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- EMDB-7349: CryoEM structure of E.coli RNA polymerase elongation complex boun... -
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Open data
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Basic information
Entry | Database: EMDB / ID: EMD-7349 | |||||||||
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Title | CryoEM structure of E.coli RNA polymerase elongation complex bound with RfaH | |||||||||
![]() | primary map | |||||||||
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![]() | RNAP / elongation complex / anti-pausing / ops / TRANSCRIPTION / transcription-dna-rna complex | |||||||||
Function / homology | ![]() regulatory RNA binding / transcription antitermination factor activity, DNA binding / translation activator activity / DNA-templated transcription elongation / bacterial-type RNA polymerase core enzyme binding / transcription elongation-coupled chromatin remodeling / RNA polymerase complex / submerged biofilm formation / cellular response to cell envelope stress / cytosolic DNA-directed RNA polymerase complex ...regulatory RNA binding / transcription antitermination factor activity, DNA binding / translation activator activity / DNA-templated transcription elongation / bacterial-type RNA polymerase core enzyme binding / transcription elongation-coupled chromatin remodeling / RNA polymerase complex / submerged biofilm formation / cellular response to cell envelope stress / 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 / transcription antitermination / DNA-templated transcription initiation / positive regulation of translation / 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 / cytoplasm / cytosol Similarity search - Function | |||||||||
Biological species | ![]() ![]() ![]() ![]() | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.7 Å | |||||||||
![]() | Kang JY / Darst SA | |||||||||
Funding support | ![]()
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![]() | ![]() Title: Structural Basis for Transcript Elongation Control by NusG Family Universal Regulators. Authors: Jin Young Kang / Rachel Anne Mooney / Yuri Nedialkov / Jason Saba / Tatiana V Mishanina / Irina Artsimovitch / Robert Landick / Seth A Darst / ![]() Abstract: NusG/RfaH/Spt5 transcription elongation factors are the only transcription regulators conserved across all life. Bacterial NusG regulates RNA polymerase (RNAP) elongation complexes (ECs) across most ...NusG/RfaH/Spt5 transcription elongation factors are the only transcription regulators conserved across all life. Bacterial NusG regulates RNA polymerase (RNAP) elongation complexes (ECs) across most genes, enhancing elongation by suppressing RNAP backtracking and coordinating ρ-dependent termination and translation. The NusG paralog RfaH engages the EC only at operon polarity suppressor (ops) sites and suppresses both backtrack and hairpin-stabilized pausing. We used single-particle cryoelectron microscopy (cryo-EM) to determine structures of ECs at ops with NusG or RfaH. Both factors chaperone base-pairing of the upstream duplex DNA to suppress backtracking, explaining stimulation of elongation genome-wide. The RfaH-opsEC structure reveals how RfaH confers operon specificity through specific recognition of an ops hairpin in the single-stranded nontemplate DNA and tighter binding to the EC to exclude NusG. Tight EC binding by RfaH sterically blocks the swiveled RNAP conformation necessary for hairpin-stabilized pausing. The universal conservation of NusG/RfaH/Spt5 suggests that the molecular mechanisms uncovered here are widespread. | |||||||||
History |
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Structure visualization
Movie |
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Structure viewer | EM map: ![]() ![]() ![]() |
Supplemental images |
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Downloads & links
-EMDB archive
Map data | ![]() | 59.9 MB | ![]() | |
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Header (meta data) | ![]() ![]() | 25.1 KB 25.1 KB | Display Display | ![]() |
Images | ![]() | 73.5 KB | ||
Filedesc metadata | ![]() | 8.8 KB | ||
Archive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 6c6sMC ![]() 7350C ![]() 7351C ![]() 6c6tC ![]() 6c6uC C: citing same article ( M: atomic model generated by this map |
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Similar structure data |
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Links
EMDB pages | ![]() ![]() |
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Related items in Molecule of the Month |
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Map
File | ![]() | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
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Annotation | primary map | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.3 Å | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Density |
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Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
CCP4 map header:
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-Supplemental data
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Sample components
+Entire : E.coli RfaH-elongation complex
+Supramolecule #1: E.coli RfaH-elongation complex
+Macromolecule #1: DNA (29-MER)
+Macromolecule #2: DNA (29-MER)
+Macromolecule #3: RNA (5'-R(*GP*CP*AP*UP*UP*CP*AP*AP*AP*GP*CP*CP*GP*AP*GP*AP*GP*GP*...
+Macromolecule #4: DNA-directed RNA polymerase subunit alpha
+Macromolecule #5: DNA-directed RNA polymerase subunit beta
+Macromolecule #6: DNA-directed RNA polymerase subunit beta'
+Macromolecule #7: DNA-directed RNA polymerase subunit omega
+Macromolecule #8: Transcription antitermination protein RfaH
+Macromolecule #9: MAGNESIUM ION
+Macromolecule #10: ZINC ION
-Experimental details
-Structure determination
Method | cryo EM |
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![]() | single particle reconstruction |
Aggregation state | particle |
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Sample preparation
Concentration | 5 mg/mL | |||||||||||||||
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Buffer | pH: 8 Component:
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Grid | Model: C-flat-1.2/1.3 4C / Material: GOLD / Support film - Material: CARBON / Support film - topology: HOLEY / Pretreatment - Type: GLOW DISCHARGE / Pretreatment - Time: 15 sec. | |||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295.15 K / Instrument: FEI VITROBOT MARK IV |
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Electron microscopy
Microscope | FEI TITAN KRIOS |
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Image recording | Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 3-50 / Number grids imaged: 2 / Number real images: 3495 / Average exposure time: 15.0 sec. / Average electron dose: 71.0 e/Å2 |
Electron beam | Acceleration voltage: 300 kV / Electron source: ![]() |
Electron optics | C2 aperture diameter: 70.0 µm / Calibrated defocus max: 3.0 µm / Calibrated defocus min: 0.5 µm / Calibrated magnification: 38462 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm / Nominal defocus max: 2.4 µm / Nominal defocus min: 0.8 µm |
Sample stage | Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Image processing
-Atomic model buiding 1
Refinement | Space: REAL / Protocol: RIGID BODY FIT |
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Output model | ![]() PDB-6c6s: |