[English] 日本語
Yorodumi
- EMDB-2176: Negative stain electron microscopy of a CSN complex -

+
Open data


ID or keywords:

Loading...

-
Basic information

Entry
Database: EMDB / ID: EMD-2176
TitleNegative stain electron microscopy of a CSN complex
Map dataCSN
Sample
  • Sample: CSN
  • Protein or peptide: Csn1
  • Protein or peptide: Csn2
  • Protein or peptide: Csn3
  • Protein or peptide: Csn4
  • Protein or peptide: Csn5
  • Protein or peptide: Csn6
  • Protein or peptide: Csn7b
  • Protein or peptide: Csn8
KeywordsCop9 Signalosome
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / negative staining / Resolution: 25.0 Å
AuthorsEnchev RI / Scott DC / da Fonseca P / Schreiber A / Monda JK / Schulman BA / Peter M / Morris EP
CitationJournal: Cell Rep / Year: 2012
Title: Structural basis for a reciprocal regulation between SCF and CSN.
Authors: Radoslav I Enchev / Daniel C Scott / Paula C A da Fonseca / Anne Schreiber / Julie K Monda / Brenda A Schulman / Matthias Peter / Edward P Morris /
Abstract: Skp1-Cul1-Fbox (SCF) E3 ligases are activated by ligation to the ubiquitin-like protein Nedd8, which is reversed by the deneddylating Cop9 signalosome (CSN). However, CSN also promotes SCF substrate ...Skp1-Cul1-Fbox (SCF) E3 ligases are activated by ligation to the ubiquitin-like protein Nedd8, which is reversed by the deneddylating Cop9 signalosome (CSN). However, CSN also promotes SCF substrate turnover through unknown mechanisms. Through biochemical and electron microscopy analyses, we determined molecular models of CSN complexes with SCF(Skp2/Cks1) and SCF(Fbw7) and found that CSN occludes both SCF functional sites-the catalytic Rbx1-Cul1 C-terminal domain and the substrate receptor. Indeed, CSN binding prevents SCF interactions with E2 enzymes and a ubiquitination substrate, and it inhibits SCF-catalyzed ubiquitin chain formation independent of deneddylation. Importantly, CSN prevents neddylation of the bound cullin, unless binding of a ubiquitination substrate triggers SCF dissociation and neddylation. Taken together, the results provide a model for how reciprocal regulation sensitizes CSN to the SCF assembly state and inhibits a catalytically competent SCF until a ubiquitination substrate drives its own degradation by displacing CSN, thereby promoting cullin neddylation and substrate ubiquitination.
History
DepositionAug 11, 2012-
Header (metadata) releaseOct 3, 2012-
Map releaseOct 3, 2012-
UpdateOct 3, 2012-
Current statusOct 3, 2012Processing site: PDBe / Status: Released

-
Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 3
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 3
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

EMD-2176.jpg

Downloads & links

-
Map

FileDownload / File: emd_2176.map.gz / Format: CCP4 / Size: 3.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCSN
Voxel sizeX=Y=Z: 3.47 Å
Density
Contour LevelBy AUTHOR: 3.0 / Movie #1: 3
Minimum - Maximum-11.70456409 - 20.460073470000001
Average (Standard dev.)0.05268458 (±0.81265736)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-50-50-50
Dimensions100100100
Spacing100100100
CellA=B=C: 347.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.473.473.47
M x/y/z100100100
origin x/y/z0.0000.0000.000
length x/y/z347.000347.000347.000
α/β/γ90.00090.00090.000
start NX/NY/NZ-184-184-183
NX/NY/NZ368368368
MAP C/R/S123
start NC/NR/NS-50-50-50
NC/NR/NS100100100
D min/max/mean-11.70520.4600.053

-
Supplemental data

-
Sample components

-
Entire : CSN

EntireName: CSN
Components
  • Sample: CSN
  • Protein or peptide: Csn1
  • Protein or peptide: Csn2
  • Protein or peptide: Csn3
  • Protein or peptide: Csn4
  • Protein or peptide: Csn5
  • Protein or peptide: Csn6
  • Protein or peptide: Csn7b
  • Protein or peptide: Csn8

-
Supramolecule #1000: CSN

SupramoleculeName: CSN / type: sample / ID: 1000
Details: The sample was purified over affinity pull-down and Superose6 size exclusion chromatography
Oligomeric state: one CSN monomer / Number unique components: 8
Molecular weightTheoretical: 320 KDa

-
Macromolecule #1: Csn1

MacromoleculeName: Csn1 / type: protein_or_peptide / ID: 1 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 53 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant plasmid: recombinant baculovirus/ MultiBac

-
Macromolecule #2: Csn2

MacromoleculeName: Csn2 / type: protein_or_peptide / ID: 2 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 52 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant plasmid: recombinant baculovirus/ MultiBac

-
Macromolecule #3: Csn3

MacromoleculeName: Csn3 / type: protein_or_peptide / ID: 3 / Details: N-terminal StrepII2x tag / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 48 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant plasmid: recombinant baculovirus/ MultiBac

-
Macromolecule #4: Csn4

MacromoleculeName: Csn4 / type: protein_or_peptide / ID: 4 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 46 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant plasmid: recombinant baculovirus/ MultiBac

-
Macromolecule #5: Csn5

MacromoleculeName: Csn5 / type: protein_or_peptide / ID: 5 / Details: N-terminal His6 tag / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 40 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant plasmid: recombinant baculovirus/ MultiBac

-
Macromolecule #6: Csn6

MacromoleculeName: Csn6 / type: protein_or_peptide / ID: 6 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 36 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant plasmid: recombinant baculovirus/ MultiBac

-
Macromolecule #7: Csn7b

MacromoleculeName: Csn7b / type: protein_or_peptide / ID: 7 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 30 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant plasmid: recombinant baculovirus/ MultiBac

-
Macromolecule #8: Csn8

MacromoleculeName: Csn8 / type: protein_or_peptide / ID: 8 / Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes
Source (natural)Organism: Homo sapiens (human) / synonym: Human
Molecular weightTheoretical: 23 KDa
Recombinant expressionOrganism: Trichoplusia ni (cabbage looper) / Recombinant plasmid: recombinant baculovirus/ MultiBac

-
Experimental details

-
Structure determination

Methodnegative staining
Processingsingle particle reconstruction
Aggregation stateparticle

-
Sample preparation

Concentration0.2 mg/mL
BufferpH: 7.8
Details: 15 mM HEPES pH 7.8, 150 mM NaCl, 1% glycerol and 1 mM DTT
StainingType: NEGATIVE
Details: Quantifoil grids (R2/2 Cu 400 mesh) coated with thin carbon floated from mica were glow-discharged for 30 seconds at 50 mA and 0.2 mbar vacuum. 3 ul purified samples were applied for 1 min ...Details: Quantifoil grids (R2/2 Cu 400 mesh) coated with thin carbon floated from mica were glow-discharged for 30 seconds at 50 mA and 0.2 mbar vacuum. 3 ul purified samples were applied for 1 min to the grids. Following two brief buffer washes, the grids were stained with 2% uranyl acetate, gently blotted using filter paper and air-dried.
VitrificationCryogen name: NONE / Instrument: OTHER

-
Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 50000
Sample stageSpecimen holder model: SIDE ENTRY, EUCENTRIC
DateJan 10, 2011
Image recordingCategory: CCD / Film or detector model: GENERIC TVIPS (4k x 4k) / Average electron dose: 100 e/Å2 / Details: data was collected on a CCD camera
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

-
Image processing

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 25.0 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: IMAGIC, Spider, EMAN, in-house / Number images used: 5165
Detailssee publication

+
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