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- PDB-6exv: Structure of mammalian RNA polymerase II elongation complex inhib... -
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Basic information
Entry | Database: PDB / ID: 6exv | ||||||||||||
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Title | Structure of mammalian RNA polymerase II elongation complex inhibited by Alpha-amanitin | ||||||||||||
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![]() | TRANSCRIPTION / Inhibitor / elongation / active site | ||||||||||||
Function / homology | ![]() B-WICH complex positively regulates rRNA expression / RNA Polymerase I Transcription Initiation / RNA Polymerase I Promoter Escape / RNA Polymerase I Transcription Termination / RNA Polymerase III Transcription Initiation From Type 1 Promoter / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Polymerase III Transcription Initiation From Type 3 Promoter / Formation of RNA Pol II elongation complex / Formation of the Early Elongation Complex / Transcriptional regulation by small RNAs ...B-WICH complex positively regulates rRNA expression / RNA Polymerase I Transcription Initiation / RNA Polymerase I Promoter Escape / RNA Polymerase I Transcription Termination / RNA Polymerase III Transcription Initiation From Type 1 Promoter / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Polymerase III Transcription Initiation From Type 3 Promoter / Formation of RNA Pol II elongation complex / Formation of the Early Elongation Complex / Transcriptional regulation by small RNAs / RNA Polymerase II Pre-transcription Events / TP53 Regulates Transcription of DNA Repair Genes / FGFR2 alternative splicing / RNA polymerase II transcribes snRNA genes / mRNA Capping / mRNA Splicing - Major Pathway / mRNA Splicing - Minor Pathway / Processing of Capped Intron-Containing Pre-mRNA / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Elongation / RNA Polymerase II Transcription Initiation And Promoter Clearance / RNA Pol II CTD phosphorylation and interaction with CE / Estrogen-dependent gene expression / Formation of TC-NER Pre-Incision Complex / Dual incision in TC-NER / Gap-filling DNA repair synthesis and ligation in TC-NER / : / : / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / organelle membrane / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / transcription elongation by RNA polymerase I / positive regulation of translational initiation / RNA polymerase I complex / RNA polymerase III complex / transcription-coupled nucleotide-excision repair / RNA polymerase III activity / RNA polymerase II, core complex / tRNA transcription by RNA polymerase III / RNA polymerase I activity / RNA polymerase II activity / core promoter sequence-specific DNA binding / translation initiation factor binding / transcription initiation at RNA polymerase II promoter / P-body / euchromatin / ribonucleoside binding / fibrillar center / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / single-stranded DNA binding / toxin activity / transcription by RNA polymerase II / nucleic acid binding / chromosome, telomeric region / single-stranded RNA binding / protein dimerization activity / nuclear speck / RNA-dependent RNA polymerase activity / nucleotide binding / chromatin binding / nucleolus / DNA binding / zinc ion binding / nucleus / metal ion binding / cytosol Similarity search - Function | ||||||||||||
Biological species | ![]() ![]() ![]() ![]() ![]() synthetic construct (others) | ||||||||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.6 Å | ||||||||||||
![]() | Liu, X. / Farnung, L. / Wigge, C. / Cramer, P. | ||||||||||||
Funding support | ![]()
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![]() | ![]() Title: Cryo-EM structure of a mammalian RNA polymerase II elongation complex inhibited by α-amanitin. Authors: Xiangyang Liu / Lucas Farnung / Christoph Wigge / Patrick Cramer / ![]() Abstract: RNA polymerase II (Pol II) is the central enzyme that transcribes eukaryotic protein-coding genes to produce mRNA. The mushroom toxin α-amanitin binds Pol II and inhibits transcription at the step ...RNA polymerase II (Pol II) is the central enzyme that transcribes eukaryotic protein-coding genes to produce mRNA. The mushroom toxin α-amanitin binds Pol II and inhibits transcription at the step of RNA chain elongation. Pol II from yeast binds α-amanitin with micromolar affinity, whereas metazoan Pol II enzymes exhibit nanomolar affinities. Here, we present the high-resolution cryo-EM structure of α-amanitin bound to and inhibited by its natural target, the mammalian Pol II elongation complex. The structure revealed that the toxin is located in a pocket previously identified in yeast Pol II but forms additional contacts with metazoan-specific residues, which explains why its affinity to mammalian Pol II is ∼3000 times higher than for yeast Pol II. Our work provides the structural basis for the inhibition of mammalian Pol II by the natural toxin α-amanitin and highlights that cryo-EM is well suited to studying interactions of a small molecule with its macromolecular target. | ||||||||||||
History |
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Structure visualization
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Structure viewer | Molecule: ![]() ![]() |
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PDBx/mmCIF format | ![]() | 752.4 KB | Display | ![]() |
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PDB format | ![]() | 604.6 KB | Display | ![]() |
PDBx/mmJSON format | ![]() | Tree view | ![]() | |
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-Validation report
Summary document | ![]() | 1.2 MB | Display | ![]() |
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Full document | ![]() | 1.2 MB | Display | |
Data in XML | ![]() | 111.2 KB | Display | |
Data in CIF | ![]() | 174.3 KB | Display | |
Arichive directory | ![]() ![]() | HTTPS FTP |
-Related structure data
Related structure data | ![]() 3981MC M: map data used to model this data C: citing same article ( |
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Similar structure data |
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Links
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Assembly
Deposited unit | ![]()
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Components
-DNA-DIRECTED RNA POLYMERASE II SUBUNIT ... , 6 types, 6 molecules ABCDGI
#1: Protein | Mass: 217450.078 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
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#2: Protein | Mass: 133201.625 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#3: Protein | Mass: 31439.074 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#4: Protein | Mass: 16331.255 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#7: Protein | Mass: 19314.283 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#9: Protein | Mass: 14541.221 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
-DNA-DIRECTED RNA POLYMERASES I, II, AND III SUBUNIT ... , 6 types, 6 molecules EFHJKL
#5: Protein | Mass: 24644.318 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
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#6: Protein | Mass: 14477.001 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#8: Protein | Mass: 17162.273 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#10: Protein | Mass: 7655.123 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#11: Protein | Mass: 13310.284 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
#12: Protein | Mass: 7018.244 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) ![]() ![]() |
-DNA chain , 2 types, 2 molecules NT
#14: DNA chain | Mass: 13303.563 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
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#16: DNA chain | Mass: 13178.483 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others) |
-Protein/peptide / RNA chain , 2 types, 2 molecules MP
-Non-polymers , 2 types, 9 molecules ![](data/chem/img/ZN.gif)
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#17: Chemical | ChemComp-ZN / #18: Chemical | ChemComp-MG / | |
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-Details
Compound details | ALPHA-AMANITIN, AN AMATOXIN, IS A DI-CYCLIC PEPTIDE. HERE, ALPHA-AMANITIN IS REPRESENTED BY THE ...ALPHA-AMANITIN, AN AMATOXIN, IS A DI-CYCLIC PEPTIDE. HERE, ALPHA-AMANITIN IS REPRESENTE |
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-Experimental details
-Experiment
Experiment | Method: ELECTRON MICROSCOPY |
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EM experiment | Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction |
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Sample preparation
Component |
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Molecular weight | Value: 0.607 MDa / Experimental value: NO | ||||||||||||||||||||||||||||||
Source (natural) |
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Source (recombinant) | Organism: synthetic construct (others) | ||||||||||||||||||||||||||||||
Buffer solution | pH: 7.6 | ||||||||||||||||||||||||||||||
Specimen | Conc.: 0.244 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: Quantifoil R2/2 | ||||||||||||||||||||||||||||||
Vitrification | Instrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277 K |
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Electron microscopy imaging
Experimental equipment | ![]() Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: FEI TITAN KRIOS |
Electron gun | Electron source: ![]() |
Electron lens | Mode: BRIGHT FIELD / Nominal magnification: 130000 X / Nominal defocus max: 2500 nm / Nominal defocus min: 1500 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE |
Specimen holder | Cryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Residual tilt: 10 mradians |
Image recording | Average exposure time: 10 sec. / Electron dose: 35 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 1 / Num. of real images: 2264 |
EM imaging optics | Energyfilter name: GIF Quantum LS / Energyfilter upper: 20 eV / Energyfilter lower: 0 eV |
Image scans | Width: 3710 / Height: 3838 / Movie frames/image: 40 |
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Processing
Software | Name: PHENIX / Version: 1.11.1_2575: / Classification: refinement | |||||||||||||||||||||||||||
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EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||||||||||||||||||||
Particle selection | Num. of particles selected: 207410 | |||||||||||||||||||||||||||
Symmetry | Point symmetry: C1 (asymmetric) | |||||||||||||||||||||||||||
3D reconstruction | Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 134512 / Symmetry type: POINT | |||||||||||||||||||||||||||
Atomic model building | B value: 53.97 / Protocol: FLEXIBLE FIT / Space: REAL | |||||||||||||||||||||||||||
Refine LS restraints |
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