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Yorodumi- EMDB-29494: CryoEM structure of E.coli transcription elongation complex bound... -
+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-29494 | ||||||||||||||||||
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Title | CryoEM structure of E.coli transcription elongation complex bound to ppGpp | ||||||||||||||||||
Map data | |||||||||||||||||||
Sample |
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Keywords | Transcription elongation / Second messager / ppGpp / DNA repair / TRANSCRIPTION-DNA-RNA complex | ||||||||||||||||||
Function / homology | Function and homology information RNA polymerase complex / submerged biofilm formation / cellular response to cell envelope stress / bacterial-type RNA polymerase core enzyme binding / cytosolic DNA-directed RNA polymerase complex / regulation of DNA-templated transcription initiation / bacterial-type flagellum assembly / bacterial-type flagellum-dependent cell motility / nitrate assimilation / transcription elongation factor complex ...RNA polymerase complex / submerged biofilm formation / cellular response to cell envelope stress / bacterial-type RNA polymerase core enzyme binding / cytosolic DNA-directed RNA polymerase complex / regulation of DNA-templated transcription initiation / bacterial-type flagellum assembly / bacterial-type flagellum-dependent cell motility / nitrate assimilation / transcription elongation factor complex / regulation of DNA-templated transcription elongation / DNA-templated transcription initiation / transcription antitermination / cell motility / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / response to heat / protein-containing complex assembly / intracellular iron ion homeostasis / protein dimerization activity / response to antibiotic / magnesium ion binding / DNA binding / zinc ion binding / membrane / cytosol / cytoplasm Similarity search - Function | ||||||||||||||||||
Biological species | Escherichia coli K-12 (bacteria) / Escherichia coli (E. coli) | ||||||||||||||||||
Method | single particle reconstruction / cryo EM / Resolution: 2.1 Å | ||||||||||||||||||
Authors | Duan W / Serganov A | ||||||||||||||||||
Funding support | United States, 5 items
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Citation | Journal: Nat Struct Mol Biol / Year: 2023 Title: Control of transcription elongation and DNA repair by alarmone ppGpp. Authors: Jacob W Weaver / Sergey Proshkin / Wenqian Duan / Vitaly Epshtein / Manjunath Gowder / Binod K Bharati / Elena Afanaseva / Alexander Mironov / Alexander Serganov / Evgeny Nudler / Abstract: Second messenger (p)ppGpp (collectively guanosine tetraphosphate and guanosine pentaphosphate) mediates bacterial adaptation to nutritional stress by modulating transcription initiation. More ...Second messenger (p)ppGpp (collectively guanosine tetraphosphate and guanosine pentaphosphate) mediates bacterial adaptation to nutritional stress by modulating transcription initiation. More recently, ppGpp has been implicated in coupling transcription and DNA repair; however, the mechanism of ppGpp engagement remained elusive. Here we present structural, biochemical and genetic evidence that ppGpp controls Escherichia coli RNA polymerase (RNAP) during elongation via a specific site that is nonfunctional during initiation. Structure-guided mutagenesis renders the elongation (but not initiation) complex unresponsive to ppGpp and increases bacterial sensitivity to genotoxic agents and ultraviolet radiation. Thus, ppGpp binds RNAP at sites with distinct functions in initiation and elongation, with the latter being important for promoting DNA repair. Our data provide insights on the molecular mechanism of ppGpp-mediated adaptation during stress, and further highlight the intricate relationships between genome stability, stress responses and transcription. | ||||||||||||||||||
History |
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-Structure visualization
Supplemental images |
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-Downloads & links
-EMDB archive
Map data | emd_29494.map.gz | 230.4 MB | EMDB map data format | |
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Header (meta data) | emd-29494-v30.xml emd-29494.xml | 29.2 KB 29.2 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_29494_fsc.xml | 13.1 KB | Display | FSC data file |
Images | emd_29494.png | 57.4 KB | ||
Filedesc metadata | emd-29494.cif.gz | 9.4 KB | ||
Others | emd_29494_half_map_1.map.gz emd_29494_half_map_2.map.gz | 226.5 MB 226.5 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-29494 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-29494 | HTTPS FTP |
-Validation report
Summary document | emd_29494_validation.pdf.gz | 998 KB | Display | EMDB validaton report |
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Full document | emd_29494_full_validation.pdf.gz | 997.5 KB | Display | |
Data in XML | emd_29494_validation.xml.gz | 21 KB | Display | |
Data in CIF | emd_29494_validation.cif.gz | 27 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-29494 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-29494 | HTTPS FTP |
-Related structure data
Related structure data | 8fvwMC 8fvrC 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_29494.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
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Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 0.825 Å | ||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
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-Supplemental data
-Half map: #2
File | emd_29494_half_map_1.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Half map: #1
File | emd_29494_half_map_2.map | ||||||||||||
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Projections & Slices |
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Density Histograms |
-Sample components
+Entire : E. coli transcription elongation complex bound to ppGpp (EC-ppGpp)
+Supramolecule #1: E. coli transcription elongation complex bound to ppGpp (EC-ppGpp)
+Macromolecule #1: DNA-directed RNA polymerase subunit alpha
+Macromolecule #2: DNA-directed RNA polymerase subunit beta
+Macromolecule #3: DNA-directed RNA polymerase subunit beta'
+Macromolecule #4: DNA-directed RNA polymerase subunit omega
+Macromolecule #5: DNA (53-MER)
+Macromolecule #7: DNA (53-MER)
+Macromolecule #6: RNA (16-MER)
+Macromolecule #8: GUANOSINE-5',3'-TETRAPHOSPHATE
+Macromolecule #9: ZINC ION
+Macromolecule #10: MAGNESIUM ION
+Macromolecule #11: SODIUM ION
+Macromolecule #12: water
-Experimental details
-Structure determination
Method | cryo EM |
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Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Concentration | 6.5 mg/mL |
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Buffer | pH: 8 Details: 20 mM HEPES, pH 8.0, 150 mM NaCl, 10 mM MgCl2, 10 uM ZnCl2, 5 mM DTT |
Grid | Model: Quantifoil R1.2/1.3 / Material: GOLD / Mesh: 300 / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 25 sec. / Pretreatment - Atmosphere: AIR / Pretreatment - Pressure: 260.0 kPa |
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 283.15 K / Instrument: FEI VITROBOT MARK IV |
Details | Elongation Complex was mixed with 0.1 mM ppGpp (TriLink) for 20 min at room temperature followed by addition of 8 mM CHAPSO. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
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Temperature | Min: 79.95 K / Max: 79.95 K |
Alignment procedure | Coma free - Residual tilt: 0.05 mrad |
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k) / Digitization - Dimensions - Width: 5760 pixel / Digitization - Dimensions - Height: 4092 pixel / Number grids imaged: 1 / Number real images: 4921 / Average exposure time: 2.0 sec. / Average electron dose: 57.54 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | C2 aperture diameter: 70.0 µm / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 1.9000000000000001 µm / Nominal defocus min: 0.9 µm / Nominal magnification: 105000 |
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 | Protocol: RIGID BODY FIT |
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Output model | PDB-8fvw: |