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- PDB-5u0p: Cryo-EM structure of the transcriptional Mediator -

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Basic information

Entry
Database: PDB / ID: 5u0p
TitleCryo-EM structure of the transcriptional Mediator
Components(Mediator complex subunit ...Mediator (coactivator)) x 16
KeywordsTRANSCRIPTION / transcription / transcriptional / Mediator / Head / Middle / Tail / Srb / RNA polymerase II / Med
Function / homologyMediator complex, subunit Med27 / SOH1 / Med6 core domain superfamily / MED22 superfamily / Mediator complex, subunit Med31 domain superfamily / Mediator complex, subunit Med7/Med21-like / Mediator complex, subunit Med21 / Mediator complex, subunit Med8, fungi / Mediator complex, subunit Med8, fungi/metazoa / Mediator complex, subunit Med17 ...Mediator complex, subunit Med27 / SOH1 / Med6 core domain superfamily / MED22 superfamily / Mediator complex, subunit Med31 domain superfamily / Mediator complex, subunit Med7/Med21-like / Mediator complex, subunit Med21 / Mediator complex, subunit Med8, fungi / Mediator complex, subunit Med8, fungi/metazoa / Mediator complex, subunit Med17 / Mediator complex, subunit Med18 / Mediator complex, subunit Med6, fungi / Mediator complex, subunit Med14 / Mediator complex, subunit Med20 / Mediator of RNA polymerase II transcription subunit 22 / Mediator complex, subunit Med7 / Mediator complex, subunit Med31 / Mediator complex, subunit Med6 / Mediator complex subunit MED14 / Mediator complex subunit 27 / Subunit 21 of Mediator complex / Mediator of RNA polymerase II transcription complex subunit 8 / Subunit 17 of Mediator complex / Med18 protein / TATA-binding related factor (TRF) of subunit 20 of Mediator complex / Surfeit locus protein 5 subunit 22 of Mediator complex / MED7 protein / MED6 mediator sub complex component / RNA polymerase II transcription coactivator activity involved in preinitiation complex assembly / chromatin silencing at centromere outer repeat region / chromosome, centromeric outer repeat region / core mediator complex / mediator complex / RNA polymerase II transcriptional preinitiation complex assembly / go:0001104: / termination of RNA polymerase II transcription / cellular protein localization / activation of protein kinase activity / transcription by RNA polymerase II / transcription coactivator activity / regulation of transcription by RNA polymerase II / transcription, DNA-templated / nucleus / cytosol / Mediator of RNA polymerase II transcription subunit 20 / Mediator of RNA polymerase II transcription subunit 6 / Mediator of RNA polymerase II transcription subunit 14 / Mediator of RNA polymerase II transcription subunit 11 / Mediator of RNA polymerase II transcription subunit 27 / Mediator of RNA polymerase II transcription subunit 18 / Mediator of RNA polymerase II transcription subunit 17 / Mediator of RNA polymerase II transcription subunit 8 / Mediator of RNA polymerase II transcription subunit 21 / Mediator of RNA polymerase II transcription subunit 7 / Mediator of RNA polymerase II transcription subunit 22 / Uncharacterized protein C24C9.04 / Mediator of RNA polymerase II transcription subunit 31
Function and homology information
Specimen sourceSchizosaccharomyces pombe (fission yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 4.4 Å resolution
AuthorsTsai, K.-L. / Yu, X. / Gopalan, S. / Chao, T.-C. / Zhang, Y. / Florens, L. / Washburn, M.P. / Murakami, K. / Conaway, R.C. / Conaway, J.W. / Asturias, F.
CitationJournal: Nature / Year: 2017
Title: Mediator structure and rearrangements required for holoenzyme formation.
Authors: Kuang-Lei Tsai / Xiaodi Yu / Sneha Gopalan / Ti-Chun Chao / Ying Zhang / Laurence Florens / Michael P Washburn / Kenji Murakami / Ronald C Conaway / Joan W Conaway / Francisco J Asturias
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Nov 26, 2016 / Release: Mar 8, 2017
RevisionDateData content typeGroupProviderType
1.0Mar 8, 2017Structure modelrepositoryInitial release
1.1Mar 15, 2017Structure modelDatabase references
1.2Apr 19, 2017Structure modelDatabase references

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Structure visualization

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Assembly

Deposited unit
N: Mediator complex subunit 14
F: Mediator complex subunit 6
H: Mediator complex subunit 8
Q: Mediator complex subunit 17
R: Mediator complex subunit 18
T: Mediator complex subunit 20
K: Mediator complex subunit 11
V: Mediator complex subunit 22
D: Mediator complex subunit 4
G: Mediator complex subunit 7
U: Mediator complex subunit 21
3: Mediator complex subunit 31
I: Mediator complex subunit 9
2: Mediator complex subunit 27
S: Mediator complex subunit 19
J: Mediator complex subunit 10


Theoretical massNumber of molelcules
Total (without water)455,87816
Polyers455,87816
Non-polymers00
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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Mediator complex subunit ... , 16 types, 16 molecules NFHQRTKVDGU3I2SJ

#1: Protein/peptide Mediator complex subunit 14 / Mediator (coactivator)


Mass: 105382.461 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q9P7Y4
#2: Protein/peptide Mediator complex subunit 6 / Mediator (coactivator)


Mass: 24928.947 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q9US45
#3: Protein/peptide Mediator complex subunit 8 / Mediator (coactivator)


Mass: 23360.068 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O94646
#4: Protein/peptide Mediator complex subunit 17 / Mediator (coactivator)


Mass: 62551.410 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: P87306
#5: Protein/peptide Mediator complex subunit 18 / Mediator (coactivator)


Mass: 24056.377 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O14198
#6: Protein/peptide Mediator complex subunit 20 / Mediator (coactivator)


Mass: 22374.775 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q10317
#7: Protein/peptide Mediator complex subunit 11 / Mediator (coactivator)


Mass: 13118.800 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q9P6Q0
#8: Protein/peptide Mediator complex subunit 22 / Mediator (coactivator)


Mass: 15396.165 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O14010
#9: Protein/peptide Mediator complex subunit 4 / Mediator (coactivator)


Mass: 20358.010 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast)
#10: Protein/peptide Mediator complex subunit 7 / Mediator (coactivator)


Mass: 43573.094 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O60104
#11: Protein/peptide Mediator complex subunit 21 / Mediator (coactivator)


Mass: 15821.031 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O94376
#12: Protein/peptide Mediator complex subunit 31 / Mediator (coactivator)


Mass: 16913.975 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q9USH1
#13: Protein/peptide Mediator complex subunit 9 / Mediator (coactivator)


Mass: 13810.964 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: O13964
#14: Protein/peptide Mediator complex subunit 27 / Mediator (coactivator)


Mass: 31132.693 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast) / References: UniProt: Q10477
#15: Protein/peptide Mediator complex subunit 19 / Mediator (coactivator)


Mass: 11847.596 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast)
#16: Protein/peptide Mediator complex subunit 10 / Mediator (coactivator)


Mass: 11251.861 Da / Num. of mol.: 1 / Source: (natural) Schizosaccharomyces pombe (fission yeast)

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / Reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Mediator complexMediator (coactivator) / Type: COMPLEX / Entity ID: 1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16 / Source: NATURAL
Source (natural)Organism: Schizosaccharomyces pombe (fission yeast)
Buffer solutionDetails: 20 mM HEPES, pH 7.4, 200 mM potassium acetate, 5 mM b-Mercaptoethanol, 0.01% NP-40
pH: 7.4
SpecimenConc.: 1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyMicroscope model: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELDBright-field microscopy / Nominal magnification: 22500 / Nominal defocus max: 4000 nm / Nominal defocus min: 1000 nm / Cs: 2.7 mm / C2 aperture diameter: 100 mm
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 7 sec. / Electron dose: 9.6 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Number of real images: 11181

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Processing

EM software
IDNameVersionCategory
1APPIONparticle selection
2LEGINON3.0image acquisition
4CTFFIND3CTF correction
10RELION1.4initial Euler assignment
11RELION1.4final Euler assignment
13RELION1.43D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1
3D reconstructionResolution: 4.4 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 42484 / Symmetry type: POINT

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