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- PDB-6tmw: Structure of the chaperonin gp146 from the bacteriophage EL (Pseu... -

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Basic information

Entry
Database: PDB / ID: 6tmw
TitleStructure of the chaperonin gp146 from the bacteriophage EL (Pseudomonas aeruginosa) in complex with ADP
ComponentsPutative GroEL-like chaperonine protein
KeywordsCHAPERONE / molecular chaperone / ATPase / chaperonin
Function / homology
Function and homology information


ATP-dependent protein folding chaperone / protein refolding / ATP binding / metal ion binding / identical protein binding
Similarity search - Function
Chaperone tailless complex polypeptide 1 (TCP-1) / 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
ADENOSINE-5'-DIPHOSPHATE / Putative GroEL-like chaperonine protein
Similarity search - Component
Biological speciesPseudomonas phage EL (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 5.91 Å
AuthorsBracher, A. / Wang, H. / Paul, S.S. / Wischnewski, N. / Hartl, F.U. / Hayer-Hartl, M.
CitationJournal: PLoS One / Year: 2020
Title: Structure and conformational cycle of a bacteriophage-encoded chaperonin.
Authors: Andreas Bracher / Simanta S Paul / Huping Wang / Nadine Wischnewski / F Ulrich Hartl / Manajit Hayer-Hartl /
Abstract: Chaperonins are ubiquitous molecular chaperones found in all domains of life. They form ring-shaped complexes that assist in the folding of substrate proteins in an ATP-dependent reaction cycle. Key ...Chaperonins are ubiquitous molecular chaperones found in all domains of life. They form ring-shaped complexes that assist in the folding of substrate proteins in an ATP-dependent reaction cycle. Key to the folding cycle is the transient encapsulation of substrate proteins by the chaperonin. Here we present a structural and functional characterization of the chaperonin gp146 (ɸEL) from the phage EL of Pseudomonas aeruginosa. ɸEL, an evolutionarily distant homolog of bacterial GroEL, is active in ATP hydrolysis and prevents the aggregation of denatured protein in a nucleotide-dependent manner. However, ɸEL failed to refold the encapsulation-dependent model substrate rhodanese and did not interact with E. coli GroES, the lid-shaped co-chaperone of GroEL. ɸEL forms tetradecameric double-ring complexes, which dissociate into single rings in the presence of ATP. Crystal structures of ɸEL (at 3.54 and 4.03 Å) in presence of ATP•BeFx revealed two distinct single-ring conformational states, both with open access to the ring cavity. One state showed uniform ATP-bound subunit conformations (symmetric state), whereas the second combined distinct ATP- and ADP-bound subunit conformations (asymmetric state). Cryo-electron microscopy of apo-ɸEL revealed a double-ring structure composed of rings in the asymmetric state (3.45 Å resolution). We propose that the phage chaperonin undergoes nucleotide-dependent conformational switching between double- and single rings and functions in aggregation prevention without substrate protein encapsulation. Thus, ɸEL may represent an evolutionarily more ancient chaperonin prior to acquisition of the encapsulation mechanism.
History
DepositionDec 5, 2019Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 22, 2020Provider: repository / Type: Initial release
Revision 1.1May 6, 2020Group: Database references / Category: citation / citation_author
Item: _citation.journal_volume / _citation.page_first ..._citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation_author.identifier_ORCID
Revision 1.2May 22, 2024Group: Data collection / Database references
Category: chem_comp_atom / chem_comp_bond ...chem_comp_atom / chem_comp_bond / database_2 / pdbx_database_related
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession / _pdbx_database_related.content_type

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Structure visualization

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Assembly

Deposited unit
A: Putative GroEL-like chaperonine protein
B: Putative GroEL-like chaperonine protein
C: Putative GroEL-like chaperonine protein
D: Putative GroEL-like chaperonine protein
E: Putative GroEL-like chaperonine protein
F: Putative GroEL-like chaperonine protein
G: Putative GroEL-like chaperonine protein
H: Putative GroEL-like chaperonine protein
I: Putative GroEL-like chaperonine protein
J: Putative GroEL-like chaperonine protein
K: Putative GroEL-like chaperonine protein
L: Putative GroEL-like chaperonine protein
M: Putative GroEL-like chaperonine protein
N: Putative GroEL-like chaperonine protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)870,63428
Polymers864,65314
Non-polymers5,98114
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: light scattering
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area47470 Å2
ΔGint-268 kcal/mol
Surface area325570 Å2
MethodPISA

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Components

#1: Protein
Putative GroEL-like chaperonine protein


Mass: 61760.941 Da / Num. of mol.: 14
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pseudomonas phage EL (virus) / Plasmid: pET22b / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q2Z0T5
#2: Chemical
ChemComp-ADP / ADENOSINE-5'-DIPHOSPHATE


Mass: 427.201 Da / Num. of mol.: 14 / Source method: obtained synthetically / Formula: C10H15N5O10P2 / Comment: ADP, energy-carrying molecule*YM
Has ligand of interestN

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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: The chaperonin gp146 from the bacteriophage EL (Pseudomonas aeruginosa) in complex with ADP
Type: COMPLEX / Entity ID: #1 / Source: RECOMBINANT
Molecular weightValue: 0.864 MDa / Experimental value: YES
Source (natural)Organism: Pseudomonas phage EL (virus)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria) / Strain: Bl21 (DE3) / Plasmid: pET22b
Buffer solutionpH: 7.5
Buffer component
IDConc.NameFormulaBuffer-ID
120 mMHEPESC8H18N2O4S1
250 mMsodium chlorideNaCl1
30.5 mMEDTAC10H16N2O81
44 mMmagnesium chlorideMgCl21
52 mMADPC10H15N5O10P21
60.04 %beta-octyl glucosideC14H28O61
SpecimenConc.: 1.125 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 300 divisions/in. / Grid type: Quantifoil R2/1
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 298 K / Details: Blot time was 2 sec.

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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
Image recordingAverage exposure time: 0.074769 sec. / Electron dose: 1.6 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 892

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Processing

EM software
IDNameVersionCategory
1Gautomatch0.56particle selection
2EPU1.3.0.77image acquisition
4CTFFIND4.1CTF correction
7UCSF Chimera1.13.1model fitting
9RELION3initial Euler assignment
10RELION3final Euler assignment
11RELION3classification
12RELION33D reconstruction
13REFMAC5.8.0155model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 429456
SymmetryPoint symmetry: D7 (2x7 fold dihedral)
3D reconstructionResolution: 5.91 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 170957 / Num. of class averages: 1 / Symmetry type: POINT
Atomic model buildingProtocol: RIGID BODY FIT / Space: REAL / Details: Jelly body refinement D7 symmetry NCS restraints

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