+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-8521 | |||||||||
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Title | 3.2 A cryo-EM ArfA-RF2 ribosome rescue complex (Structure II) | |||||||||
Map data | ||||||||||
Sample |
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Keywords | ARFA RF2 ribosome rescue complex / RIBOSOME | |||||||||
Function / homology | Function and homology information translation release factor activity, codon specific / negative regulation of cytoplasmic translational initiation / stringent response / ribosomal large subunit binding / ornithine decarboxylase inhibitor activity / transcription antitermination factor activity, RNA binding / misfolded RNA binding / Group I intron splicing / RNA folding / transcriptional attenuation ...translation release factor activity, codon specific / negative regulation of cytoplasmic translational initiation / stringent response / ribosomal large subunit binding / ornithine decarboxylase inhibitor activity / transcription antitermination factor activity, RNA binding / misfolded RNA binding / Group I intron splicing / RNA folding / transcriptional attenuation / endoribonuclease inhibitor activity / RNA-binding transcription regulator activity / positive regulation of ribosome biogenesis / negative regulation of cytoplasmic translation / four-way junction DNA binding / translational termination / DnaA-L2 complex / translation repressor activity / negative regulation of translational initiation / negative regulation of DNA-templated DNA replication initiation / regulation of mRNA stability / rescue of stalled ribosome / mRNA regulatory element binding translation repressor activity / ribosome assembly / positive regulation of RNA splicing / assembly of large subunit precursor of preribosome / transcription elongation factor complex / cytosolic ribosome assembly / regulation of DNA-templated transcription elongation / DNA endonuclease activity / ribosomal large subunit assembly / transcription antitermination / response to reactive oxygen species / translational initiation / regulation of cell growth / DNA-templated transcription termination / maintenance of translational fidelity / response to radiation / mRNA 5'-UTR binding / large ribosomal subunit / ribosome biogenesis / ribosome binding / regulation of translation / ribosomal small subunit biogenesis / 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 / cytoplasmic translation / tRNA binding / molecular adaptor activity / rRNA binding / negative regulation of translation / ribosome / structural constituent of ribosome / translation / ribonucleoprotein complex / response to antibiotic / negative regulation of DNA-templated transcription / mRNA binding / DNA binding / RNA binding / zinc ion binding / membrane / metal ion binding / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Escherichia coli (E. coli) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.2 Å | |||||||||
Authors | Demo G / Svidritskiy E | |||||||||
Funding support | United States, 2 items
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Citation | Journal: Elife / Year: 2017 Title: Mechanism of ribosome rescue by ArfA and RF2. Authors: Gabriel Demo / Egor Svidritskiy / Rohini Madireddy / Ruben Diaz-Avalos / Timothy Grant / Nikolaus Grigorieff / Duncan Sousa / Andrei A Korostelev / Abstract: ArfA rescues ribosomes stalled on truncated mRNAs by recruiting release factor RF2, which normally binds stop codons to catalyze peptide release. We report two 3.2 Å resolution cryo-EM structures - ...ArfA rescues ribosomes stalled on truncated mRNAs by recruiting release factor RF2, which normally binds stop codons to catalyze peptide release. We report two 3.2 Å resolution cryo-EM structures - determined from a single sample - of the 70S ribosome with ArfA•RF2 in the A site. In both states, the ArfA C-terminus occupies the mRNA tunnel downstream of the A site. One state contains a compact inactive RF2 conformation. Ordering of the ArfA N-terminus in the second state rearranges RF2 into an extended conformation that docks the catalytic GGQ motif into the peptidyl-transferase center. Our work thus reveals the structural dynamics of ribosome rescue. The structures demonstrate how ArfA 'senses' the vacant mRNA tunnel and activates RF2 to mediate peptide release without a stop codon, allowing stalled ribosomes to be recycled. | |||||||||
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_8521.map.gz | 36.6 MB | EMDB map data format | |
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Header (meta data) | emd-8521-v30.xml emd-8521.xml | 77.7 KB 77.7 KB | Display Display | EMDB header |
Images | emd_8521.png | 87.3 KB | ||
Filedesc metadata | emd-8521.cif.gz | 14.8 KB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-8521 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-8521 | HTTPS FTP |
-Validation report
Summary document | emd_8521_validation.pdf.gz | 533.3 KB | Display | EMDB validaton report |
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Full document | emd_8521_full_validation.pdf.gz | 532.9 KB | Display | |
Data in XML | emd_8521_validation.xml.gz | 7.1 KB | Display | |
Data in CIF | emd_8521_validation.cif.gz | 8.1 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8521 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-8521 | HTTPS FTP |
-Related structure data
Related structure data | 5u9fMC 8522C 5u9gC C: citing same article (ref.) M: atomic model generated by this map |
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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_8521.map.gz / Format: CCP4 / Size: 125 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Voxel size | X=Y=Z: 1.215 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 : ArfA-RF2 ribosome rescue complex
+Supramolecule #1: ArfA-RF2 ribosome rescue complex
+Macromolecule #1: 16S ribosomal RNA
+Macromolecule #2: 23S ribosomal RNA
+Macromolecule #3: 5S ribosomal RNA
+Macromolecule #5: fMet-tRNA (P- and E-site)
+Macromolecule #38: truncated mRNA
+Macromolecule #4: Alternative ribosome-rescue factor A
+Macromolecule #6: 50S ribosomal protein L1
+Macromolecule #7: 50S ribosomal protein L2
+Macromolecule #8: 50S ribosomal protein L3
+Macromolecule #9: 50S ribosomal protein L4
+Macromolecule #10: 50S ribosomal protein L5
+Macromolecule #11: 50S ribosomal protein L6
+Macromolecule #12: 50S ribosomal protein L9
+Macromolecule #13: 50S ribosomal protein L10
+Macromolecule #14: 50S ribosomal protein L11
+Macromolecule #15: 50S ribosomal protein L13
+Macromolecule #16: 50S ribosomal protein L14
+Macromolecule #17: 50S ribosomal protein L15
+Macromolecule #18: 50S ribosomal protein L16
+Macromolecule #19: 50S ribosomal protein L17
+Macromolecule #20: 50S ribosomal protein L18
+Macromolecule #21: 50S ribosomal protein L19
+Macromolecule #22: 50S ribosomal protein L20
+Macromolecule #23: 50S ribosomal protein L21
+Macromolecule #24: 50S ribosomal protein L22
+Macromolecule #25: 50S ribosomal protein L23
+Macromolecule #26: 50S ribosomal protein L24
+Macromolecule #27: 50S ribosomal protein L25
+Macromolecule #28: 50S ribosomal protein L27
+Macromolecule #29: 50S ribosomal protein L28
+Macromolecule #30: 50S ribosomal protein L29
+Macromolecule #31: 50S ribosomal protein L30
+Macromolecule #32: 50S ribosomal protein L31
+Macromolecule #33: 50S ribosomal protein L32
+Macromolecule #34: 50S ribosomal protein L33
+Macromolecule #35: 50S ribosomal protein L34
+Macromolecule #36: 50S ribosomal protein L35
+Macromolecule #37: 50S ribosomal protein L36
+Macromolecule #39: Peptide chain release factor RF2
+Macromolecule #40: 30S ribosomal protein S2
+Macromolecule #41: 30S ribosomal protein S3
+Macromolecule #42: 30S ribosomal protein S4
+Macromolecule #43: 30S ribosomal protein S5
+Macromolecule #44: 30S ribosomal protein S6
+Macromolecule #45: 30S ribosomal protein S7
+Macromolecule #46: 30S ribosomal protein S8
+Macromolecule #47: 30S ribosomal protein S9
+Macromolecule #48: 30S ribosomal protein S10
+Macromolecule #49: 30S ribosomal protein S11
+Macromolecule #50: 30S ribosomal protein S12
+Macromolecule #51: 30S ribosomal protein S13
+Macromolecule #52: 30S ribosomal protein S14
+Macromolecule #53: 30S ribosomal protein S15
+Macromolecule #54: 30S ribosomal protein S16
+Macromolecule #55: 30S ribosomal protein S17
+Macromolecule #56: 30S ribosomal protein S18
+Macromolecule #57: 30S ribosomal protein S19
+Macromolecule #58: 30S ribosomal protein S20
+Macromolecule #59: 30S ribosomal protein S21
+Macromolecule #60: MAGNESIUM ION
+Macromolecule #61: ZINC ION
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7 Component:
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Grid | Model: C-flat-2/2 / Material: COPPER / Mesh: 200 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: PLASMA CLEANING / Pretreatment - Time: 20 sec. / Pretreatment - Atmosphere: OTHER Details: Using a Solarus 950 plasma cleaning system. The forward RF target was set to 7w. | ||||||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 277.15 K / Instrument: FEI VITROBOT MARK IV |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: DIRECT ELECTRON DE-20 (5k x 3k) / Detector mode: COUNTING / Digitization - Frames/image: 2-27 / Number grids imaged: 1 / Number real images: 3760 / Average electron dose: 1.2 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 3.0 µm / Nominal defocus min: 0.5 µm / Nominal magnification: 29000 |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
+Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: OTHER / Overall B value: 100 / Target criteria: Correlation coefficient |
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Output model | PDB-5u9f: |