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- EMDB-14332: 1.58 A STRUCTURE OF HUMAN APOFERRITIN OBTAINED FROM TITAN KRIOS 2... -

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Basic information

Entry
Database: EMDB / ID: EMD-14332
Title1.58 A STRUCTURE OF HUMAN APOFERRITIN OBTAINED FROM TITAN KRIOS 2 AT eBIC, DLS UNDER COMMISSIONING SESSION CM26464-2
Map dataApoF structure post processed in Relion
Sample
  • Complex: human Apoferritin
    • Protein or peptide: Ferritin heavy chain, N-terminally processed
  • Ligand: SODIUM ION
  • Ligand: water
KeywordsProtein standard / METAL BINDING PROTEIN
Function / homology
Function and homology information


iron ion sequestering activity / ferritin complex / Scavenging by Class A Receptors / negative regulation of ferroptosis / Golgi Associated Vesicle Biogenesis / ferroxidase / autolysosome / ferroxidase activity / intracellular sequestering of iron ion / negative regulation of fibroblast proliferation ...iron ion sequestering activity / ferritin complex / Scavenging by Class A Receptors / negative regulation of ferroptosis / Golgi Associated Vesicle Biogenesis / ferroxidase / autolysosome / ferroxidase activity / intracellular sequestering of iron ion / negative regulation of fibroblast proliferation / autophagosome / ferric iron binding / Iron uptake and transport / ferrous iron binding / tertiary granule lumen / iron ion transport / intracellular iron ion homeostasis / ficolin-1-rich granule lumen / immune response / iron ion binding / negative regulation of cell population proliferation / Neutrophil degranulation / extracellular exosome / extracellular region / identical protein binding / membrane / nucleus / cytosol / cytoplasm
Similarity search - Function
Ferritin iron-binding regions signature 1. / Ferritin iron-binding regions signature 2. / Ferritin, conserved site / Ferritin / Ferritin-like diiron domain / Ferritin-like diiron domain profile. / Ferritin/DPS protein domain / Ferritin-like domain / Ferritin-like / Ferritin-like superfamily
Similarity search - Domain/homology
Ferritin heavy chain
Similarity search - Component
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 1.58 Å
AuthorsClare DK
Funding support United Kingdom, 1 items
OrganizationGrant numberCountry
Not fundedcm26464-2 United Kingdom
CitationJournal: Faraday Discuss / Year: 2022
Title: Application of super-resolution and correlative double sampling in cryo-electron microscopy.
Authors: Yuewen Sheng / Peter J Harrison / Vinod Vogirala / Zhengyi Yang / Claire Strain-Damerell / Thomas Frosio / Benjamin A Himes / C Alistair Siebert / Peijun Zhang / Daniel K Clare /
Abstract: Developments in cryo-EM have allowed atomic or near-atomic resolution structure determination to become routine in single particle analysis (SPA). However, near-atomic resolution structures ...Developments in cryo-EM have allowed atomic or near-atomic resolution structure determination to become routine in single particle analysis (SPA). However, near-atomic resolution structures determined using cryo-electron tomography and sub-tomogram averaging (cryo-ET STA) are much less routine. In this paper, we show that collecting cryo-ET STA data using the same conditions as SPA, with both correlated double sampling (CDS) and the super-resolution mode, allowed apoferritin to be reconstructed out to the physical Nyquist frequency of the images. Even with just two tilt series, STA yields an apoferritin map at 2.9 Å resolution. These results highlight the exciting potential of cryo-ET STA in the future of protein structure determination. While processing SPA data recorded in super-resolution mode may yield structures surpassing the physical Nyquist limit, processing cryo-ET STA data in the super-resolution mode gave no additional resolution benefit. We further show that collecting SPA data in the super-resolution mode, with CDS activated, reduces the estimated -factor, leading to a reduction in the number of particles required to reach a target resolution without compromising the data size on disk and the area imaged in SerialEM. However, collecting SPA data in CDS does reduce throughput, given that a similar resolution structure, with a slightly larger -factor, is achievable with optimised parameters for speed in EPU (without CDS).
History
DepositionFeb 11, 2022-
Header (metadata) releaseJun 22, 2022-
Map releaseJun 22, 2022-
UpdateJul 5, 2023-
Current statusJul 5, 2023Processing site: PDBe / Status: Released

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Structure visualization

Supplemental images

emd_14332.png

Downloads & links

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Map

FileDownload / File: emd_14332.map.gz / Format: CCP4 / Size: 824 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationApoF structure post processed in Relion
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.66 Å/pix.
x 600 pix.
= 393.6 Å		0.66 Å/pix.
x 600 pix.
= 393.6 Å		0.66 Å/pix.
x 600 pix.
= 393.6 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.656 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.025
Minimum - Maximum-0.05789371 - 0.19448964
Average (Standard dev.)0.00006207643 (±0.002771542)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions600600600
Spacing600600600
CellA=B=C: 393.6 Å
α=β=γ: 90.0 °

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Supplemental data

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Half map: Half map

Fileemd_14332_half_map_1.map
AnnotationHalf map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Half map

Fileemd_14332_half_map_2.map
AnnotationHalf map
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 Apoferritin

EntireName: human Apoferritin
Components
  • Complex: human Apoferritin
    • Protein or peptide: Ferritin heavy chain, N-terminally processed
  • Ligand: SODIUM ION
  • Ligand: water

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Supramolecule #1: human Apoferritin

SupramoleculeName: human Apoferritin / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1 / Details: Heavy chain
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 500 KDa

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Macromolecule #1: Ferritin heavy chain, N-terminally processed

MacromoleculeName: Ferritin heavy chain, N-terminally processed / type: protein_or_peptide / ID: 1 / Number of copies: 24 / Enantiomer: LEVO
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 20.21857 KDa
Recombinant expressionOrganism: Escherichia coli (E. coli)
SequenceString:
STSQVRQNYH QDSEAAINRQ INLELYASYV YLSMSYYFDR DDVALKNFAK YFLHQSHEER EHAEKLMKLQ NQRGGRIFLQ DIQKPD(CSX)DD WESGLNAMEC ALHLEKNVNQ SLLELHKLAT DKNDPHLCDF IETHYLNEQV KAIKELGDHV TNLRKMG AP ESGLAEYLFD KHTLG

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Macromolecule #2: SODIUM ION

MacromoleculeName: SODIUM ION / type: ligand / ID: 2 / Number of copies: 24
Molecular weightTheoretical: 22.99 Da

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Macromolecule #3: water

MacromoleculeName: water / type: ligand / ID: 3 / Number of copies: 3384 / Formula: HOH
Molecular weightTheoretical: 18.015 Da
Chemical component information

ChemComp-HOH:
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

Concentration0.01 mg/mL
BufferpH: 7.5 / Details: 50 mM Tris-HCl, pH 7.5, 100 mM NaCl, 1 mM TCEP
GridModel: Quantifoil R2/2 / Material: COPPER / Mesh: 300 / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 40 sec.
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV / Details: blot time of 2.5 seconds.
DetailsThe apoferritin sample was applied to in-house graphene-coated Quantifoil r2/2 EM grid

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsEnergy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 2295 / Average exposure time: 2.5 sec. / Average electron dose: 34.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 50.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 130000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Startup modelType of model: OTHER
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: O (octahedral) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 1.58 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3.1) / Number images used: 250000
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

6z6u

RefinementSpace: RECIPROCAL / Protocol: FLEXIBLE FIT / Target criteria: Correlation coefficient
Output model

PDB-7r5o:
1.58 A STRUCTURE OF HUMAN APOFERRITIN OBTAINED FROM TITAN KRIOS 2 AT eBIC, DLS UNDER COMMISSIONING SESSION CM26464-2

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