+Open data
-Basic information
Entry | Database: EMDB / ID: EMD-28794 | |||||||||
---|---|---|---|---|---|---|---|---|---|---|
Title | EsN-dhsU36mm1 local map (map 2) | |||||||||
Map data | Raw map | |||||||||
Sample |
| |||||||||
Keywords | promoter-bound / initiation / TRANSCRIPTION | |||||||||
Biological species | Escherichia coli (E. coli) / Aquifex aeolicus (bacteria) | |||||||||
Method | single particle reconstruction / cryo EM / Resolution: 3.0 Å | |||||||||
Authors | Mueller AU / Chen J / Darst SA | |||||||||
Funding support | United States, 1 items
| |||||||||
Citation | Journal: Proc Natl Acad Sci U S A / Year: 2023 Title: A general mechanism for transcription bubble nucleation in bacteria. Authors: Andreas U Mueller / James Chen / Mengyu Wu / Courtney Chiu / B Tracy Nixon / Elizabeth A Campbell / Seth A Darst / Abstract: Bacterial transcription initiation requires σ factors for nucleation of the transcription bubble. The canonical housekeeping σ factor, σ, nucleates DNA melting via recognition of conserved bases ...Bacterial transcription initiation requires σ factors for nucleation of the transcription bubble. The canonical housekeeping σ factor, σ, nucleates DNA melting via recognition of conserved bases of the promoter -10 motif, which are unstacked and captured in pockets of σ. By contrast, the mechanism of transcription bubble nucleation and formation during the unrelated σ-mediated transcription initiation is poorly understood. Herein, we combine structural and biochemical approaches to establish that σ, like σ, captures a flipped, unstacked base in a pocket formed between its N-terminal region I (RI) and extra-long helix features. Strikingly, RI inserts into the nascent bubble to stabilize the nucleated bubble prior to engagement of the obligate ATPase activator. Our data suggest a general paradigm of transcription initiation that requires σ factors to nucleate an early melted intermediate prior to productive RNA synthesis. | |||||||||
History |
|
-Structure visualization
Supplemental images |
---|
-Downloads & links
-EMDB archive
Map data | emd_28794.map.gz | 32.2 MB | EMDB map data format | |
---|---|---|---|---|
Header (meta data) | emd-28794-v30.xml emd-28794.xml | 22.1 KB 22.1 KB | Display Display | EMDB header |
FSC (resolution estimation) | emd_28794_fsc.xml | 8.9 KB | Display | FSC data file |
Images | emd_28794.png | 78.1 KB | ||
Masks | emd_28794_msk_1.map emd_28794_msk_2.map | 64 MB 64 MB | Mask map | |
Filedesc metadata | emd-28794.cif.gz | 4.9 KB | ||
Others | emd_28794_additional_1.map.gz emd_28794_additional_2.map.gz emd_28794_additional_3.map.gz emd_28794_half_map_1.map.gz emd_28794_half_map_2.map.gz | 59.8 MB 2.4 MB 3 MB 59.4 MB 59.4 MB | ||
Archive directory | http://ftp.pdbj.org/pub/emdb/structures/EMD-28794 ftp://ftp.pdbj.org/pub/emdb/structures/EMD-28794 | HTTPS FTP |
-Validation report
Summary document | emd_28794_validation.pdf.gz | 878.9 KB | Display | EMDB validaton report |
---|---|---|---|---|
Full document | emd_28794_full_validation.pdf.gz | 878.5 KB | Display | |
Data in XML | emd_28794_validation.xml.gz | 16.5 KB | Display | |
Data in CIF | emd_28794_validation.cif.gz | 21.2 KB | Display | |
Arichive directory | https://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28794 ftp://ftp.pdbj.org/pub/emdb/validation_reports/EMD-28794 | HTTPS FTP |
-Related structure data
-Links
EMDB pages | EMDB (EBI/PDBe) / EMDataResource |
---|---|
Related items in Molecule of the Month |
-Map
File | Download / File: emd_28794.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES) | ||||||||||||||||||||||||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Raw map | ||||||||||||||||||||||||||||||||||||
Projections & slices | Image control
Images are generated by Spider. | ||||||||||||||||||||||||||||||||||||
Voxel size | X=Y=Z: 1.083 Å | ||||||||||||||||||||||||||||||||||||
Density |
| ||||||||||||||||||||||||||||||||||||
Symmetry | Space group: 1 | ||||||||||||||||||||||||||||||||||||
Details | EMDB XML:
|
-Supplemental data
-Mask #1
File | emd_28794_msk_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Mask #2
File | emd_28794_msk_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Projections & Slices |
| ||||||||||||
Density Histograms |
-Additional map: B-factor sharpened map (-110.1)
File | emd_28794_additional_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | B-factor sharpened map (-110.1) | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Additional map: Locally filtered map (blocfilt)
File | emd_28794_additional_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Locally filtered map (blocfilt) | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Additional map: Local resolution map (blocres)
File | emd_28794_additional_3.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Local resolution map (blocres) | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: Half map A
File | emd_28794_half_map_1.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Half map A | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Half map: Half map B
File | emd_28794_half_map_2.map | ||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Annotation | Half map B | ||||||||||||
Projections & Slices |
| ||||||||||||
Density Histograms |
-Sample components
-Entire : EsN-dhsU36mm1
Entire | Name: EsN-dhsU36mm1 |
---|---|
Components |
|
-Supramolecule #1: EsN-dhsU36mm1
Supramolecule | Name: EsN-dhsU36mm1 / type: complex / ID: 1 / Parent: 0 |
---|
-Supramolecule #2: RNA polymerase complex
Supramolecule | Name: RNA polymerase complex / type: complex / ID: 2 / Parent: 1 |
---|---|
Source (natural) | Organism: Escherichia coli (E. coli) |
-Supramolecule #3: DNA
Supramolecule | Name: DNA / type: complex / ID: 3 / Parent: 1 |
---|---|
Source (natural) | Organism: Aquifex aeolicus (bacteria) / Synthetically produced: Yes |
-Experimental details
-Structure determination
Method | cryo EM |
---|---|
Processing | single particle reconstruction |
Aggregation state | particle |
-Sample preparation
Buffer | pH: 8 Component:
| |||||||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Grid | Model: C-flat-1.2/1.3 / Material: GOLD / Mesh: 400 / Support film - Material: CARBON / Support film - topology: HOLEY | |||||||||||||||
Vitrification | Cryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 295 K / Instrument: FEI VITROBOT MARK IV Details: Octyl beta-D-glucopyranoside (beta-OG) was added to the sample to 0.1%w/v final concentration (from 10x stock) just prior to plunge vitrification. |
-Electron microscopy
Microscope | FEI TITAN KRIOS |
---|---|
Specialist optics | Energy filter - Name: GIF Bioquantum / Energy filter - Slit width: 20 eV |
Image recording | Film or detector model: GATAN K3 (6k x 4k) / Number real images: 12294 / Average exposure time: 2.0 sec. / Average electron dose: 51.0 e/Å2 Details: dose-fractionation with 0.05 seconds per frame (=40 frames) |
Electron beam | Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN |
Electron optics | Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Nominal defocus max: 2.5 µm / Nominal defocus min: 1.0 µm |
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |