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Yorodumi- EMDB-29688: Time-resolved cryo-EM study of the 70S recycling by the HflX:1st ... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-29688 | |||||||||
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Title | Time-resolved cryo-EM study of the 70S recycling by the HflX:1st intermediate | |||||||||
Map data | i70SHflX-I (First intermediate) | |||||||||
Sample |
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Keywords | Recycling / Time-resolved Cryo-EM / 70S / HflX / RIBOSOME | |||||||||
Function / homology | Function and homology information transcription elongation-coupled chromatin remodeling / ribosome assembly / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / 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 / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / 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 | Escherichia coli (E. coli) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.2 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_29688.map.gz | 130.2 MB | EMDB map data format | |
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Header (meta data) | emd-29688-v30.xml emd-29688.xml | 62.7 KB 62.7 KB | Display Display | EMDB header |
Images | emd_29688.png | 58.9 KB | ||
Filedesc metadata | emd-29688.cif.gz | 12.8 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-29688 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-29688 | HTTPS FTP |
-Validation report
Summary document | emd_29688_validation.pdf.gz | 504.3 KB | Display | EMDB validaton report |
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Full document | emd_29688_full_validation.pdf.gz | 503.9 KB | Display | |
Data in XML | emd_29688_validation.xml.gz | 6.8 KB | Display | |
Data in CIF | emd_29688_validation.cif.gz | 7.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-29688 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-29688 | HTTPS FTP |
-Related structure data
Related structure data | 8g34MC 8g2uC 8g31C 8g38C C: citing same article (ref.) M: atomic model generated by this map |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_29688.map.gz / Format: CCP4 / Size: 149.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | i70SHflX-I (First intermediate) | ||||||||||||||||||||
Voxel size | X=Y=Z: 1.03 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Sample components
+Entire : Apo 70S 1st intermediate
+Supramolecule #1: Apo 70S 1st intermediate
+Macromolecule #1: 50S ribosomal protein L32
+Macromolecule #2: 50S ribosomal protein L33
+Macromolecule #3: 50S ribosomal protein L34
+Macromolecule #4: 50S ribosomal protein L35
+Macromolecule #5: 50S ribosomal protein L36
+Macromolecule #8: 50S ribosomal protein L2
+Macromolecule #9: 50S ribosomal protein L3
+Macromolecule #10: 50S ribosomal protein L4
+Macromolecule #11: 50S ribosomal protein L5
+Macromolecule #12: 50S ribosomal protein L6
+Macromolecule #13: 50S ribosomal protein L13
+Macromolecule #14: 50S ribosomal protein L14
+Macromolecule #15: 50S ribosomal protein L15
+Macromolecule #16: 50S ribosomal protein L16
+Macromolecule #17: 50S ribosomal protein L17
+Macromolecule #18: 50S ribosomal protein L18
+Macromolecule #19: 50S ribosomal protein L19
+Macromolecule #20: 50S ribosomal protein L20
+Macromolecule #21: Ribosomal protein L21
+Macromolecule #22: 50S ribosomal protein L22
+Macromolecule #23: 50S ribosomal protein L23
+Macromolecule #24: 50S ribosomal protein L24
+Macromolecule #25: 50S ribosomal protein L25
+Macromolecule #26: 50S ribosomal protein L27
+Macromolecule #27: 50S ribosomal protein L28
+Macromolecule #28: 50S ribosomal protein L29
+Macromolecule #29: 50S ribosomal protein L30
+Macromolecule #30: GTPase HflX
+Macromolecule #31: 30S ribosomal protein S3
+Macromolecule #32: 30S ribosomal protein S7
+Macromolecule #33: 30S ribosomal protein S9
+Macromolecule #34: 30S ribosomal protein S10
+Macromolecule #35: 30S ribosomal protein S13
+Macromolecule #36: 30S ribosomal protein S14
+Macromolecule #37: 30S ribosomal protein S19
+Macromolecule #38: Transcription termination/antitermination protein NusG
+Macromolecule #39: 30S ribosomal protein S2
+Macromolecule #40: 30S ribosomal protein S4
+Macromolecule #41: 30S ribosomal protein S5
+Macromolecule #42: 30S ribosomal protein S6, non-modified isoform
+Macromolecule #43: 30S ribosomal protein S8
+Macromolecule #44: 30S ribosomal protein S11
+Macromolecule #45: 30S ribosomal protein S12
+Macromolecule #46: 30S ribosomal protein S15
+Macromolecule #47: 30S ribosomal protein S16
+Macromolecule #48: 30S ribosomal protein S17
+Macromolecule #49: 30S ribosomal protein S18
+Macromolecule #50: 30S ribosomal protein S20
+Macromolecule #51: 30S ribosomal protein S21 (Fragment)
+Macromolecule #6: 5S
+Macromolecule #7: 23S
+Macromolecule #52: 16S
-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 |