+Open data
-Basic information
Entry | Database: PDB / ID: 7kpw | ||||||
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Title | Structure of the H-lobe of yeast CKM | ||||||
Components | Mediator of RNA polymerase II transcription subunit 12 | ||||||
Keywords | TRANSCRIPTION / Mediator / Cdk8 / Med13 / Med12 / CycC / CDK / Argonaute / RNA Polymerase II / PIWI | ||||||
Function / homology | Function and homology information positive regulation of transcription by galactose / CKM complex / mediator complex / transcription coactivator activity / negative regulation of transcription by RNA polymerase II / positive regulation of transcription by RNA polymerase II Similarity search - Function | ||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.9 Å | ||||||
Authors | Li, Y.C. / Chao, T.C. / Kim, H.J. / Cholko, T. / Chen, S.F. / Nakanishi, K. / Chang, C.E. / Murakami, K. / Garcia, B.A. / Boyer, T.G. / Tsai, K.L. | ||||||
Citation | Journal: Sci Adv / Year: 2021 Title: Structure and noncanonical Cdk8 activation mechanism within an Argonaute-containing Mediator kinase module. Authors: Yi-Chuan Li / Ti-Chun Chao / Hee Jong Kim / Timothy Cholko / Shin-Fu Chen / Guojie Li / Laura Snyder / Kotaro Nakanishi / Chia-En Chang / Kenji Murakami / Benjamin A Garcia / Thomas G Boyer / Kuang-Lei Tsai / Abstract: The Cdk8 kinase module (CKM) in Mediator, comprising Med13, Med12, CycC, and Cdk8, regulates RNA polymerase II transcription through kinase-dependent and -independent functions. Numerous pathogenic ...The Cdk8 kinase module (CKM) in Mediator, comprising Med13, Med12, CycC, and Cdk8, regulates RNA polymerase II transcription through kinase-dependent and -independent functions. Numerous pathogenic mutations causative for neurodevelopmental disorders and cancer congregate in CKM subunits. However, the structure of the intact CKM and the mechanism by which Cdk8 is non-canonically activated and functionally affected by oncogenic CKM alterations are poorly understood. Here, we report a cryo-electron microscopy structure of CKM that redefines prior CKM structural models and explains the mechanism of Med12-dependent Cdk8 activation. Med12 interacts extensively with CycC and activates Cdk8 by stabilizing its activation (T-)loop through conserved Med12 residues recurrently mutated in human tumors. Unexpectedly, Med13 has a characteristic Argonaute-like bi-lobal architecture. These findings not only provide a structural basis for understanding CKM function and pathological dysfunction, but also further impute a previously unknown regulatory mechanism of Mediator in transcriptional modulation through its Med13 Argonaute-like features. | ||||||
History |
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-Structure visualization
Movie |
Movie viewer |
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Structure viewer | Molecule: MolmilJmol/JSmol |
-Downloads & links
-Download
PDBx/mmCIF format | 7kpw.cif.gz | 145.8 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7kpw.ent.gz | 90.5 KB | Display | PDB format |
PDBx/mmJSON format | 7kpw.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7kpw_validation.pdf.gz | 1.1 MB | Display | wwPDB validaton report |
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Full document | 7kpw_full_validation.pdf.gz | 1.1 MB | Display | |
Data in XML | 7kpw_validation.xml.gz | 37.9 KB | Display | |
Data in CIF | 7kpw_validation.cif.gz | 57.8 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/kp/7kpw ftp://data.pdbj.org/pub/pdb/validation_reports/kp/7kpw | HTTPS FTP |
-Related structure data
Related structure data | 22990MC 7kpvC 7kpxC M: map data used to model this data C: citing same article (ref.) |
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Similar structure data |
-Links
-Assembly
Deposited unit |
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-Components
#1: Protein | Mass: 167049.812 Da / Num. of mol.: 1 Source method: isolated from a genetically manipulated source Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / Gene: SRB8, GIG1, MED12, NUT6, SSN5, YCR081W, YCR81W/YCR80W / Production host: Saccharomyces cerevisiae S288C (yeast) / References: UniProt: P25648 |
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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 |
-Sample preparation
Component | Name: yeast CDK8 complex / Type: COMPLEX / Entity ID: all / Source: NATURAL |
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Molecular weight | Value: 0.43 MDa / Experimental value: NO |
Source (natural) | Organism: Saccharomyces cerevisiae S288C (yeast) |
Buffer solution | pH: 7.4 / Details: 25 mM Hepes pH 7.4, 200 mM NaCl, and 0.005% NP-40 |
Specimen | Conc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES |
Specimen support | Details: unspecified |
Vitrification | Instrument: FEI VITROBOT MARK III / Cryogen name: ETHANE / Humidity: 100 % |
-Electron microscopy imaging
Experimental equipment | Model: Titan Krios / Image courtesy: FEI Company |
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Microscopy | Model: TFS KRIOS |
Electron gun | Electron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM |
Electron lens | Mode: BRIGHT FIELD / Cs: 2.7 mm / C2 aperture diameter: 70 µm |
Specimen holder | Cryogen: NITROGEN |
Image recording | Electron dose: 65 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) / Num. of real images: 15075 |
Image scans | Movie frames/image: 40 |
-Processing
Software | Name: PHENIX / Version: 1.15.2_3472: / 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: 815542 | ||||||||||||||||||||||||
3D reconstruction | Resolution: 4.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 36691 / Symmetry type: POINT | ||||||||||||||||||||||||
Refine LS restraints |
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