+Open data
-Basic information
Entry | Database: PDB / ID: 6r7m | |||||||||
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Title | Tobacco Mosaic Virus (TMV) | |||||||||
Components | Capsid protein | |||||||||
Keywords | VIRUS / tobacco / mosaic / TMV / cryo-EM / cryoWriter / microfluidic | |||||||||
Function / homology | Function and homology information helical viral capsid / structural molecule activity / identical protein binding Similarity search - Function | |||||||||
Biological species | Tobacco mosaic virus | |||||||||
Method | ELECTRON MICROSCOPY / helical reconstruction / cryo EM / Resolution: 1.92 Å | |||||||||
Authors | Schmidli, C. / Albiez, S. / Rima, L. / Righetto, R. / Mohammed, I. / Oliva, P. / Kovacik, L. / Stahlberg, H. / Braun, T. | |||||||||
Funding support | Switzerland, 2items
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Citation | Journal: Proc Natl Acad Sci U S A / Year: 2019 Title: Microfluidic protein isolation and sample preparation for high-resolution cryo-EM. Authors: Claudio Schmidli / Stefan Albiez / Luca Rima / Ricardo Righetto / Inayatulla Mohammed / Paolo Oliva / Lubomir Kovacik / Henning Stahlberg / Thomas Braun / Abstract: High-resolution structural information is essential to understand protein function. Protein-structure determination needs a considerable amount of protein, which can be challenging to produce, often ...High-resolution structural information is essential to understand protein function. Protein-structure determination needs a considerable amount of protein, which can be challenging to produce, often involving harsh and lengthy procedures. In contrast, the several thousand to a few million protein particles required for structure determination by cryogenic electron microscopy (cryo-EM) can be provided by miniaturized systems. Here, we present a microfluidic method for the rapid isolation of a target protein and its direct preparation for cryo-EM. Less than 1 μL of cell lysate is required as starting material to solve the atomic structure of the untagged, endogenous human 20S proteasome. Our work paves the way for high-throughput structure determination of proteins from minimal amounts of cell lysate and opens more opportunities for the isolation of sensitive, endogenous protein complexes. | |||||||||
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 | 6r7m.cif.gz | 38.9 KB | Display | PDBx/mmCIF format |
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PDB format | pdb6r7m.ent.gz | 25.9 KB | Display | PDB format |
PDBx/mmJSON format | 6r7m.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 6r7m_validation.pdf.gz | 706.9 KB | Display | wwPDB validaton report |
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Full document | 6r7m_full_validation.pdf.gz | 706.4 KB | Display | |
Data in XML | 6r7m_validation.xml.gz | 20 KB | Display | |
Data in CIF | 6r7m_validation.cif.gz | 28.1 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/r7/6r7m ftp://data.pdbj.org/pub/pdb/validation_reports/r7/6r7m | HTTPS FTP |
-Related structure data
Related structure data | 4628MC 4738C 6r70C 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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2 |
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Symmetry | Helical symmetry: (Circular symmetry: 1 / Dyad axis: no / N subunits divisor: 1 / Num. of operations: 40 / Rise per n subunits: 1.41 Å / Rotation per n subunits: 22.03 °) |
-Components
#1: Protein | Mass: 17091.998 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Tobacco mosaic virus / References: UniProt: A0A348G6U1, UniProt: P69687*PLUS |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: HELICAL ARRAY / 3D reconstruction method: helical reconstruction |
-Sample preparation
Component | Name: Tobacco mosaic virus / Type: VIRUS / Entity ID: all / Source: NATURAL |
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Molecular weight | Value: 39.5 MDa / Experimental value: NO |
Source (natural) | Organism: Tobacco mosaic virus |
Details of virus | Empty: NO / Enveloped: NO / Isolate: STRAIN / Type: VIRUS-LIKE PARTICLE |
Natural host | Organism: Nicotiana tabacum |
Virus shell | Name: CP / Diameter: 180 nm |
Buffer solution | pH: 7.4 |
Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Grid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: Quantifoil R1.2/1.3 |
Vitrification | Cryogen name: ETHANE / Details: CryoWriter |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Cs: 2.7 mm / C2 aperture diameter: 70 µm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Average exposure time: 6 sec. / Electron dose: 72 e/Å2 / Detector mode: SUPER-RESOLUTION / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 523 |
Image scans | Width: 7676 / Height: 7420 / Movie frames/image: 30 / Used frames/image: 0-30 |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||||||||||
Helical symmerty | Angular rotation/subunit: 22.03 ° / Axial rise/subunit: 1.41 Å / Axial symmetry: C1 | |||||||||||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 2676 | |||||||||||||||||||||||||||||||||||
3D reconstruction | Resolution: 1.92 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 52776 / Algorithm: BACK PROJECTION / Symmetry type: HELICAL | |||||||||||||||||||||||||||||||||||
Atomic model building | PDB-ID: 6I5A Accession code: 6I5A / Source name: PDB / Type: experimental model |