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Yorodumi- PDB-8h40: Cryo-EM structure of the transcription activation complex NtcA-TAC -
+Open data
-Basic information
Entry | Database: PDB / ID: 8h40 | |||||||||||||||
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Title | Cryo-EM structure of the transcription activation complex NtcA-TAC | |||||||||||||||
Components |
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Keywords | TRANSCRIPTION / Transcription activation complex | |||||||||||||||
Function / homology | Function and homology information sigma factor activity / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / DNA-binding transcription factor activity / DNA-templated transcription / magnesium ion binding ...sigma factor activity / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / DNA-binding transcription factor activity / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / cytoplasm Similarity search - Function | |||||||||||||||
Biological species | Anabaena (bacteria) | |||||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å | |||||||||||||||
Authors | Han, S.J. / Jiang, Y.L. / You, L.L. / Shen, L.Q. / Wu, X.X. / Yang, F. / Kong, W.W. / Chen, Z.P. / Zhang, Y. / Zhou, C.Z. | |||||||||||||||
Funding support | China, 4items
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Citation | Journal: Nat Struct Mol Biol / Year: 2024 Title: DNA looping mediates cooperative transcription activation. Authors: Shu-Jing Han / Yong-Liang Jiang / Lin-Lin You / Li-Qiang Shen / Xiaoxian Wu / Feng Yang / Ning Cui / Wen-Wen Kong / Hui Sun / Ke Zhou / Hui-Chao Meng / Zhi-Peng Chen / Yuxing Chen / Yu Zhang / Cong-Zhao Zhou / Abstract: Transcription factors respond to multilevel stimuli and co-occupy promoter regions of target genes to activate RNA polymerase (RNAP) in a cooperative manner. To decipher the molecular mechanism, here ...Transcription factors respond to multilevel stimuli and co-occupy promoter regions of target genes to activate RNA polymerase (RNAP) in a cooperative manner. To decipher the molecular mechanism, here we report two cryo-electron microscopy structures of Anabaena transcription activation complexes (TACs): NtcA-TAC composed of RNAP holoenzyme, promoter and a global activator NtcA, and NtcA-NtcB-TAC comprising an extra context-specific regulator, NtcB. Structural analysis showed that NtcA binding makes the promoter DNA bend by ∼50°, which facilitates RNAP to contact NtcB at the distal upstream NtcB box. The sequential binding of NtcA and NtcB induces looping back of promoter DNA towards RNAP, enabling the assembly of a fully activated TAC bound with two activators. Together with biochemical assays, we propose a 'DNA looping' mechanism of cooperative transcription activation in bacteria. | |||||||||||||||
History |
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-Structure visualization
Structure viewer | Molecule: MolmilJmol/JSmol |
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-Downloads & links
-Download
PDBx/mmCIF format | 8h40.cif.gz | 850.7 KB | Display | PDBx/mmCIF format |
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PDB format | pdb8h40.ent.gz | 628.9 KB | Display | PDB format |
PDBx/mmJSON format | 8h40.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/h4/8h40 ftp://data.pdbj.org/pub/pdb/validation_reports/h4/8h40 | HTTPS FTP |
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-Related structure data
Related structure data | 34476MC 8h3vC 8h3zC C: citing same article (ref.) M: map data used to model this data |
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Similar structure data | Similarity search - Function & homologyF&H Search |
-Links
-Assembly
Deposited unit |
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1 |
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-Components
-DNA chain , 2 types, 2 molecules 12
#1: DNA chain | Mass: 38723.902 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Anabaena (bacteria) |
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#2: DNA chain | Mass: 38571.723 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Anabaena (bacteria) |
-DNA-directed RNA polymerase subunit ... , 5 types, 6 molecules ABCDEF
#3: Protein | Mass: 126781.531 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Anabaena (bacteria) / Strain: PCC 7120 / SAG 25.82 / UTEX 2576 / Gene: rpoB, alr1594 / Production host: Escherichia coli (E. coli) / References: UniProt: P22703, DNA-directed RNA polymerase | ||||
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#4: Protein | Mass: 147137.484 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Anabaena (bacteria) / Strain: PCC 7120 / SAG 25.82 / UTEX 2576 / Gene: rpoC2, alr1596 / Production host: Escherichia coli (E. coli) / References: UniProt: P22705, DNA-directed RNA polymerase | ||||
#5: Protein | Mass: 26166.613 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Anabaena (bacteria) / Strain: PCC 7120 / SAG 25.82 / UTEX 2576 / Gene: rpoA, all4191 / Production host: Escherichia coli (E. coli) / References: UniProt: Q8YPK3, DNA-directed RNA polymerase #6: Protein | | Mass: 70653.055 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Anabaena (bacteria) / Strain: PCC 7120 / SAG 25.82 / UTEX 2576 / Gene: rpoC1, alr1595 / Production host: Escherichia coli (E. coli) / References: UniProt: P22704, DNA-directed RNA polymerase #7: Protein | | Mass: 9143.490 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Anabaena (bacteria) / Strain: PCC 7120 / SAG 25.82 / UTEX 2576 / Gene: rpoZ, asr4648 / Production host: Escherichia coli (E. coli) / References: UniProt: Q8YNB9, DNA-directed RNA polymerase |
-Protein , 2 types, 3 molecules GXY
#8: Protein | Mass: 45717.027 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Anabaena (bacteria) / Strain: PCC 7120 / SAG 25.82 / UTEX 2576 / Gene: sigA, rpoD, all5263 / Production host: Escherichia coli (E. coli) / References: UniProt: P26683 |
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#9: Protein | Mass: 24951.266 Da / Num. of mol.: 2 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Anabaena (bacteria) / Strain: PCC 7120 / SAG 25.82 / UTEX 2576 / Gene: ntcA, bifA, alr4392 / Production host: Escherichia coli (E. coli) / References: UniProt: P0A4U6 |
-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
-Sample preparation
Component | Name: transcription activation complex with the global regulator NtcA Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES | |||||||||||||||||||||||||
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Molecular weight | Value: 0.58 MDa / Experimental value: YES | |||||||||||||||||||||||||
Source (natural) | Organism: Anabaena (bacteria) / Strain: PCC. 7120 | |||||||||||||||||||||||||
Source (recombinant) | Organism: Escherichia coli (E. coli) | |||||||||||||||||||||||||
Buffer solution | pH: 7.5 Details: 10 mM HEPES pH 7.5, 100 mM KCl, 5 mM MgCl2 and 2 mM DTT | |||||||||||||||||||||||||
Buffer component |
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Specimen | Conc.: 15 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES | |||||||||||||||||||||||||
Specimen support | Grid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: UltrAuFoil R1.2/1.3 | |||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK I / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS / Details: Preliminary grid screening was performed manually. |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 2200 nm / Nominal defocus min: 1200 nm / Cs: 2.7 mm |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER |
Image recording | Electron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k) / Num. of grids imaged: 1 / Num. of real images: 1462 Details: Images were collected in movie-mode at 32 frames per second. |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | ||||||||||||||||||||||||
Particle selection | Num. of particles selected: 555921 | ||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | ||||||||||||||||||||||||
3D reconstruction | Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 45239 / Symmetry type: POINT | ||||||||||||||||||||||||
Atomic model building | Protocol: RIGID BODY FIT | ||||||||||||||||||||||||
Refine LS restraints |
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