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Yorodumi- EMDB-29844: Time-resolved cryo-EM study of the 70S recycling by the HflX:2nd ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-29844 | |||||||||
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Title | Time-resolved cryo-EM study of the 70S recycling by the HflX:2nd Intermediate, 50S focused and 30S subtracted | |||||||||
Map data | Time-resolved cryo-EM study of the 70S recycling by the HflX:2nd Intermediate, 50S focused and 30S subtracted | |||||||||
Sample |
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Keywords | Recycling / Time-resolved Cryo-EM / 70S / HflX / RIBOSOME | |||||||||
Biological species | Escherichia coli (E. coli) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.14 Å | |||||||||
Authors | Bhattacharjee S / Brown PZ / Frank J | |||||||||
Funding support | United States, 1 items
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Citation | Journal: Cell / Year: 2024 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
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_29844.map.gz | 141.8 MB | EMDB map data format | |
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Header (meta data) | emd-29844-v30.xml emd-29844.xml | 12.6 KB 12.6 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_29844_fsc.xml | 12.1 KB | Display | FSC data file |
Images | emd_29844.png | 91.5 KB | ||
Masks | emd_29844_msk_1.map | 149.9 MB | Mask map | |
Filedesc metadata | emd-29844.cif.gz | 4 KB | ||
Others | emd_29844_half_map_1.map.gz emd_29844_half_map_2.map.gz | 139 MB 139 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-29844 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-29844 | HTTPS FTP |
-Validation report
Summary document | emd_29844_validation.pdf.gz | 1.3 MB | Display | EMDB validaton report |
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Full document | emd_29844_full_validation.pdf.gz | 1.3 MB | Display | |
Data in XML | emd_29844_validation.xml.gz | 19.6 KB | Display | |
Data in CIF | emd_29844_validation.cif.gz | 25.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-29844 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-29844 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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-Map
File | Download / File: emd_29844.map.gz / Format: CCP4 / Size: 149.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Time-resolved cryo-EM study of the 70S recycling by the HflX:2nd Intermediate, 50S focused and 30S subtracted | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.02529 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_29844_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: Time-resolved cryo-EM study of the 70S recycling by...
File | emd_29844_half_map_1.map | ||||||||||||
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Annotation | Time-resolved cryo-EM study of the 70S recycling by the HflX:2nd Intermediate, 50S focused and 30S subtracted, half-1 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Time-resolved cryo-EM study of the 70S recycling by...
File | emd_29844_half_map_2.map | ||||||||||||
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Annotation | Time-resolved cryo-EM study of the 70S recycling by the HflX:2nd Intermediate, 50S focused and 30S subtracted, half-2 | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
-Entire : 50S 2nd intermediate
Entire | Name: 50S 2nd intermediate |
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Components |
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-Supramolecule #1: 50S 2nd intermediate
Supramolecule | Name: 50S 2nd intermediate / type: complex / ID: 1 / Parent: 0 |
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Source (natural) | Organism: Escherichia coli (E. coli) |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | cell |
-Sample preparation
Buffer | pH: 7.5 |
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Vitrification | Cryogen name: ETHANE |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K3 (6k x 4k) / Average electron dose: 58.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: DARK FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |