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- EMDB-1046: ATP-bound states of GroEL captured by cryo-electron microscopy. -

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Basic information

Entry
Database: EMDB / ID: EMD-1046
TitleATP-bound states of GroEL captured by cryo-electron microscopy.
Map dataGroEL with GroES and ADP bound to one ring, and ATP bound to the other ring
Sample
  • Sample: GroES-ADP7-GroEL-ATP7 from E.coli
  • Protein or peptide: GroEL
  • Protein or peptide: GroES
Function / homology
Function and homology information


GroEL-GroES complex / chaperonin ATPase / virion assembly / chaperone cofactor-dependent protein refolding / protein folding chaperone / isomerase activity / ATP-dependent protein folding chaperone / response to radiation / unfolded protein binding / protein folding ...GroEL-GroES complex / chaperonin ATPase / virion assembly / chaperone cofactor-dependent protein refolding / protein folding chaperone / isomerase activity / ATP-dependent protein folding chaperone / response to radiation / unfolded protein binding / protein folding / response to heat / protein-folding chaperone binding / protein refolding / magnesium ion binding / ATP hydrolysis activity / ATP binding / membrane / metal ion binding / identical protein binding / cytosol / cytoplasm
Similarity search - Function
Chaperonin GroES, conserved site / Chaperonins cpn10 signature. / Chaperonin 10 Kd subunit / GroES chaperonin family / GroES chaperonin superfamily / Chaperonin 10 Kd subunit / Chaperonin Cpn60, conserved site / Chaperonins cpn60 signature. / Chaperonin Cpn60/GroEL / GroEL-like equatorial domain superfamily ...Chaperonin GroES, conserved site / Chaperonins cpn10 signature. / Chaperonin 10 Kd subunit / GroES chaperonin family / GroES chaperonin superfamily / Chaperonin 10 Kd subunit / Chaperonin Cpn60, conserved site / Chaperonins cpn60 signature. / Chaperonin Cpn60/GroEL / GroEL-like equatorial domain superfamily / TCP-1-like chaperonin intermediate domain superfamily / GroEL-like apical domain superfamily / TCP-1/cpn60 chaperonin family / Chaperonin Cpn60/GroEL/TCP-1 family / GroES-like superfamily
Similarity search - Domain/homology
Co-chaperonin GroES / Chaperonin GroEL / Chaperonin GroEL / Co-chaperonin GroES
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 23.5 Å
AuthorsSaibil HR
CitationJournal: Cell / Year: 2001
Title: ATP-bound states of GroEL captured by cryo-electron microscopy.
Authors: N A Ranson / G W Farr / A M Roseman / B Gowen / W A Fenton / A L Horwich / H R Saibil /
Abstract: The chaperonin GroEL drives its protein-folding cycle by cooperatively binding ATP to one of its two rings, priming that ring to become folding-active upon GroES binding, while simultaneously ...The chaperonin GroEL drives its protein-folding cycle by cooperatively binding ATP to one of its two rings, priming that ring to become folding-active upon GroES binding, while simultaneously discharging the previous folding chamber from the opposite ring. The GroEL-ATP structure, determined by cryo-EM and atomic structure fitting, shows that the intermediate domains rotate downward, switching their intersubunit salt bridge contacts from substrate binding to ATP binding domains. These observations, together with the effects of ATP binding to a GroEL-GroES-ADP complex, suggest structural models for the ATP-induced reduction in affinity for polypeptide and for cooperativity. The model for cooperativity, based on switching of intersubunit salt bridge interactions around the GroEL ring, may provide general insight into cooperativity in other ring complexes and molecular machines.
History
DepositionMar 13, 2003-
Header (metadata) releaseMay 16, 2003-
Map releaseMay 16, 2003-
UpdateOct 24, 2012-
Current statusOct 24, 2012Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.08
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.08
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-1gru
  • Surface level: 0.08
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
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Supplemental images

Downloads & links

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Map

FileDownload / File: emd_1046.map.gz / Format: CCP4 / Size: 7.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationGroEL with GroES and ADP bound to one ring, and ATP bound to the other ring
Voxel sizeX=Y=Z: 2.8 Å
Density
Contour LevelBy AUTHOR: 0.029 / Movie #1: 0.08
Minimum - Maximum-0.04720771 - 0.27609465
Average (Standard dev.)0.00606513 (±0.02973022)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions128128128
Spacing128128128
CellA=B=C: 358.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.82.82.8
M x/y/z128128128
origin x/y/z0.0000.0000.000
length x/y/z358.400358.400358.400
α/β/γ90.00090.00090.000
start NX/NY/NZ0052
NX/NY/NZ12812855
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS128128128
D min/max/mean-0.0470.2760.006

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Supplemental data

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Sample components

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Entire : GroES-ADP7-GroEL-ATP7 from E.coli

EntireName: GroES-ADP7-GroEL-ATP7 from E.coli
Components
  • Sample: GroES-ADP7-GroEL-ATP7 from E.coli
  • Protein or peptide: GroEL
  • Protein or peptide: GroES

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Supramolecule #1000: GroES-ADP7-GroEL-ATP7 from E.coli

SupramoleculeName: GroES-ADP7-GroEL-ATP7 from E.coli / type: sample / ID: 1000
Details: The complexes were prepared by pre-forming a GroEL-GroES-ADP complex, with 100 uM ADP, then adding an excess of single ring GroEL (SR1) to trap any released GroES; then 1 mM ATP was added. ...Details: The complexes were prepared by pre-forming a GroEL-GroES-ADP complex, with 100 uM ADP, then adding an excess of single ring GroEL (SR1) to trap any released GroES; then 1 mM ATP was added. Any GroEL-GroES complexes remaining have ADP in the GroES-bound ring and ATP in the other ring, modelling the in vivo ATP-binding reaction.
Oligomeric state: 14-mer / Number unique components: 2
Molecular weightExperimental: 1 MDa / Theoretical: 1 MDa

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Macromolecule #1: GroEL

MacromoleculeName: GroEL / type: protein_or_peptide / ID: 1 / Name.synonym: Chaperonin 60 / Number of copies: 1 / Oligomeric state: 14-mer / Recombinant expression: Yes
Source (natural)Organism: Escherichia coli (E. coli) / Cell: E. coli
Molecular weightExperimental: 800 KDa / Theoretical: 800 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)

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Macromolecule #2: GroES

MacromoleculeName: GroES / type: protein_or_peptide / ID: 2 / Name.synonym: Chaperonin 10 / Number of copies: 1 / Oligomeric state: 7-mer / Recombinant expression: Yes
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightExperimental: 70 KDa / Theoretical: 70 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)

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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

Concentration0.8 mg/mL
BufferpH: 7.5
Details: 12.5 mM HEPES, 5 mM KCl, 5 mM MgCl2, 100 uM ADP, then 2x molar excess of single ring mutant of GroEL (SR1), then 1 mM ATP just before vitrification. See sample details for an explanation of ...Details: 12.5 mM HEPES, 5 mM KCl, 5 mM MgCl2, 100 uM ADP, then 2x molar excess of single ring mutant of GroEL (SR1), then 1 mM ATP just before vitrification. See sample details for an explanation of the experimental design.
GridDetails: holey carbon film
VitrificationCryogen name: ETHANE / Chamber temperature: 100 K / Instrument: HOMEMADE PLUNGER / Details: Vitrification instrument: self made
Timed resolved state: The sample was vitrified within a few seconds of manually mixing in ATP.
Method: Blot for 1 second before plunging

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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 / Cs: 2 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 50000
Sample stageSpecimen holder: Eucentric / Specimen holder model: GATAN LIQUID NITROGEN
TemperatureAverage: 105 K
Image recordingCategory: FILM / Film or detector model: KODAK SO-163 FILM / Digitization - Scanner: ZEISS SCAI / Digitization - Sampling interval: 14 µm / Number real images: 71 / Average electron dose: 20 e/Å2 / Od range: 1 / Bits/pixel: 8
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: CTF multiplication and merging of 2D averages
Final angle assignmentDetails: Full coverage around a single axis, using mainly side views
Final reconstructionApplied symmetry - Point group: C7 (7 fold cyclic) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 23.5 Å / Resolution method: FSC 0.5 CUT-OFF / Software - Name: Spider / Details: Filtered back projection / Number images used: 1448

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Atomic model buiding 1

Initial modelPDB ID:
SoftwareName: Manual fitting in O
DetailsProtocol: Rigid body. The only movement relative to the crystal structure of GroEL-ES-ADP seen at this resolution is a twist of the free apical domains.
RefinementSpace: REAL / Protocol: RIGID BODY FIT
Output model

PDB-1gru:
SOLUTION STRUCTURE OF GROES-ADP7-GROEL-ATP7 COMPLEX BY CRYO-EM

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