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- EMDB-10417: GroEL map obtained using the Preassis method for grid preparation -

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Basic information

Entry
Database: EMDB / ID: EMD-10417
TitleGroEL map obtained using the Preassis method for grid preparation
Map dataGroEL map obtained using the Preassis method for sample preparation
Sample
  • Complex: GroEL
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)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsZhao J / Xu H / Carroni M / Zou X
Funding support Sweden, 2 items
OrganizationGrant numberCountry
Knut and Alice Wallenberg Foundation Sweden
Swedish Research Council Sweden
CitationJournal: Nat Commun / Year: 2021
Title: A simple pressure-assisted method for MicroED specimen preparation.
Authors: Jingjing Zhao / Hongyi Xu / Hugo Lebrette / Marta Carroni / Helena Taberman / Martin Högbom / Xiaodong Zou /
Abstract: Micro-crystal electron diffraction (MicroED) has shown great potential for structure determination of macromolecular crystals too small for X-ray diffraction. However, specimen preparation remains a ...Micro-crystal electron diffraction (MicroED) has shown great potential for structure determination of macromolecular crystals too small for X-ray diffraction. However, specimen preparation remains a major bottleneck. Here, we report a simple method for preparing MicroED specimens, named Preassis, in which excess liquid is removed through an EM grid with the assistance of pressure. We show the ice thicknesses can be controlled by tuning the pressure in combination with EM grids with appropriate carbon hole sizes. Importantly, Preassis can handle a wide range of protein crystals grown in various buffer conditions including those with high viscosity, as well as samples with low crystal concentrations. Preassis is a simple and universal method for MicroED specimen preparation, and will significantly broaden the applications of MicroED.
History
DepositionOct 28, 2019-
Header (metadata) releaseNov 18, 2020-
Map releaseNov 18, 2020-
UpdateOct 6, 2021-
Current statusOct 6, 2021Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.015
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.015
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10417.png

Downloads & links

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Map

FileDownload / File: emd_10417.map.gz / Format: CCP4 / Size: 163.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationGroEL map obtained using the Preassis method for sample preparation
Voxel sizeX=Y=Z: 0.83 Å
Density
Contour LevelBy AUTHOR: 0.0137 / Movie #1: 0.015
Minimum - Maximum-0.06091923 - 0.108942464
Average (Standard dev.)0.00043294486 (±0.00422564)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions350350350
Spacing350350350
CellA=B=C: 290.5 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.830.830.83
M x/y/z350350350
origin x/y/z0.0000.0000.000
length x/y/z290.500290.500290.500
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS350350350
D min/max/mean-0.0610.1090.000

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Supplemental data

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Mask #1

Fileemd_10417_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: postprocessed unmasked GroEL map obtained using the Preassis...

Fileemd_10417_additional_1.map
Annotationpostprocessed unmasked GroEL map obtained using the Preassis method for sample preparation
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 2 GroEL map obtained using the...

Fileemd_10417_half_map_1.map
AnnotationHalf map 2 GroEL map obtained using the Preassis method for sample preparation
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1 GroEL map obtained using the...

Fileemd_10417_half_map_2.map
AnnotationHalf map 1 GroEL map obtained using the Preassis method for sample preparation
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

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Supramolecule #1: GroEL

SupramoleculeName: GroEL / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Escherichia coli (E. coli)
Recombinant expressionOrganism: Escherichia coli (E. coli)

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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

BufferpH: 7.4
VitrificationCryogen name: ETHANE
DetailsSpecimen prepared using the novel freezing system Preassis

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K2 QUANTUM (4k x 4k) / Average electron dose: 40.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER / Details: ab-initio SGD
Initial angle assignmentType: MAXIMUM LIKELIHOOD
Final angle assignmentType: MAXIMUM LIKELIHOOD
Final reconstructionApplied symmetry - Point group: D7 (2x7 fold dihedral) / Resolution.type: BY AUTHOR / Resolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 59985
FSC plot (resolution estimation)

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