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- EMDB-15940: CryoEM structure of GroEL-ADP.BeF3-Rubisco. -

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Basic information

Entry
Database: EMDB / ID: EMD-15940
TitleCryoEM structure of GroEL-ADP.BeF3-Rubisco.
Map dataCryoEM reconstruction of GroEL-ADP.BeFx-Rubisco.
Sample
  • Complex: GroEL-ADP.BeFx-Rubisco
    • Protein or peptide: Chaperonin GroEL
  • Ligand: BERYLLIUM TRIFLUORIDE ION
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: POTASSIUM IONPotassium
  • Ligand: water
KeywordsGroEL / nucleotide / Rubisco / Chameleon / CHAPERONE
Function / homology
Function and homology information


GroEL-GroES complex / chaperonin ATPase / virion assembly / chaperone cofactor-dependent protein refolding / isomerase activity / ATP-dependent protein folding chaperone / response to radiation / unfolded protein binding / protein folding / response to heat ...GroEL-GroES complex / chaperonin ATPase / virion assembly / chaperone cofactor-dependent protein refolding / isomerase activity / ATP-dependent protein folding chaperone / response to radiation / unfolded protein binding / protein folding / response to heat / protein refolding / magnesium ion binding / ATP hydrolysis activity / ATP binding / membrane / identical protein binding / cytosol
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 speciesEscherichia coli (E. coli) / Escherichia coli K-12 (bacteria)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsGardner S / Saibil HR
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Biotechnology and Biological Sciences Research Council (BBSRC)BB/M009513/1 United Kingdom
CitationJournal: Proc Natl Acad Sci U S A / Year: 2023
Title: Structural basis of substrate progression through the bacterial chaperonin cycle.
Authors: Scott Gardner / Michele C Darrow / Natalya Lukoyanova / Konstantinos Thalassinos / Helen R Saibil /
Abstract: The bacterial chaperonin GroEL-GroES promotes protein folding through ATP-regulated cycles of substrate protein binding, encapsulation, and release. Here, we have used cryoEM to determine structures ...The bacterial chaperonin GroEL-GroES promotes protein folding through ATP-regulated cycles of substrate protein binding, encapsulation, and release. Here, we have used cryoEM to determine structures of GroEL, GroEL-ADP·BeF, and GroEL-ADP·AlF-GroES all complexed with the model substrate Rubisco. Our structures provide a series of snapshots that show how the conformation and interactions of non-native Rubisco change as it proceeds through the GroEL-GroES reaction cycle. We observe specific charged and hydrophobic GroEL residues forming strong initial contacts with non-native Rubisco. Binding of ATP or ADP·BeF to GroEL-Rubisco results in the formation of an intermediate GroEL complex displaying striking asymmetry in the ATP/ADP·BeF-bound ring. In this ring, four GroEL subunits bind Rubisco and the other three are in the GroES-accepting conformation, suggesting how GroEL can recruit GroES without releasing bound substrate. Our cryoEM structures of stalled GroEL-ADP·AlF-Rubisco-GroES complexes show Rubisco folding intermediates interacting with GroEL-GroES via different sets of residues.
History
DepositionOct 11, 2022-
Header (metadata) releaseDec 27, 2023-
Map releaseDec 27, 2023-
UpdateDec 27, 2023-
Current statusDec 27, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15940.png

Downloads & links

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Map

FileDownload / File: emd_15940.map.gz / Format: CCP4 / Size: 343 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM reconstruction of GroEL-ADP.BeFx-Rubisco.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.22 Å/pix.
x 448 pix.
= 546.291 Å		1.22 Å/pix.
x 448 pix.
= 546.291 Å		1.22 Å/pix.
x 448 pix.
= 546.291 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.2194 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0218
Minimum - Maximum-0.029073583 - 0.15881711
Average (Standard dev.)0.000022599463 (±0.003434118)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions448448448
Spacing448448448
CellA=B=C: 546.2912 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_15940_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: CryoEM reconstruction of GroEL-ADP.BeFx-Rubisco.

Fileemd_15940_half_map_1.map
AnnotationCryoEM reconstruction of GroEL-ADP.BeFx-Rubisco.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: CryoEM reconstruction of GroEL-ADP.BeFx-Rubisco.

Fileemd_15940_half_map_2.map
AnnotationCryoEM reconstruction of GroEL-ADP.BeFx-Rubisco.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : GroEL-ADP.BeFx-Rubisco

EntireName: GroEL-ADP.BeFx-Rubisco
Components
  • Complex: GroEL-ADP.BeFx-Rubisco
    • Protein or peptide: Chaperonin GroEL
  • Ligand: BERYLLIUM TRIFLUORIDE ION
  • Ligand: ADENOSINE-5'-DIPHOSPHATE
  • Ligand: MAGNESIUM ION
  • Ligand: POTASSIUM IONPotassium
  • Ligand: water

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Supramolecule #1: GroEL-ADP.BeFx-Rubisco

SupramoleculeName: GroEL-ADP.BeFx-Rubisco / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 802 KDa

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Macromolecule #1: Chaperonin GroEL

MacromoleculeName: Chaperonin GroEL / type: protein_or_peptide / ID: 1 / Number of copies: 14 / Enantiomer: LEVO / EC number: chaperonin ATPase
Source (natural)Organism: Escherichia coli K-12 (bacteria) / Strain: K-12
Molecular weightTheoretical: 57.391711 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MAAKDVKFGN DARVKMLRGV NVLADAVKVT LGPKGRNVVL DKSFGAPTIT KDGVSVAREI ELEDKFENMG AQMVKEVASK ANDAAGDGT TTATVLAQAI ITEGLKAVAA GMNPMDLKRG IDKAVTAAVE ELKALSVPCS DSKAIAQVGT ISANSDETVG K LIAEAMDK ...String:
MAAKDVKFGN DARVKMLRGV NVLADAVKVT LGPKGRNVVL DKSFGAPTIT KDGVSVAREI ELEDKFENMG AQMVKEVASK ANDAAGDGT TTATVLAQAI ITEGLKAVAA GMNPMDLKRG IDKAVTAAVE ELKALSVPCS DSKAIAQVGT ISANSDETVG K LIAEAMDK VGKEGVITVE DGTGLQDELD VVEGMQFDRG YLSPYFINKP ETGAVELESP FILLADKKIS NIREMLPVLE AV AKAGKPL LIIAEDVEGE ALATLVVNTM RGIVKVAAVK APGFGDRRKA MLQDIATLTG GTVISEEIGM ELEKATLEDL GQA KRVVIN KDTTTIIDGV GEEAAIQGRV AQIRQQIEEA TSDYDREKLQ ERVAKLAGGV AVIKVGAATE VEMKEKKARV EDAL HATRA AVEEGVVAGG GVALIRVASK LADLRGQNED QNVGIKVALR AMEAPLRQIV LNCGEEPSVV ANTVKGGDGN YGYNA ATEE YGNMIDMGIL DPTKVTRSAL QYAASVAGLM ITTECMVTDL PKNDAADLGA AGGMGGMGGM GGMM

UniProtKB: Chaperonin GroEL

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Macromolecule #2: BERYLLIUM TRIFLUORIDE ION

MacromoleculeName: BERYLLIUM TRIFLUORIDE ION / type: ligand / ID: 2 / Number of copies: 7 / Formula: BEF
Molecular weightTheoretical: 66.007 Da
Chemical component information

ChemComp-BEF:
BERYLLIUM TRIFLUORIDE ION

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Macromolecule #3: ADENOSINE-5'-DIPHOSPHATE

MacromoleculeName: ADENOSINE-5'-DIPHOSPHATE / type: ligand / ID: 3 / Number of copies: 14 / Formula: ADP
Molecular weightTheoretical: 427.201 Da
Chemical component information

ChemComp-ADP:
ADENOSINE-5'-DIPHOSPHATE / ADP, energy-carrying molecule*YM / Adenosine diphosphate

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Macromolecule #4: MAGNESIUM ION

MacromoleculeName: MAGNESIUM ION / type: ligand / ID: 4 / Number of copies: 14 / Formula: MG
Molecular weightTheoretical: 24.305 Da

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Macromolecule #5: POTASSIUM ION

MacromoleculeName: POTASSIUM ION / type: ligand / ID: 5 / Number of copies: 14 / Formula: K
Molecular weightTheoretical: 39.098 Da

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Macromolecule #6: water

MacromoleculeName: water / type: ligand / ID: 6 / Number of copies: 7 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
WATER / Water

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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

Concentration2.7 mg/mL
BufferpH: 7.5
VitrificationCryogen name: ETHANE / Chamber humidity: 80 % / Chamber temperature: 293 K / Instrument: SPOTITON
Details: The grid was prepared using a chameleon (SPT Labtech)..

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.4000000000000001 µm
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Number grids imaged: 2 / Average electron dose: 40.2 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 7712622
Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

1ss8

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 3D classificationNumber classes: 8 / Avg.num./class: 25000 / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. 3.3.2)
Final reconstructionNumber classes used: 2 / Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 202582
FSC plot (resolution estimation)

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