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- EMDB-15939: CryoEM structure of nucleotide-free GroEL-Rubisco. -

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Basic information

Entry
Database: EMDB / ID: EMD-15939
TitleCryoEM structure of nucleotide-free GroEL-Rubisco.
Map dataCryoEM structure of GroEL bound to non-native Rubisco.
Sample
  • Complex: GroEL
    • Protein or peptide: Chaperonin GroEL
KeywordsGroEL / Rubisco / Chaperone / complex
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 / identical protein binding / membrane / 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)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.4 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) releaseOct 25, 2023-
Map releaseOct 25, 2023-
UpdateDec 27, 2023-
Current statusDec 27, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_15939.png

Downloads & links

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Map

FileDownload / File: emd_15939.map.gz / Format: CCP4 / Size: 216 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationCryoEM structure of GroEL bound to non-native Rubisco.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.34 Å/pix.
x 384 pix.
= 514.56 Å		1.34 Å/pix.
x 384 pix.
= 514.56 Å		1.34 Å/pix.
x 384 pix.
= 514.56 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.34 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0317
Minimum - Maximum-0.09837879 - 0.16313978
Average (Standard dev.)0.00020874589 (±0.003962051)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions384384384
Spacing384384384
CellA=B=C: 514.56 Å
α=β=γ: 90.0 °

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Supplemental data

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Mask #1

Fileemd_15939_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: CryoEM structure of GroEL bound to non-native Rubisco.

Fileemd_15939_half_map_1.map
AnnotationCryoEM structure of GroEL bound to non-native Rubisco.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: CryoEM structure of GroEL bound to non-native Rubisco.

Fileemd_15939_half_map_2.map
AnnotationCryoEM structure of GroEL bound to non-native 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

EntireName: GroEL
Components
  • Complex: GroEL
    • Protein or peptide: Chaperonin GroEL

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Supramolecule #1: GroEL

SupramoleculeName: GroEL / type: complex / ID: 1 / Parent: 0 / Macromolecule list: all
Source (natural)Organism: Escherichia coli (E. coli) / Strain: K12
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 (E. coli) / Strain: K12
Molecular weightTheoretical: 57.260504 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: AAKDVKFGND ARVKMLRGVN VLADAVKVTL GPKGRNVVLD KSFGAPTITK DGVSVAREIE LEDKFENMGA QMVKEVASKA NDAAGDGTT TATVLAQAII TEGLKAVAAG MNPMDLKRGI DKAVTAAVEE LKALSVPCSD SKAIAQVGTI SANSDETVGK L IAEAMDKV ...String:
AAKDVKFGND ARVKMLRGVN VLADAVKVTL GPKGRNVVLD KSFGAPTITK DGVSVAREIE LEDKFENMGA QMVKEVASKA NDAAGDGTT TATVLAQAII TEGLKAVAAG MNPMDLKRGI DKAVTAAVEE LKALSVPCSD SKAIAQVGTI SANSDETVGK L IAEAMDKV GKEGVITVED GTGLQDELDV VEGMQFDRGY LSPYFINKPE TGAVELESPF ILLADKKISN IREMLPVLEA VA KAGKPLL IIAEDVEGEA LATLVVNTMR GIVKVAAVKA PGFGDRRKAM LQDIATLTGG TVISEEIGME LEKATLEDLG QAK RVVINK DTTTIIDGVG EEAAIQGRVA QIRQQIEEAT SDYDREKLQE RVAKLAGGVA VIKVGAATEV EMKEKKARVE DALH ATRAA VEEGVVAGGG VALIRVASKL ADLRGQNEDQ NVGIKVALRA MEAPLRQIVL NCGEEPSVVA NTVKGGDGNY GYNAA TEEY GNMIDMGILD PTKVTRSALQ YAASVAGLMI TTECMVTDLP KNDAADLGAA GGMGGMGGMG GMM

UniProtKB: Chaperonin GroEL

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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
Image recordingFilm or detector model: GATAN K2 BASE (4k x 4k) / Number grids imaged: 2 / Average electron dose: 40.2 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 70.0 µm / Illumination mode: SPOT SCAN / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 1.4000000000000001 µm
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: PDB ENTRY
PDB model - PDB ID:

1oel

Final reconstructionNumber classes used: 2 / Applied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 4.4 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 65453
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final 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)
FSC plot (resolution estimation)

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