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- EMDB-6802: Near-atomic resolution reconstruction of mixed over-/under-focuse... -

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Basic information

Entry
Database: EMDB / ID: 6802
TitleNear-atomic resolution reconstruction of mixed over-/under-focused apoferritin
Map data
SampleNear-atomic resolution reconstruction of over-focused apoferritin:
Function / homologyFerritin-like superfamily / Ferritin-like / Ferritin-like diiron domain / Ferritin/DPS protein domain / Ferritin / Ferritin, conserved site / Ferritin-like domain / Ferritin iron-binding regions signature 2. / Ferritin iron-binding regions signature 1. / Ferritin-like diiron domain profile. ...Ferritin-like superfamily / Ferritin-like / Ferritin-like diiron domain / Ferritin/DPS protein domain / Ferritin / Ferritin, conserved site / Ferritin-like domain / Ferritin iron-binding regions signature 2. / Ferritin iron-binding regions signature 1. / Ferritin-like diiron domain profile. / Scavenging by Class A Receptors / Golgi Associated Vesicle Biogenesis / Neutrophil degranulation / Iron uptake and transport / iron ion import / autolysosome / intracellular ferritin complex / ferroxidase / intracellular sequestering of iron ion / ferroxidase activity / negative regulation of fibroblast proliferation / ferric iron binding / cellular iron ion homeostasis / tertiary granule lumen / ficolin-1-rich granule lumen / immune response / iron ion binding / negative regulation of cell proliferation / neutrophil degranulation / extracellular exosome / extracellular region / nucleus / cytosol / Ferritin heavy chain
Function and homology information
SourceHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / 3 Å resolution
AuthorsFan X / Zhao LY
CitationJournal: Structure / Year: 2017
Title: Near-Atomic Resolution Structure Determination in Over-Focus with Volta Phase Plate by Cs-Corrected Cryo-EM.
Authors: Xiao Fan / Lingyun Zhao / Chuan Liu / Jin-Can Zhang / Kelong Fan / Xiyun Yan / Hai-Lin Peng / Jianlin Lei / Hong-Wei Wang
Abstract: Volta phase plate (VPP) is a recently developed transmission electron microscope (TEM) apparatus that can significantly enhance the image contrast of biological samples in cryoelectron microscopy, ...Volta phase plate (VPP) is a recently developed transmission electron microscope (TEM) apparatus that can significantly enhance the image contrast of biological samples in cryoelectron microscopy, and therefore provide the possibility to solve structures of relatively small macromolecules at high-resolution. In this work, we performed theoretical analysis and found that using phase plate on objective lens spherical aberration (Cs)-corrected TEM may gain some interesting optical properties, including the over-focus imaging of macromolecules. We subsequently evaluated the imaging strategy of frozen-hydrated apo-ferritin with VPP on a Cs-corrected TEM and obtained the structure of apo-ferritin at near-atomic resolution from both under- and over-focused dataset, illustrating the feasibility and new potential of combining VPP with Cs-corrected TEM for high-resolution cryo-EM.
DateDeposition: Jul 21, 2017 / Header (metadata) release: Nov 22, 2017 / Map release: Nov 22, 2017 / Last update: Nov 22, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.08
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.08
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_6802.map.gz (map file in CCP4 format, 55297 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
240 pix
0.88 Å/pix.
= 211.2 Å
240 pix
0.88 Å/pix.
= 211.2 Å
240 pix
0.88 Å/pix.
= 211.2 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.88 Å
Density
Contour Level:0.08 (by author), 0.08 (movie #1):
Minimum - Maximum-0.14438719 - 0.29314175
Average (Standard dev.)0.00074269285 (0.020843051)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions240240240
Origin000
Limit239239239
Spacing240240240
CellA=B=C: 211.2 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.880.880.88
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z211.200211.200211.200
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.1440.2930.001

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Supplemental data

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Sample components

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Entire Near-atomic resolution reconstruction of over-focused apoferritin

EntireName: Near-atomic resolution reconstruction of over-focused apoferritin
Number of components: 1

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Component #1: protein, Near-atomic resolution reconstruction of over-focused ap...

ProteinName: Near-atomic resolution reconstruction of over-focused apoferritin
Recombinant expression: No
SourceSpecies: Homo sapiens (human)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria) / Strain: BL21(DE3)

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionSpecimen conc.: 1 mg/ml / pH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 283 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 25 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 0.001 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: FEI FALCON II (4k x 4k)

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Image acquisition

Image acquisitionSampling size: 14 microns
Raw dataEMPIAR-10122 (Title: Mixed under-/over-focused data collected by VPP-Cs-corrector coupled EM.
Data size: 34.4
Data #1: Dose-weighted image from MotionCor2 for Mixed under-/over-focused data [micrographs - single frame])

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 63015
3D reconstructionSoftware: RELION / Resolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot
(resolution estimation)

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