+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-3133 | |||||||||
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Title | Cryo-EM structures of the 50S ribosome subunit bound with HflX | |||||||||
Map data | Cryo-EM structures of the 50S ribosome subunit bound with HflX | |||||||||
Sample |
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Keywords | Ribosome rescue | |||||||||
Function / homology | Function and homology information ribosome disassembly / guanosine tetraphosphate binding / stringent response / ribosomal large subunit binding / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity / positive regulation of ribosome biogenesis / negative regulation of cytoplasmic translation / rescue of stalled ribosome ...ribosome disassembly / guanosine tetraphosphate binding / stringent response / ribosomal large subunit binding / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity / positive regulation of ribosome biogenesis / negative regulation of cytoplasmic translation / rescue of stalled ribosome / translational termination / DnaA-L2 complex / negative regulation of translational initiation / translation repressor activity / negative regulation of DNA-templated DNA replication initiation / ribosome assembly / mRNA regulatory element binding translation repressor activity / response to reactive oxygen species / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly / regulation of cell growth / DNA-templated transcription termination / response to radiation / mRNA 5'-UTR binding / ribosomal large subunit assembly / large ribosomal subunit rRNA binding / ribosome binding / large ribosomal subunit / cytoplasmic translation / 5S rRNA binding / cytosolic large ribosomal subunit / response to heat / transferase activity / tRNA binding / negative regulation of translation / rRNA binding / ribosome / structural constituent of ribosome / translation / response to antibiotic / mRNA binding / GTPase activity / negative regulation of DNA-templated transcription / GTP binding / ATP hydrolysis activity / DNA binding / RNA binding / zinc ion binding / ATP binding / metal ion binding / cytosol / cytoplasm Similarity search - Function | |||||||||
Biological species | Escherichia coli K-12 (bacteria) / Escherichia coli (E. coli) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 4.5 Å | |||||||||
Authors | Zhang Y / Mandava CS / Cao W / Li X / Zhang D / Li N / Zhang X / Qin Y / Mi K / Lei J ...Zhang Y / Mandava CS / Cao W / Li X / Zhang D / Li N / Zhang X / Qin Y / Mi K / Lei J / Sanyal S / Gao N | |||||||||
Citation | Journal: Nat Struct Mol Biol / Year: 2015 Title: HflX is a ribosome-splitting factor rescuing stalled ribosomes under stress conditions. Authors: Yanqing Zhang / Chandra Sekhar Mandava / Wei Cao / Xiaojing Li / Dejiu Zhang / Ningning Li / Yixiao Zhang / Xiaoxiao Zhang / Yan Qin / Kaixia Mi / Jianlin Lei / Suparna Sanyal / Ning Gao / Abstract: Adverse cellular conditions often lead to nonproductive translational stalling and arrest of ribosomes on mRNAs. Here, we used fast kinetics and cryo-EM to characterize Escherichia coli HflX, a ...Adverse cellular conditions often lead to nonproductive translational stalling and arrest of ribosomes on mRNAs. Here, we used fast kinetics and cryo-EM to characterize Escherichia coli HflX, a GTPase with unknown function. Our data reveal that HflX is a heat shock-induced ribosome-splitting factor capable of dissociating vacant as well as mRNA-associated ribosomes with deacylated tRNA in the peptidyl site. Structural data demonstrate that the N-terminal effector domain of HflX binds to the peptidyl transferase center in a strikingly similar manner as that of the class I release factors and induces dramatic conformational changes in central intersubunit bridges, thus promoting subunit dissociation. Accordingly, loss of HflX results in an increase in stalled ribosomes upon heat shock. These results suggest a primary role of HflX in rescuing translationally arrested ribosomes under stress conditions. | |||||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | EM map: SurfViewMolmilJmol/JSmol |
Supplemental images |
-Downloads & links
-EMDB archive
Map data | emd_3133.map.gz | 95.8 MB | EMDB map data format | |
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Header (meta data) | emd-3133-v30.xml emd-3133.xml | 11.5 KB 11.5 KB | Display Display | EMDB header |
Images | emd_3133.png | 160.7 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-3133 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-3133 | HTTPS FTP |
-Related structure data
Related structure data | 5adyMC M: atomic model generated by this map C: citing same article (ref.) |
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Similar structure data |
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
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Related items in Molecule of the Month |
-Map
File | Download / File: emd_3133.map.gz / Format: CCP4 / Size: 100.6 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | Cryo-EM structures of the 50S ribosome subunit bound with HflX | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.1659 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
-Sample components
-Entire : 50S-HflX complex
Entire | Name: 50S-HflX complex |
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Components |
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-Supramolecule #1000: 50S-HflX complex
Supramolecule | Name: 50S-HflX complex / type: sample / ID: 1000 / Number unique components: 2 |
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Molecular weight | Experimental: 1.5 MDa / Theoretical: 1.5 MDa |
-Supramolecule #1: prokaryotic 50S ribosome subunit
Supramolecule | Name: prokaryotic 50S ribosome subunit / type: complex / ID: 1 / Name.synonym: 50S subunit / Recombinant expression: No / Ribosome-details: ribosome-prokaryote: LSU 50S |
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Source (natural) | Organism: Escherichia coli K-12 (bacteria) / Location in cell: Cytoplasm |
Molecular weight | Experimental: 1.5 MDa / Theoretical: 1.5 MDa |
-Macromolecule #1: HflX
Macromolecule | Name: HflX / type: protein_or_peptide / ID: 1 Details: GMPPNP was bound with HflX in the 50S-HflX complex. Number of copies: 1 / Oligomeric state: monomer / Recombinant expression: Yes |
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Source (natural) | Organism: Escherichia coli (E. coli) / Location in cell: Cytoplasm |
Molecular weight | Experimental: 50 KDa / Theoretical: 50 KDa |
Recombinant expression | Organism: Escherichia coli BL21 (bacteria) / Recombinant plasmid: pET28a |
Sequence | UniProtKB: GTPase HflX |
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.5 / Details: 20mM Tris-HCl, 100mM NH4Cl, 10mM MgCl2 |
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Grid | Details: 200 mesh copper grid with thin carbon support, glow discharged |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK IV / Method: Blot for 1 seconds before plunging |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.70 mm / Nominal defocus max: 4.0 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 75000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Date | Jan 7, 2012 |
Image recording | Category: CCD / Film or detector model: FEI EAGLE (4k x 4k) / Number real images: 6022 / Average electron dose: 20 e/Å2 |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
-Image processing
CTF correction | Details: Each particle |
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Final reconstruction | Applied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 4.5 Å / Resolution method: OTHER / Software - Name: Spider, Relion / Number images used: 384206 |
-Atomic model buiding 1
Initial model | PDB ID: 3fik |
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Software | Name: MDFF |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
Output model | PDB-5ady: |
-Atomic model buiding 2
Initial model | PDB ID: |
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Software | Name: MDFF |
Details | Atomic model of E. coli HflX was modeled from the crystal structure of Sulfolobus solfataricus HflX (PDB id: 3KXI).The homology modeling was performed with MODELLER. The model of the CTD of E. coli HflX was independently modeled by I-TASSER (template PDB code: 2WBM, residues 164-232). The switch I region disordered in the crystal structure of S. solfataricus HflX was modeled using the crystal structure of S. thermophilus NFeoB (PDB id: 3B1X) as a template. GMPPNP was derived from a previous model (PDB id: 3B1X) and docked into the atomic model of the E. coli HflX. |
Refinement | Space: REAL / Protocol: FLEXIBLE FIT |
Output model | PDB-5ady: |