+Open data
-Basic information
Entry | Database: PDB / ID: 7o6e | |||||||||
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Title | 2.12 A cryo-EM structure of Mycobacterium tuberculosis Ferritin | |||||||||
Components | Ferritin BfrB | |||||||||
Keywords | METAL TRANSPORT / Iron storage / Ferroxidase / Bacterial Ferritin / Octahedral symmetry. | |||||||||
Function / homology | Function and homology information Mtb iron assimilation by chelation / response to nitrosative stress / encapsulin nanocompartment / iron ion sequestering activity / ferroxidase / intracellular sequestering of iron ion / ferroxidase activity / peptidoglycan-based cell wall / ferric iron binding / ferrous iron binding ...Mtb iron assimilation by chelation / response to nitrosative stress / encapsulin nanocompartment / iron ion sequestering activity / ferroxidase / intracellular sequestering of iron ion / ferroxidase activity / peptidoglycan-based cell wall / ferric iron binding / ferrous iron binding / iron ion transport / response to hypoxia / extracellular region / plasma membrane / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Mycobacterium tuberculosis H37Rv (bacteria) | |||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.1 Å | |||||||||
Authors | Gijsbers, A. / Zhang, Y. / Gao, Y. / Peters, P.J. / Ravelli, R.B.G. | |||||||||
Funding support | European Union, Netherlands, 2items
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Citation | Journal: Acta Crystallogr D Struct Biol / Year: 2021 Title: Mycobacterium tuberculosis ferritin: a suitable workhorse protein for cryo-EM development. Authors: Abril Gijsbers / Yue Zhang / Ye Gao / Peter J Peters / Raimond B G Ravelli / Abstract: The use of cryo-EM continues to expand worldwide and calls for good-quality standard proteins with simple protocols for their production. Here, a straightforward expression and purification protocol ...The use of cryo-EM continues to expand worldwide and calls for good-quality standard proteins with simple protocols for their production. Here, a straightforward expression and purification protocol is presented that provides an apoferritin, bacterioferritin B (BfrB), from Mycobacterium tuberculosis with high yield and purity. A 2.12 Å resolution cryo-EM structure of BfrB is reported, showing the typical cage-like oligomer constituting of 24 monomers related by 432 symmetry. However, it also contains a unique C-terminal extension (164-181), which loops into the cage region of the shell and provides extra stability to the protein. Part of this region was ambiguous in previous crystal structures but could be built within the cryo-EM map. These findings and this protocol could serve the growing cryo-EM community in characterizing and pushing the limits of their electron microscopes and workflows. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7o6e.cif.gz | 734.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7o6e.ent.gz | 613.1 KB | Display | PDB format |
PDBx/mmJSON format | 7o6e.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7o6e_validation.pdf.gz | 1.3 MB | Display | wwPDB validaton report |
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Full document | 7o6e_full_validation.pdf.gz | 1.4 MB | Display | |
Data in XML | 7o6e_validation.xml.gz | 106.8 KB | Display | |
Data in CIF | 7o6e_validation.cif.gz | 136.3 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/o6/7o6e ftp://data.pdbj.org/pub/pdb/validation_reports/o6/7o6e | HTTPS FTP |
-Related structure data
Related structure data | 12738MC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
#1: Protein | Mass: 20463.936 Da / Num. of mol.: 24 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Mycobacterium tuberculosis H37Rv (bacteria) Gene: bfrB / Plasmid: pRSET / Production host: Escherichia coli BL21(DE3) (bacteria) / Variant (production host): C41 / References: UniProt: P9WNE5, ferroxidase #2: Water | ChemComp-HOH / | |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: Mycobacterium tuberculosis ferritin / Type: ORGANELLE OR CELLULAR COMPONENT / Details: Mycobacterium tuberculosis ferritin / Entity ID: #1 / Source: RECOMBINANT | |||||||||||||||
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Molecular weight | Value: 0.490 MDa / Experimental value: NO | |||||||||||||||
Source (natural) | Organism: Mycobacterium tuberculosis H37Rv (bacteria) / Strain: H37Rv | |||||||||||||||
Source (recombinant) | Organism: Escherichia coli BL21(DE3) (bacteria) / Strain: C41 / Plasmid: pRSET | |||||||||||||||
Buffer solution | pH: 8 | |||||||||||||||
Buffer component |
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Specimen | Conc.: 11 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES / Details: This sample was monodisperse. | |||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil | |||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS Details: Basic direct alignments were done as well as astigmatism and coma alignment using AutoCTF |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Calibrated magnification: 76757 X / Nominal defocus max: 1400 nm / Nominal defocus min: 400 nm / Calibrated defocus min: 200 nm / Calibrated defocus max: 2000 nm / Cs: 2.7 mm / C2 aperture diameter: 50 µm / Alignment procedure: OTHER |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature (max): 105 K / Temperature (min): 93 K |
Image recording | Average exposure time: 1.3 sec. / Electron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2518 Details: Images were acquired in superresolution counting mode at a speed of 64 movies/hour and stored as tiff lzw non-gain normalised |
EM imaging optics | Energyfilter name: GIF Bioquantum / Energyfilter slit width: 20 eV |
Image scans | Sampling size: 5 µm / Width: 5760 / Height: 4092 |
-Processing
Software |
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 249563 | ||||||||||||||||||||||||||||||||||||
Symmetry | Point symmetry: O (octahedral) | ||||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 2.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 163568 / Algorithm: FOURIER SPACE / Symmetry type: POINT | ||||||||||||||||||||||||||||||||||||
Atomic model building | B value: 28.1 / Protocol: OTHER / Space: REAL / Target criteria: Correlation coefficient | ||||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 3QD8 Pdb chain-ID: A / Accession code: 3QD8 / Pdb chain residue range: 10-181 / Source name: PDB / Type: experimental model | ||||||||||||||||||||||||||||||||||||
Refinement | Cross valid method: NONE Stereochemistry target values: GeoStd + Monomer Library + CDL v1.2 | ||||||||||||||||||||||||||||||||||||
Displacement parameters | Biso mean: 28.43 Å2 | ||||||||||||||||||||||||||||||||||||
Refine LS restraints |
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