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- PDB-8wu4: Cryo-EM structure of native H. thermoluteolus TH-1 GroEL -

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Basic information

Entry
Database: PDB / ID: 8wu4
TitleCryo-EM structure of native H. thermoluteolus TH-1 GroEL
ComponentsChaperonin GroEL
KeywordsCHAPERONE / Chaperonin / ATPase / protein folding
Function / homology
Function and homology information


chaperonin ATPase / isomerase activity / ATP-dependent protein folding chaperone / unfolded protein binding / protein refolding / ATP binding / cytoplasm
Similarity search - Function
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
Similarity search - Domain/homology
Biological speciesHydrogenophilus thermoluteolus (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.3 Å
AuthorsLiao, Z. / Gopalasingam, C.C. / Kameya, M. / Gerle, C. / Shigematsu, H. / Ishii, M. / Arakawa, T. / Fushinobu, S.
Funding support Japan, 2items
OrganizationGrant numberCountry
Japan Society for the Promotion of Science (JSPS)23H00322 Japan
Japan Science and TechnologyJPMJSP2108 Japan
CitationJournal: Structure / Year: 2024
Title: Structural insights into thermophilic chaperonin complexes.
Authors: Zengwei Liao / Chai C Gopalasingam / Masafumi Kameya / Christoph Gerle / Hideki Shigematsu / Masaharu Ishii / Takatoshi Arakawa / Shinya Fushinobu /
Abstract: Group I chaperonins are dual heptamer protein complexes that play significant roles in protein homeostasis. The structure and function of the Escherichia coli chaperonin are well characterized. ...Group I chaperonins are dual heptamer protein complexes that play significant roles in protein homeostasis. The structure and function of the Escherichia coli chaperonin are well characterized. However, the dynamic properties of chaperonins, such as large ATPase-dependent conformational changes by binding of lid-like co-chaperonin GroES, have made structural analyses challenging, and our understanding of these changes during the turnover of chaperonin complex formation is limited. In this study, we used single-particle cryogenic electron microscopy to investigate the structures of GroES-bound chaperonin complexes from the thermophilic hydrogen-oxidizing bacteria Hydrogenophilus thermoluteolus and Hydrogenobacter thermophilus in the presence of ATP and AMP-PNP. We captured the structure of an intermediate state chaperonin complex, designated as an asymmetric football-shaped complex, and performed analyses to decipher the dynamic structural variations. Our structural analyses of inter- and intra-subunit communications revealed a unique mechanism of complex formation through the binding of a second GroES to a bullet-shaped complex.
History
DepositionOct 20, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 27, 2024Provider: repository / Type: Initial release
Revision 1.1Jun 19, 2024Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Chaperonin GroEL
B: Chaperonin GroEL
C: Chaperonin GroEL
D: Chaperonin GroEL
E: Chaperonin GroEL
F: Chaperonin GroEL
G: Chaperonin GroEL
H: Chaperonin GroEL
I: Chaperonin GroEL
J: Chaperonin GroEL
K: Chaperonin GroEL
L: Chaperonin GroEL
M: Chaperonin GroEL
N: Chaperonin GroEL


Theoretical massNumber of molelcules
Total (without water)781,71914
Polymers781,71914
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein
Chaperonin GroEL / 60 kDa chaperonin / Chaperonin-60 / Cpn60


Mass: 55837.070 Da / Num. of mol.: 14 / Source method: isolated from a natural source / Source: (natural) Hydrogenophilus thermoluteolus (bacteria) / Strain: TH-1 / References: UniProt: A0A2Z6DW38, chaperonin ATPase

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Native H. thermoluteolus GroEL / Type: COMPLEX / Entity ID: all / Source: NATURAL
Molecular weightValue: 809 kDa/nm / Experimental value: NO
Source (natural)Organism: Hydrogenophilus thermoluteolus (bacteria) / Strain: TH-1
Buffer solutionpH: 7.4
Buffer component
IDConc.NameFormulaBuffer-ID
120 mM4-(2-hydroxyethyl)-1-piperazineethanesulfonic acidC8H18N2O4S1
2100 mMPotassium chlorideKCl1
35 mMMagnesium ChlorideMgCl21
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: PIB-10, hard mode / Grid material: COPPER / Grid mesh size: 200 divisions/in. / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K / Details: blot force 10, blot time 15 s

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Electron microscopy imaging

Experimental equipment
Model: Talos Arctica / Image courtesy: FEI Company
MicroscopyModel: FEI TALOS ARCTICA
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 200 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal magnification: 120000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Specimen holderCryogen: NITROGEN
Image recordingAverage exposure time: 4.95 sec. / Electron dose: 50 e/Å2 / Film or detector model: FEI FALCON IV (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 3418
Image scansWidth: 4096 / Height: 4096

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Processing

EM software
IDNameVersionCategoryDetails
2EPUimage acquisition
4cryoSPARC4.3CTF correctionPatch CTF
7UCSF ChimeraX1.6model fitting
9cryoSPARC4.3initial Euler assignment
10cryoSPARC4.3final Euler assignment
11cryoSPARC4.3classification
12cryoSPARC4.33D reconstruction
13PHENIX1.2.0model refinementreal_space_refinement
14ISOLDE1.6model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: D7 (2x7 fold dihedral)
3D reconstructionResolution: 3.3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 40366 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL / Details: Initial fitting was done using UCSF ChimeraX
Atomic model buildingSource name: AlphaFold / Type: in silico model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00255146
ELECTRON MICROSCOPYf_angle_d0.47174438
ELECTRON MICROSCOPYf_dihedral_angle_d3.6847784
ELECTRON MICROSCOPYf_chiral_restr0.048988
ELECTRON MICROSCOPYf_plane_restr0.0049730

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