+Open data
-Basic information
Entry | Database: PDB / ID: 7kpx | ||||||
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Title | Structure of the yeast CKM | ||||||
Components |
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Keywords | TRANSCRIPTION / Mediator / Kinase / Cdk8 / Med13 / Med12 / CycC / CDK / Argonaute / RNA Polymerase II / PIWI. | ||||||
Function / homology | Function and homology information negative regulation of filamentous growth / positive regulation of transcription by galactose / CKM complex / nuclear-transcribed mRNA catabolic process, non-stop decay / mediator complex / positive regulation of transcription from RNA polymerase II promoter by galactose / [RNA-polymerase]-subunit kinase / cyclin-dependent protein serine/threonine kinase regulator activity / RNA polymerase II core promoter sequence-specific DNA binding / cyclin-dependent kinase ...negative regulation of filamentous growth / positive regulation of transcription by galactose / CKM complex / nuclear-transcribed mRNA catabolic process, non-stop decay / mediator complex / positive regulation of transcription from RNA polymerase II promoter by galactose / [RNA-polymerase]-subunit kinase / cyclin-dependent protein serine/threonine kinase regulator activity / RNA polymerase II core promoter sequence-specific DNA binding / cyclin-dependent kinase / cyclin-dependent protein serine/threonine kinase activity / RNA polymerase II CTD heptapeptide repeat kinase activity / meiotic cell cycle / protein destabilization / transcription coactivator activity / protein kinase activity / phosphorylation / protein serine kinase activity / negative regulation of transcription by RNA polymerase II / positive regulation of transcription by RNA polymerase II / ATP binding / nucleus / metal ion binding Similarity search - Function | ||||||
Biological species | Saccharomyces cerevisiae (brewer's yeast) | ||||||
Method | ELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.4 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. | ||||||
Funding support | United States, 1items
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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 | 7kpx.cif.gz | 505.6 KB | Display | PDBx/mmCIF format |
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PDB format | pdb7kpx.ent.gz | 384.3 KB | Display | PDB format |
PDBx/mmJSON format | 7kpx.json.gz | Tree view | PDBx/mmJSON format | |
Others | Other downloads |
-Validation report
Summary document | 7kpx_validation.pdf.gz | 1.4 MB | Display | wwPDB validaton report |
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Full document | 7kpx_full_validation.pdf.gz | 1.4 MB | Display | |
Data in XML | 7kpx_validation.xml.gz | 85.7 KB | Display | |
Data in CIF | 7kpx_validation.cif.gz | 130.9 KB | Display | |
Arichive directory | https://data.pdbj.org/pub/pdb/validation_reports/kp/7kpx ftp://data.pdbj.org/pub/pdb/validation_reports/kp/7kpx | HTTPS FTP |
-Related structure data
Related structure data | 22991MC 7kpvC 7kpwC 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: 62940.906 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c References: UniProt: P39073, cyclin-dependent kinase, [RNA-polymerase]-subunit kinase |
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#2: Protein | Mass: 41136.512 Da / Num. of mol.: 1 / Source method: isolated from a natural source Details: Authors introduced a tag to the C-terminus of the chain B protein in the yeast genome. So the promoter of the chain B is still its endogenous promoter. They did not use a plasmid for protein expression Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P47821 |
#3: Protein | Mass: 167049.812 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P25648 |
#4: Protein | Mass: 160175.156 Da / Num. of mol.: 1 / Source method: isolated from a natural source Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast) Strain: ATCC 204508 / S288c / References: UniProt: P38931 |
-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 |
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 |
Image recording | Electron dose: 65 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 QUANTUM (4k x 4k) |
-Processing
EM software |
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CTF correction | Type: PHASE FLIPPING AND AMPLITUDE CORRECTION | |||||||||
Particle selection | Num. of particles selected: 815542 | |||||||||
3D reconstruction | Resolution: 4.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 138178 / Symmetry type: POINT |