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- PDB-8wux: Cryo-EM structure of H. thermophilus GroEL-GroES bullet complex -

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Basic information

Entry
Database: PDB / ID: 8wux
TitleCryo-EM structure of H. thermophilus GroEL-GroES bullet complex
Components
  • Chaperonin GroEL
  • Co-chaperonin GroES
KeywordsCHAPERONE / Chaperonin / ATPase / protein folding
Function / homology
Function and homology information


chaperonin ATPase / protein folding chaperone / isomerase activity / ATP-dependent protein folding chaperone / unfolded protein binding / protein refolding / ATP binding / cytoplasm
Similarity search - Function
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 ...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
PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER / : / Co-chaperonin GroES / Chaperonin GroEL
Similarity search - Component
Biological speciesHydrogenobacter thermophilus TK-6 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.6 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 21, 2023Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Mar 27, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Chaperonin GroEL
H: Chaperonin GroEL
a: Co-chaperonin GroES
B: Chaperonin GroEL
I: Chaperonin GroEL
b: Co-chaperonin GroES
C: Chaperonin GroEL
J: Chaperonin GroEL
c: Co-chaperonin GroES
D: Chaperonin GroEL
K: Chaperonin GroEL
d: Co-chaperonin GroES
E: Chaperonin GroEL
L: Chaperonin GroEL
e: Co-chaperonin GroES
F: Chaperonin GroEL
M: Chaperonin GroEL
f: Co-chaperonin GroES
G: Chaperonin GroEL
N: Chaperonin GroEL
g: Co-chaperonin GroES
hetero molecules


Theoretical massNumber of molelcules
Total (without water)873,19456
Polymers865,49421
Non-polymers7,70135
Water0
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: 56600.918 Da / Num. of mol.: 14
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Hydrogenobacter thermophilus TK-6 (bacteria)
Strain: TK-6 / Gene: groL, groEL, HTH_1794 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: D3DK86, chaperonin ATPase
#2: Protein
Co-chaperonin GroES / 10 kDa chaperonin / Chaperonin-10 / Cpn10


Mass: 10440.112 Da / Num. of mol.: 7
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Hydrogenobacter thermophilus TK-6 (bacteria)
Strain: TK-6 / Gene: groS, groES, HTH_1793 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: D3DK85
#3: Chemical
ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 14 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION
#4: Chemical
ChemComp-ANP / PHOSPHOAMINOPHOSPHONIC ACID-ADENYLATE ESTER


Mass: 506.196 Da / Num. of mol.: 14 / Source method: obtained synthetically / Formula: C10H17N6O12P3 / Feature type: SUBJECT OF INVESTIGATION / Comment: AMP-PNP, energy-carrying molecule analogue*YM
#5: Chemical
ChemComp-K / POTASSIUM ION


Mass: 39.098 Da / Num. of mol.: 7 / Source method: obtained synthetically / Formula: K / Feature type: SUBJECT OF INVESTIGATION
Has ligand of interestY

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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

Component
IDNameTypeEntity IDParent-IDSource
1H. thermophilus GroEL-GroES bullet complexCOMPLEX#1-#20RECOMBINANT
2chaperonin GroELCOMPLEX#11RECOMBINANT
3co-chaperonin GroESCOMPLEX#21RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)Organism: Hydrogenobacter thermophilus TK-6 (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.4
Buffer component
IDConc.NameBuffer-ID
120 mMHEPES1
2100 mMpotassium chloride1
35 mMmagnesium chloride1
42 mMadenylyl-imidodiphosphate1
51 mMdithiothreitol1
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: glow discharged at 7 Pa, 10 mA, 10 sec by a JEOL JEC-3000FC auto fine coater
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: 281 K

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Electron microscopy imaging

MicroscopyModel: JEOL CRYO ARM 300
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 60000 X / Nominal defocus max: 1600 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm
Specimen holderCryogen: NITROGEN
Image recordingAverage exposure time: 2.264 sec. / Electron dose: 49.97 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 5650

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Processing

EM software
IDNameVersionCategoryDetails
1cryoSPARCparticle selection
2SerialEMimage acquisition
4cryoSPARC4.3CTF correction
7UCSF ChimeraX1.6model fitting
9cryoSPARC4.3initial Euler assignment
10cryoSPARC4.3final Euler assignment
11cryoSPARC4.3classification
12cryoSPARC4.33D reconstruction
13ISOLDE1.6model refinement
14PHENIX1.2model refinementreal_space_refine
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C7 (7 fold cyclic)
3D reconstructionResolution: 2.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 20238 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT
Atomic model buildingPDB-ID: 7pbj

7pbj


Accession code: 7pbj / Source name: PDB / Type: experimental model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00361607
ELECTRON MICROSCOPYf_angle_d0.58183258
ELECTRON MICROSCOPYf_dihedral_angle_d4.5828575
ELECTRON MICROSCOPYf_chiral_restr0.04210010
ELECTRON MICROSCOPYf_plane_restr0.00310668

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