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- EMDB-2634: Electron cryo-microscopy of yeast transcriptional Mediator -

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Basic information

Entry
Database: EMDB / ID: EMD-2634
TitleElectron cryo-microscopy of yeast transcriptional Mediator
Map dataReconstruction of yeast transcriptional Mediator
Sample
  • Sample: yeast transcriptional Mediator
  • Protein or peptide: Mediator
Keywordstranscription / mediator / Srb / RNAPII / regulation / activator
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 25.0 Å
AuthorsTsai KL / Tomomori-Sato C / Sato S / Conaway RC / Conaway JW / Asturias FJ
CitationJournal: Cell / Year: 2014
Title: Subunit architecture and functional modular rearrangements of the transcriptional mediator complex.
Authors: Kuang-Lei Tsai / Chieri Tomomori-Sato / Shigeo Sato / Ronald C Conaway / Joan W Conaway / Francisco J Asturias /
Abstract: The multisubunit Mediator, comprising ∼30 distinct proteins, plays an essential role in gene expression regulation by acting as a bridge between DNA-binding transcription factors and the RNA ...The multisubunit Mediator, comprising ∼30 distinct proteins, plays an essential role in gene expression regulation by acting as a bridge between DNA-binding transcription factors and the RNA polymerase II (RNAPII) transcription machinery. Efforts to uncover the Mediator mechanism have been hindered by a poor understanding of its structure, subunit organization, and conformational rearrangements. By overcoming biochemical and image analysis hurdles, we obtained accurate EM structures of yeast and human Mediators. Subunit localization experiments, docking of partial X-ray structures, and biochemical analyses resulted in comprehensive mapping of yeast Mediator subunits and a complete reinterpretation of our previous Mediator organization model. Large-scale Mediator rearrangements depend on changes at the interfaces between previously described Mediator modules, which appear to be facilitated by factors conducive to transcription initiation. Conservation across eukaryotes of Mediator structure, subunit organization, and RNA polymerase II interaction suggest conservation of fundamental aspects of the Mediator mechanism.
History
DepositionApr 23, 2014-
Header (metadata) releaseMay 28, 2014-
Map releaseMay 28, 2014-
UpdateJun 18, 2014-
Current statusJun 18, 2014Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.5
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.5
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

EMD-2634_yME...

Downloads & links

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Map

FileDownload / File: emd_2634.map.gz / Format: CCP4 / Size: 4.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationReconstruction of yeast transcriptional Mediator
Voxel sizeX=Y=Z: 4.84 Å
Density
Contour LevelBy EMDB: 0.35 / Movie #1: 0.5
Minimum - Maximum-0.0000004 - 4.56570101
Average (Standard dev.)0.02234847 (±0.16090581)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions108108108
Spacing108108108
CellA=B=C: 522.72003 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z4.844.844.84
M x/y/z108108108
origin x/y/z0.0000.0000.000
length x/y/z522.720522.720522.720
α/β/γ90.00090.00090.000
start NX/NY/NZ-184-184-183
NX/NY/NZ368368368
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS108108108
D min/max/mean-0.0004.5660.022

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Supplemental data

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Sample components

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Entire : yeast transcriptional Mediator

EntireName: yeast transcriptional Mediator
Components
  • Sample: yeast transcriptional Mediator
  • Protein or peptide: Mediator

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Supramolecule #1000: yeast transcriptional Mediator

SupramoleculeName: yeast transcriptional Mediator / type: sample / ID: 1000 / Number unique components: 1
Molecular weightTheoretical: 1 MDa

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Macromolecule #1: Mediator

MacromoleculeName: Mediator / type: protein_or_peptide / ID: 1 / Recombinant expression: No
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: S228C
Molecular weightTheoretical: 1 MDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

Concentration0.03 mg/mL
VitrificationCryogen name: ETHANE / Instrument: HOMEMADE PLUNGER

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Nominal defocus max: 3.7 µm / Nominal defocus min: 0.5 µm
Sample stageSpecimen holder model: GATAN LIQUID NITROGEN
DateAug 18, 2013
Image recordingCategory: CCD / Film or detector model: GATAN K2 (4k x 4k)
Details: Every image is the average of 25 frames recorded by the direct electron detector
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

CTF correctionDetails: Each particle
Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Algorithm: OTHER / Resolution.type: BY AUTHOR / Resolution: 25.0 Å / Resolution method: OTHER / Software - Name: EMAN2/SPARX / Number images used: 13006

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Atomic model buiding 1

Initial modelPDB ID:

4gwq

SoftwareName: Chimera
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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