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- PDB-8g2u: Time-resolved cryo-EM study of the 70S recycling by the HflX:cont... -
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Open data
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Basic information
Entry | Database: PDB / ID: 8g2u | ||||||
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Title | Time-resolved cryo-EM study of the 70S recycling by the HflX:control-apo-70S at 900ms | ||||||
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![]() | RIBOSOME / Recycling / Time-resolved Cryo-EM / 70S / HflX | ||||||
Function / homology | ![]() transcription elongation-coupled chromatin remodeling / ribosome assembly / assembly of large subunit precursor of preribosome / regulation of DNA-templated transcription elongation / cytosolic ribosome assembly / transcription antitermination / DNA-templated transcription termination / ribosomal large subunit assembly / mRNA 5'-UTR binding / large ribosomal subunit ...transcription elongation-coupled chromatin remodeling / ribosome assembly / assembly of large subunit precursor of preribosome / regulation of DNA-templated transcription elongation / cytosolic ribosome assembly / transcription antitermination / DNA-templated transcription termination / ribosomal large subunit assembly / mRNA 5'-UTR binding / large ribosomal subunit / ribosomal small subunit assembly / small ribosomal subunit / small ribosomal subunit rRNA binding / transferase activity / 5S rRNA binding / large ribosomal subunit rRNA binding / cytosolic small ribosomal subunit / cytosolic large ribosomal subunit / tRNA binding / cytoplasmic translation / rRNA binding / negative regulation of translation / ribosome / structural constituent of ribosome / ribonucleoprotein complex / translation / response to antibiotic / mRNA binding / membrane / cytoplasm / cytosol Similarity search - Function | ||||||
Biological species | ![]() ![]() | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3 Å | ||||||
![]() | Bhattacharjee, S. / Brown, P.Z. / Frank, J. | ||||||
Funding support | ![]()
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![]() | ![]() Title: Time resolution in cryo-EM using a PDMS-based microfluidic chip assembly and its application to the study of HflX-mediated ribosome recycling. Authors: Sayan Bhattacharjee / Xiangsong Feng / Suvrajit Maji / Prikshat Dadhwal / Zhening Zhang / Zuben P Brown / Joachim Frank / ![]() Abstract: The rapid kinetics of biological processes and associated short-lived conformational changes pose a significant challenge in attempts to structurally visualize biomolecules during a reaction in real ...The rapid kinetics of biological processes and associated short-lived conformational changes pose a significant challenge in attempts to structurally visualize biomolecules during a reaction in real time. Conventionally, on-pathway intermediates have been trapped using chemical modifications or reduced temperature, giving limited insights. Here, we introduce a time-resolved cryo-EM method using a reusable PDMS-based microfluidic chip assembly with high reactant mixing efficiency. Coating of PDMS walls with SiO virtually eliminates non-specific sample adsorption and ensures maintenance of the stoichiometry of the reaction, rendering it highly reproducible. In an operating range from 10 to 1,000 ms, the device allows us to follow in vitro reactions of biological molecules at resolution levels in the range of 3 Å. By employing this method, we show the mechanism of progressive HflX-mediated splitting of the 70S E. coli ribosome in the presence of the GTP via capture of three high-resolution reaction intermediates within 140 ms. | ||||||
History |
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Structure visualization
Structure viewer | Molecule: ![]() ![]() |
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Downloads & links
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Download
PDBx/mmCIF format | ![]() | 3 MB | Display | ![]() |
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PDB format | ![]() | Display | ![]() | |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
Others | ![]() |
-Validation report
Summary document | ![]() | 1.5 MB | Display | ![]() |
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Full document | ![]() | 1.6 MB | Display | |
Data in XML | ![]() | 204.4 KB | Display | |
Data in CIF | ![]() | 353.7 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 29681MC ![]() 8g31C ![]() 8g34C ![]() 8g38C M: map data used to model this data C: citing same article ( |
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Similar structure data | Similarity search - Function & homology ![]() |
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Links
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Assembly
Deposited unit | ![]()
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Components
+50S ribosomal protein ... , 26 types, 26 molecules 01234CDEFGJKLMNOPQSTUVWXYZ
-RNA chain , 3 types, 3 molecules ABv
#6: RNA chain | Mass: 37848.555 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
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#7: RNA chain | Mass: 941306.188 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#51: RNA chain | Mass: 498725.406 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
-Protein , 2 types, 2 molecules Ru
#21: Protein | Mass: 11586.374 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
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#50: Protein | Mass: 6639.455 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
-30S ribosomal protein ... , 20 types, 20 molecules abcdefghijklmnopqrst
#30: Protein | Mass: 24253.943 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
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#31: Protein | Mass: 23078.785 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#32: Protein | Mass: 23383.002 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#33: Protein | Mass: 15804.282 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#34: Protein | Mass: 11669.371 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#35: Protein | Mass: 16861.523 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#36: Protein | Mass: 14015.361 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#37: Protein | Mass: 14554.882 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#38: Protein | Mass: 11196.988 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#39: Protein | Mass: 12487.200 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#40: Protein | Mass: 13636.961 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#41: Protein | Mass: 12625.753 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#42: Protein | Mass: 11489.390 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#43: Protein | Mass: 10159.621 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#44: Protein | Mass: 9207.572 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#45: Protein | Mass: 9263.946 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#46: Protein | Mass: 6466.477 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#47: Protein | Mass: 9057.626 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#48: Protein | Mass: 9506.190 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#49: Protein | Mass: 6067.081 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: CELL / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component | Name: Apo 70S at 900 ms / Type: RIBOSOME / Details: control experiment / Entity ID: all / Source: NATURAL |
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Source (natural) | Organism: ![]() ![]() |
Buffer solution | pH: 7.5 |
Specimen | Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Vitrification | Cryogen name: ETHANE |
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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: ![]() |
Electron lens | Mode: DARK FIELD / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm |
Image recording | Electron dose: 58 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) |
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Processing
EM software |
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Image processing | Details: High-pass filtered | ||||||||||||
CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||
3D reconstruction | Resolution: 3 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 140338 / Symmetry type: POINT |