvrD-like DNA helicase, C-terminal / P-loop containing nucleoside triphosphate hydrolase / UvrD-like helicase C-terminal domain / UvrD/AddA helicase, N-terminal / PD-(D/E)XK endonuclease-like domain, AddAB-type / RecC, C-terminal / RecBCD enzyme subunit RecD, N-terminal domain
RecBCD enzyme subunit RecD / RecBCD enzyme subunit RecC / RecBCD enzyme subunit RecB
Biological speciesEscherichia coli (E. coli) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.8 Å
AuthorsCheng K / Wilkinson M / Wigley DB
CitationJournal: Nat Struct Mol Biol / Year: 2020
Title: A conformational switch in response to Chi converts RecBCD from phage destruction to DNA repair.
Authors: Kaiying Cheng / Martin Wilkinson / Yuriy Chaban / Dale B Wigley /
Abstract: The RecBCD complex plays key roles in phage DNA degradation, CRISPR array acquisition (adaptation) and host DNA repair. The switch between these roles is regulated by a DNA sequence called Chi. We ...The RecBCD complex plays key roles in phage DNA degradation, CRISPR array acquisition (adaptation) and host DNA repair. The switch between these roles is regulated by a DNA sequence called Chi. We report cryo-EM structures of the Escherichia coli RecBCD complex bound to several different DNA forks containing a Chi sequence, including one in which Chi is recognized and others in which it is not. The Chi-recognized structure shows conformational changes in regions of the protein that contact Chi and reveals a tortuous path taken by the DNA. Sequence specificity arises from interactions with both the RecC subunit and the sequence itself. These structures provide molecular details for how Chi is recognized and insights into the changes that occur in response to Chi binding that switch RecBCD from bacteriophage destruction and CRISPR spacer acquisition to constructive host DNA repair.
Validation ReportSummary, Full report, XML, About validation report
History
DepositionAug 13, 2019-
Header (metadata) releaseJan 1, 2020-
Map releaseJan 1, 2020-
UpdateJan 29, 2020-
Current statusJan 29, 2020Processing site: PDBe / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.01
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.01
  • Imaged by UCSF Chimera
  • Download
  • Surface view with fitted model
  • Atomic models: PDB-6sjg
  • Surface level: 0.01
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

-
Map

FileDownload / File: emd_10217.map.gz / Format: CCP4 / Size: 52.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.05 Å/pix.
x 240 pix.
= 251.52 Å
1.05 Å/pix.
x 240 pix.
= 251.52 Å
1.05 Å/pix.
x 240 pix.
= 251.52 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.048 Å
Density
Contour LevelBy AUTHOR: 0.01 / Movie #1: 0.01
Minimum - Maximum-0.023008011 - 0.05779179
Average (Standard dev.)0.00001524390 (±0.002595359)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 251.51999 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.0481.0481.048
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z251.520251.520251.520
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.0230.0580.000

-
Supplemental data

-
Sample components

+
Entire RecBCD bound to a forked DNA substrate that does not contain Chi

EntireName: RecBCD bound to a forked DNA substrate that does not contain Chi
Number of components: 7

+
Component #1: protein, RecBCD bound to a forked DNA substrate that does not con...

ProteinName: RecBCD bound to a forked DNA substrate that does not contain Chi
Recombinant expression: No
MassTheoretical: 355 kDa

+
Component #2: protein, RecBCD

ProteinName: RecBCD / Recombinant expression: No
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

+
Component #3: protein, DNA

ProteinName: DNA / Recombinant expression: No
SourceSpecies: synthetic construct (others)
Source (engineered)Expression System: synthetic construct (others)

+
Component #4: protein, RecBCD enzyme subunit RecB

ProteinName: RecBCD enzyme subunit RecB / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 134.123688 kDa
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

+
Component #5: protein, RecBCD enzyme subunit RecC

ProteinName: RecBCD enzyme subunit RecC / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 128.974102 kDa
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

+
Component #6: protein, RecBCD enzyme subunit RecD

ProteinName: RecBCD enzyme subunit RecD / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 66.990367 kDa
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

+
Component #7: nucleic-acid, Forked DNA substrate

nucleic acidName: Forked DNA substrate / Class: DNA
Details: We generated a forked DNA substrate with single-stranded overhangs by annealing two synthesised oligonucleotides
Structure: OTHER / Synthetic: No
Sequence: (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DG)(DA)(DG)(DC)(DG) (DA)(DC)(DT)(DG)(DC)(DA)(DC)(DT)(DA)(DC) (DA)(DA)(DC)(DA)(DG)(DA)(DA)(DC)(DC)(DA) (DT)(DG)(DG)(DT)(DT)(DC) ...Sequence:
(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DG)(DA)(DG)(DC)(DG) (DA)(DC)(DT)(DG)(DC)(DA)(DC)(DT)(DA)(DC) (DA)(DA)(DC)(DA)(DG)(DA)(DA)(DC)(DC)(DA) (DT)(DG)(DG)(DT)(DT)(DC)(DT)(DG)(DT)(DT) (DG)(DT)(DA)(DG)(DT)(DG)(DC)(DA)(DG)(DT) (DC)(DG)(DC)(DT)(DC)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT)(DT) (DT)(DT)(DT)(DT)(DT)
MassTheoretical: 26.049602 kDa
SourceSpecies: synthetic construct (others)

-
Experimental details

-
Sample preparation

SpecimenSpecimen state: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.2 mg/mL / pH: 8
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277 K / Humidity: 90 % / Details: 1s blot before plunging.

-
Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 45 e/Å2 / Illumination mode: FLOOD BEAM
LensMagnification: 130000.0 X (nominal), 47710.0 X (calibrated) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 1400.0 - 2600.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

-
Image acquisition

Image acquisitionNumber of digital images: 788 / Sampling size: 5 µm

-
Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 62812
3D reconstructionSoftware: RELION / Resolution: 3.8 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot (resolution estimation)

-
Atomic model buiding

Modeling #1Refinement space: REAL
Details: Rounds of manual structure editing in coot and real-space refinement with Phenix
Input PDB model: 6SJF
Output model

+
About Yorodumi

-
News

-
Aug 12, 2020. New: Covid-19 info

New: Covid-19 info

  • 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. New: 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. (see PDBe EMDB page)
  • 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 at PDBe / 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. See below links for details.
  • 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

+
Jun 16, 2017. Omokage search with filter

Omokage search with filter

  • Result of Omokage search can be filtered by keywords and the database types

Related info.:Omokage search

Read more

-
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.

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