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Yorodumi- EMDB-28197: Escherichia coli 70S ribosome bound to thermorubin, deacylated P-... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-28197 | |||||||||
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Title | Escherichia coli 70S ribosome bound to thermorubin, deacylated P-site tRNAfMet and aminoacylated A-site Phe-tRNA | |||||||||
Map data | Escherichia coli 70S ribosome bound to thermorubin, deacylated P-site tRNAfMet and aminoacylated A-site Phe-tRNA. | |||||||||
Sample |
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Keywords | Antibiotic / Thermorubin / cryo-EM / Ribosome | |||||||||
Function / homology | Function and homology information negative regulation of cytoplasmic translational initiation / positive regulation of ribosome biogenesis / DnaA-L2 complex / negative regulation of translational initiation / negative regulation of DNA-templated DNA replication initiation / translational initiation / ribosome assembly / mRNA regulatory element binding translation repressor activity / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly ...negative regulation of cytoplasmic translational initiation / positive regulation of ribosome biogenesis / DnaA-L2 complex / negative regulation of translational initiation / negative regulation of DNA-templated DNA replication initiation / translational initiation / ribosome assembly / mRNA regulatory element binding translation repressor activity / assembly of large subunit precursor of preribosome / cytosolic ribosome assembly / transcription antitermination / regulation of cell growth / DNA-templated transcription termination / maintenance of translational fidelity / ribosomal large subunit assembly / mRNA 5'-UTR binding / large ribosomal subunit / ribosome binding / 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 / tRNA binding / cytoplasmic translation / rRNA binding / negative regulation of translation / ribosome / structural constituent of ribosome / ribonucleoprotein complex / translation / response to antibiotic / mRNA binding / RNA binding / zinc ion binding / membrane / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | Escherichia coli (E. coli) / Escherichia phage T4 (virus) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.7 Å | |||||||||
Authors | Rybak MY / Gagnon MG | |||||||||
Funding support | United States, 2 items
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Citation | Journal: Nucleic Acids Res / Year: 2023 Title: Insights into the molecular mechanism of translation inhibition by the ribosome-targeting antibiotic thermorubin. Authors: Madhura N Paranjpe / Valeria I Marina / Aleksandr A Grachev / Tinashe P Maviza / Olga A Tolicheva / Alena Paleskava / Ilya A Osterman / Petr V Sergiev / Andrey L Konevega / Yury S Polikanov ...Authors: Madhura N Paranjpe / Valeria I Marina / Aleksandr A Grachev / Tinashe P Maviza / Olga A Tolicheva / Alena Paleskava / Ilya A Osterman / Petr V Sergiev / Andrey L Konevega / Yury S Polikanov / Matthieu G Gagnon / Abstract: Thermorubin (THR) is an aromatic anthracenopyranone antibiotic active against both Gram-positive and Gram-negative bacteria. It is known to bind to the 70S ribosome at the intersubunit bridge B2a and ...Thermorubin (THR) is an aromatic anthracenopyranone antibiotic active against both Gram-positive and Gram-negative bacteria. It is known to bind to the 70S ribosome at the intersubunit bridge B2a and was thought to inhibit factor-dependent initiation of translation and obstruct the accommodation of tRNAs into the A site. Here, we show that thermorubin causes ribosomes to stall in vivo and in vitro at internal and termination codons, thereby allowing the ribosome to initiate protein synthesis and translate at least a few codons before stalling. Our biochemical data show that THR affects multiple steps of translation elongation with a significant impact on the binding stability of the tRNA in the A site, explaining premature cessation of translation. Our high-resolution crystal and cryo-EM structures of the 70S-THR complex show that THR can co-exist with P- and A-site tRNAs, explaining how ribosomes can elongate in the presence of the drug. Remarkable is the ability of THR to arrest ribosomes at the stop codons. Our data suggest that by causing structural re-arrangements in the decoding center, THR interferes with the accommodation of tRNAs or release factors into the ribosomal A site. | |||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_28197.map.gz | 255.8 MB | EMDB map data format | |
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Header (meta data) | emd-28197-v30.xml emd-28197.xml | 79.6 KB 79.6 KB | Display Display | EMDB header |
Images | emd_28197.png | 210.3 KB | ||
Masks | emd_28197_msk_1.map | 512 MB | Mask map | |
Filedesc metadata | emd-28197.cif.gz | 15.4 KB | ||
Others | emd_28197_additional_1.map.gz emd_28197_half_map_1.map.gz emd_28197_half_map_2.map.gz | 264.2 MB 475.1 MB 475.1 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-28197 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-28197 | HTTPS FTP |
-Validation report
Summary document | emd_28197_validation.pdf.gz | 1005.1 KB | Display | EMDB validaton report |
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Full document | emd_28197_full_validation.pdf.gz | 1004.7 KB | Display | |
Data in XML | emd_28197_validation.xml.gz | 19 KB | Display | |
Data in CIF | emd_28197_validation.cif.gz | 22.7 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28197 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28197 | HTTPS FTP |
-Related structure data
Related structure data | 8ekcMC 8ekbC M: atomic model generated by this map C: citing same article (ref.) |
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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_28197.map.gz / Format: CCP4 / Size: 512 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||
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Annotation | Escherichia coli 70S ribosome bound to thermorubin, deacylated P-site tRNAfMet and aminoacylated A-site Phe-tRNA. | ||||||||||||||||||||
Voxel size | X=Y=Z: 0.86 Å | ||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Mask #1
File | emd_28197_msk_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Additional map: Escherichia coli 70S ribosome bound to thermorubin, deacylated...
File | emd_28197_additional_1.map | ||||||||||||
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Annotation | Escherichia coli 70S ribosome bound to thermorubin, deacylated P-site tRNAfMet and aminoacylated A-site Phe-tRNA. Sharpened map. | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Escherichia coli 70S ribosome bound to thermorubin, deacylated...
File | emd_28197_half_map_1.map | ||||||||||||
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Annotation | Escherichia coli 70S ribosome bound to thermorubin, deacylated P-site tRNAfMet and aminoacylated A-site Phe-tRNA. Half-map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Half map: Escherichia coli 70S ribosome bound to thermorubin, deacylated...
File | emd_28197_half_map_2.map | ||||||||||||
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Annotation | Escherichia coli 70S ribosome bound to thermorubin, deacylated P-site tRNAfMet and aminoacylated A-site Phe-tRNA. Half-map | ||||||||||||
Projections & Slices |
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Density Histograms |
-Sample components
+Entire : Escherichia coli 70S ribosome bound to thermorubin, deacylated P-...
+Supramolecule #1: Escherichia coli 70S ribosome bound to thermorubin, deacylated P-...
+Macromolecule #1: 16S Ribosomal RNA
+Macromolecule #22: A-site phenylalanine tRNA
+Macromolecule #23: P-site initiator tRNA
+Macromolecule #24: M-F-Stop mRNA
+Macromolecule #25: 23S Ribosomal RNA
+Macromolecule #26: 5S Ribosomal RNA
+Macromolecule #2: 30S ribosomal protein S2
+Macromolecule #3: 30S ribosomal protein S3
+Macromolecule #4: 30S ribosomal protein S4
+Macromolecule #5: 30S ribosomal protein S5
+Macromolecule #6: 30S ribosomal protein S6
+Macromolecule #7: 30S ribosomal protein S7
+Macromolecule #8: 30S ribosomal protein S8
+Macromolecule #9: 30S ribosomal protein S9
+Macromolecule #10: 30S ribosomal protein S10
+Macromolecule #11: 30S ribosomal protein S11
+Macromolecule #12: 30S ribosomal protein S12
+Macromolecule #13: 30S ribosomal protein S13
+Macromolecule #14: 30S ribosomal protein S14
+Macromolecule #15: 30S ribosomal protein S15
+Macromolecule #16: 30S ribosomal protein S16
+Macromolecule #17: 30S ribosomal protein S17
+Macromolecule #18: 30S ribosomal protein S18
+Macromolecule #19: 30S ribosomal protein S19
+Macromolecule #20: 30S ribosomal protein S20
+Macromolecule #21: 30S ribosomal protein S21
+Macromolecule #27: 50S ribosomal protein L2
+Macromolecule #28: 50S ribosomal protein L3
+Macromolecule #29: 50S ribosomal protein L4
+Macromolecule #30: 50S ribosomal protein L5
+Macromolecule #31: 50S ribosomal protein L6
+Macromolecule #32: 50S ribosomal protein L9
+Macromolecule #33: 50S ribosomal protein L13
+Macromolecule #34: 50S ribosomal protein L14
+Macromolecule #35: 50S Ribosomal Protein L15
+Macromolecule #36: 50S ribosomal protein L16
+Macromolecule #37: 50S ribosomal protein L17
+Macromolecule #38: 50S ribosomal protein L18
+Macromolecule #39: 50S ribosomal protein L19
+Macromolecule #40: 50S ribosomal protein L20
+Macromolecule #41: Ribosomal protein L21
+Macromolecule #42: 50S ribosomal protein L22
+Macromolecule #43: 50S ribosomal protein L23
+Macromolecule #44: 50S ribosomal protein L24
+Macromolecule #45: 50S ribosomal protein L25
+Macromolecule #46: 50S ribosomal protein L27
+Macromolecule #47: 50S ribosomal protein L28
+Macromolecule #48: 50S ribosomal protein L29
+Macromolecule #49: 50S ribosomal protein L30
+Macromolecule #50: 50S ribosomal protein L31
+Macromolecule #51: 50S ribosomal protein L32
+Macromolecule #52: 50S ribosomal protein L33
+Macromolecule #53: 50S ribosomal protein L34
+Macromolecule #54: 50S ribosomal protein L35
+Macromolecule #55: 50S ribosomal protein L36
+Macromolecule #56: MAGNESIUM ION
+Macromolecule #57: Thermorubin
+Macromolecule #58: PHENYLALANINE
+Macromolecule #59: ZINC ION
+Macromolecule #60: water
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 7.4 |
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Grid | Model: Quantifoil R2/1 / Material: GOLD / Mesh: 200 / Support film - Material: GOLD / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 30 sec. / Pretreatment - Atmosphere: OTHER |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 85 % / Chamber temperature: 295 K / Instrument: LEICA EM GP |
-Electron microscopy
Microscope | TFS KRIOS |
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Software | Name: EPU (ver. 2.11.1) |
Image recording | Film or detector model: FEI FALCON III (4k x 4k) / Number grids imaged: 1 / Number real images: 10032 / Average exposure time: 1.0 sec. / Average electron dose: 40.0 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: 2.3000000000000003 µm / Nominal defocus min: 1.0 µm / Nominal magnification: 96000 |
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
Initial model | PDB ID: Chain - Source name: PDB / Chain - Initial model type: experimental model |
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Software | Name: UCSF Chimera (ver. 1.14) |
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
Output model | PDB-8ekc: |