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- EMDB-22347: CryoEM structure of human light chain apoferritin calculated from... -

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Basic information

Entry
Database: EMDB / ID: EMD-22347
TitleCryoEM structure of human light chain apoferritin calculated from EER movies (super-resolution experiment, 2x supersampling with random subpixel information)
Map dataSharpened map
Sample
  • Complex: Human light chain apoferritin
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 2.8 Å
AuthorsGuo H / Rubinstein J
Funding support Canada, 2 items
OrganizationGrant numberCountry
Natural Sciences and Engineering Research Council (NSERC, Canada) Canada
Canada Excellence Research Chair Award Canada
Citation
Journal: IUCrJ / Year: 2020
Title: Electron-event representation data enable efficient cryoEM file storage with full preservation of spatial and temporal resolution.
Authors: Hui Guo / Erik Franken / Yuchen Deng / Samir Benlekbir / Garbi Singla Lezcano / Bart Janssen / Lingbo Yu / Zev A Ripstein / Yong Zi Tan / John L Rubinstein /
Abstract: Direct detector device (DDD) cameras have revolutionized electron cryomicroscopy (cryoEM) with their high detective quantum efficiency (DQE) and output of movie data. A high ratio of camera frame ...Direct detector device (DDD) cameras have revolutionized electron cryomicroscopy (cryoEM) with their high detective quantum efficiency (DQE) and output of movie data. A high ratio of camera frame rate (frames per second) to camera exposure rate (electrons per pixel per second) allows electron counting, which further improves the DQE and enables the recording of super-resolution information. Movie output also allows the correction of specimen movement and compensation for radiation damage. However, these movies come at the cost of producing large volumes of data. It is common practice to sum groups of successive camera frames to reduce the final frame rate, and therefore the file size, to one suitable for storage and image processing. This reduction in the temporal resolution of the camera requires decisions to be made during data acquisition that may result in the loss of information that could have been advantageous during image analysis. Here, experimental analysis of a new electron-event representation (EER) data format for electron-counting DDD movies is presented, which is enabled by new hardware developed by Thermo Fisher Scientific for their Falcon DDD cameras. This format enables the recording of DDD movies at the raw camera frame rate without sacrificing either spatial or temporal resolution. Experimental data demonstrate that the method retains super-resolution information and allows the correction of specimen movement at the physical frame rate of the camera while maintaining manageable file sizes. The EER format will enable the development of new methods that can utilize the full spatial and temporal resolution of DDD cameras.
#1: Journal: Biorxiv / Year: 2020
Title: Electron Event Representation (EER) data enables efficient cryoEM file storage with full preservation of spatial and temporal resolution
Authors: Guo H / Franken E / Deng Y / Benlekbir S / Lezcano GS / Janssen B / Yu L / Ripstein ZA / Tan YZ / Rubinstein JL
History
DepositionJul 24, 2020-
Header (metadata) releaseAug 5, 2020-
Map releaseAug 5, 2020-
UpdateSep 30, 2020-
Current statusSep 30, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.8
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.8
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_22347.png

Downloads & links

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Map

FileDownload / File: emd_22347.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationSharpened map
Voxel sizeX=Y=Z: 0.82 Å
Density
Contour LevelBy AUTHOR: 0.8 / Movie #1: 0.8
Minimum - Maximum-1.3597962 - 3.1129324
Average (Standard dev.)-0.00040820372 (±0.16319762)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions320320320
Spacing320320320
CellA=B=C: 262.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.820.820.82
M x/y/z320320320
origin x/y/z0.0000.0000.000
length x/y/z262.400262.400262.400
α/β/γ90.00090.00090.000
start NX/NY/NZ212211153
NX/NY/NZ80102186
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS320320320
D min/max/mean-1.3603.113-0.000

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Supplemental data

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Mask #1

Fileemd_22347_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 1

Fileemd_22347_half_map_1.map
AnnotationHalf map 1
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map 2

Fileemd_22347_half_map_2.map
AnnotationHalf map 2
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Human light chain apoferritin

EntireName: Human light chain apoferritin
Components
  • Complex: Human light chain apoferritin

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Supramolecule #1: Human light chain apoferritin

SupramoleculeName: Human light chain apoferritin / type: complex / ID: 1 / Parent: 0
Source (natural)Organism: Homo sapiens (human)
Recombinant expressionOrganism: Homo sapiens (human) / Recombinant cell: HEK293F
Molecular weightTheoretical: 480 KDa

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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

Concentration10 mg/mL
BufferpH: 8
GridModel: Homemade / Material: COPPER/RHODIUM / Support film - Material: CARBON / Support film - topology: HOLEY / Support film - Film thickness: 30.0 nm / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Atmosphere: OTHER
VitrificationCryogen name: ETHANE / Chamber humidity: 90 % / Chamber temperature: 295 K / Instrument: GATAN CRYOPLUNGE 3

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Calibrated defocus max: 1.1 µm / Calibrated defocus min: 0.6 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal magnification: 47000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Image recordingFilm or detector model: FEI FALCON IV (4k x 4k) / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Number grids imaged: 1 / Number real images: 100 / Average exposure time: 24.0 sec. / Average electron dose: 45.0 e/Å2
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 247312
CTF correctionSoftware - Name: cryoSPARC (ver. 2.15)
Initial angle assignmentType: PROJECTION MATCHING / Software - Name: cryoSPARC (ver. 2.15)
Final 3D classificationNumber classes: 50 / Avg.num./class: 4375 / Software - Name: cryoSPARC (ver. 2.15)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: cryoSPARC (ver. 2.15)
Final reconstructionApplied symmetry - Point group: O (octahedral) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 2.8 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: cryoSPARC (ver. 2.15) / Number images used: 214410
FSC plot (resolution estimation)

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