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- EMDB-1753: YEAST RNA POLYMERASE III -

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Basic information

Entry
Database: EMDB / ID: EMD-1753
TitleYEAST RNA POLYMERASE III
Map dataRNA-Polymerase III in its free form
Sample
  • Sample: RNA polymerase III
  • Protein or peptide: RNA polymerase III
KeywordsTranscription / RNA polymerase / tRNA
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 21.0 Å
AuthorsVannini A / Ringel R / Kusser AG / Berninghausen O / Kassavetis GA / Cramer P
CitationJournal: Cell / Year: 2010
Title: Molecular basis of RNA polymerase III transcription repression by Maf1.
Authors: Alessandro Vannini / Rieke Ringel / Anselm G Kusser / Otto Berninghausen / George A Kassavetis / Patrick Cramer /
Abstract: RNA polymerase III (Pol III) transcribes short RNAs required for cell growth. Under stress conditions, the conserved protein Maf1 rapidly represses Pol III transcription. We report the crystal ...RNA polymerase III (Pol III) transcribes short RNAs required for cell growth. Under stress conditions, the conserved protein Maf1 rapidly represses Pol III transcription. We report the crystal structure of Maf1 and cryo-electron microscopic structures of Pol III, an active Pol III-DNA-RNA complex, and a repressive Pol III-Maf1 complex. Binding of DNA and RNA causes ordering of the Pol III-specific subcomplex C82/34/31 that is required for transcription initiation. Maf1 binds the Pol III clamp and rearranges C82/34/31 at the rim of the active center cleft. This impairs recruitment of Pol III to a complex of promoter DNA with the initiation factors Brf1 and TBP and thus prevents closed complex formation. Maf1 does however not impair binding of a DNA-RNA scaffold and RNA synthesis. These results explain how Maf1 specifically represses transcription initiation from Pol III promoters and indicate that Maf1 also prevents reinitiation by binding Pol III during transcription elongation.
History
DepositionJun 30, 2010-
Header (metadata) releaseJul 23, 2010-
Map releaseOct 8, 2010-
UpdateDec 11, 2013-
Current statusDec 11, 2013Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0035
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.0035
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

1753.tif

Downloads & links

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Map

FileDownload / File: emd_1753.map.gz / Format: CCP4 / Size: 2.7 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationRNA-Polymerase III in its free form
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
3.31 Å/pix.
x 90 pix.
= 297.9 Å		3.31 Å/pix.
x 90 pix.
= 297.9 Å		3.31 Å/pix.
x 90 pix.
= 297.9 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 3.31 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0035 / Movie #1: 0.0035
Minimum - Maximum-0.00569051 - 0.253301
Average (Standard dev.)0.00280515 (±0.0138798)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions909090
Spacing909090
CellA=B=C: 297.9 Å
α=β=γ: 90 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.313.313.31
M x/y/z909090
origin x/y/z0.0000.0000.000
length x/y/z297.900297.900297.900
α/β/γ90.00090.00090.000
start NX/NY/NZ-100-100-100
NX/NY/NZ200200200
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS909090
D min/max/mean-0.0060.2530.003

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Supplemental data

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Sample components

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Entire : RNA polymerase III

EntireName: RNA polymerase III
Components
  • Sample: RNA polymerase III
  • Protein or peptide: RNA polymerase III

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Supramolecule #1000: RNA polymerase III

SupramoleculeName: RNA polymerase III / type: sample / ID: 1000 / Details: The sample was monodisperse / Oligomeric state: Monomeric / Number unique components: 1
Molecular weightExperimental: 700 KDa / Theoretical: 700 KDa / Method: Native Mass Spectrometry

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Macromolecule #1: RNA polymerase III

MacromoleculeName: RNA polymerase III / type: protein_or_peptide / ID: 1 / Name.synonym: RNA polymerase III / Number of copies: 1 / Oligomeric state: Monomeric / Recombinant expression: Yes
Source (natural)Organism: Saccharomyces cerevisiae (brewer's yeast) / Strain: NZ-16 / synonym: Baker's Yeast / Location in cell: Nucleus
Molecular weightExperimental: 700 KDa / Theoretical: 700 KDa
Recombinant expressionOrganism: Saccharomyces cerevisiae (brewer's yeast)

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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

Concentration0.1 mg/mL
BufferpH: 7.8
Details: 20mM Hepes pH 7.8, 50 mM Ammonium Sulfate, 0.1 mM MgCl2, 0.01 mM ZnCl2, 5mM DTT
GridDetails: Quantifoil R3/3 2nm carbon pre-coated holey grids
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 184 K / Instrument: OTHER / Details: Vitrification instrument: Vitrobot, FEI

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Electron microscopy

MicroscopeFEI TECNAI F20
Electron beamAcceleration voltage: 120 kV / Electron source: LAB6
Electron opticsIllumination mode: OTHER / Imaging mode: OTHER
Sample stageSpecimen holder: Eucentric / Specimen holder model: GATAN LIQUID NITROGEN
Experimental equipment
Model: Tecnai F20 / Image courtesy: FEI Company

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Image processing

Final reconstructionApplied symmetry - Point group: C1 (asymmetric) / Resolution.type: BY AUTHOR / Resolution: 21.0 Å / Resolution method: FSC 0.5 CUT-OFF / Number images used: 20480

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