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- EMDB-7534: Yeast RNA polymerase III intermediate open complex (iOC) -

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Basic information

Entry
Database: EMDB / ID: EMD-7534
TitleYeast RNA polymerase III intermediate open complex (iOC)
Map dataprimary map
Sample
  • Complex: Yeast RNA polymerase III intermediate open complex (iOC)
Biological speciesSaccharomyces cerevisiae S288c (yeast)
Methodsingle particle reconstruction / cryo EM / Resolution: 8.5 Å
AuthorsHan Y / He Y
Funding support United States, 6 items
OrganizationGrant numberCountry
Northwestern UniversityCornew Innovation Award United States
American Cancer SocietyIRG-15-173-21 United States
Chicago Biomedical Consortium with support from the Searle Funds at The Chicago Community TrustCatalyst Award United States
National Science Foundation XSEDE programCHE110042 United States
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM110387 United States
National Energy Research Scientific Computing CenterDE-AC02-05CH11231 United States
CitationJournal: Cell Discov / Year: 2018
Title: Structural visualization of RNA polymerase III transcription machineries.
Authors: Yan Han / Chunli Yan / Susan Fishbain / Ivaylo Ivanov / Yuan He /
Abstract: RNA polymerase III (Pol III) transcription initiation requires the action of the transcription factor IIIB (TFIIIB) and is highly regulated. Here, we determine the structures of Pol III pre- ...RNA polymerase III (Pol III) transcription initiation requires the action of the transcription factor IIIB (TFIIIB) and is highly regulated. Here, we determine the structures of Pol III pre-initiation complexes (PICs) using single particle cryo-electron microscopy (cryo-EM). We observe stable Pol III-TFIIIB complexes using nucleic acid scaffolds mimicking various functional states, in which TFIIIB tightly encircles the upstream promoter DNA. There is an intricate interaction between TFIIIB and Pol III, which stabilizes the winged-helix domains of the C34 subunit of Pol III over the active site cleft. The architecture of Pol III PIC more resembles that of the Pol II PIC than the Pol I PIC. In addition, we also obtain a 3D reconstruction of Pol III in complex with TFIIIB using the elongation complex (EC) scaffold, shedding light on the mechanism of facilitated recycling of Pol III prior to transcription re-initiation.
History
DepositionMar 8, 2018-
Header (metadata) releaseMar 21, 2018-
Map releaseAug 22, 2018-
UpdateApr 1, 2020-
Current statusApr 1, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.09
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.09
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_7534.png

Downloads & links

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Map

FileDownload / File: emd_7534.map.gz / Format: CCP4 / Size: 11.4 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Annotationprimary map
Voxel sizeX=Y=Z: 2.23 Å
Density
Contour LevelBy AUTHOR: 0.09 / Movie #1: 0.09
Minimum - Maximum-0.12150958 - 0.32923743
Average (Standard dev.)0.0019044195 (±0.021076402)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions144144144
Spacing144144144
CellA=B=C: 321.12 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z2.232.232.23
M x/y/z144144144
origin x/y/z0.0000.0000.000
length x/y/z321.120321.120321.120
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ281156
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS144144144
D min/max/mean-0.1220.3290.002

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Supplemental data

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Sample components

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Entire : Yeast RNA polymerase III intermediate open complex (iOC)

EntireName: Yeast RNA polymerase III intermediate open complex (iOC)
Components
  • Complex: Yeast RNA polymerase III intermediate open complex (iOC)

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Supramolecule #1: Yeast RNA polymerase III intermediate open complex (iOC)

SupramoleculeName: Yeast RNA polymerase III intermediate open complex (iOC)
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#21
Source (natural)Organism: Saccharomyces cerevisiae S288c (yeast)
Molecular weightTheoretical: 900 KDa

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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

BufferpH: 7.9
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV

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Electron microscopy

MicroscopeJEOL 3200FS
Electron beamAcceleration voltage: 200 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Average electron dose: 77.2 e/Å2

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Image processing

CTF correctionDetails: CTF amplitude correction was performed following 3D auto refinement in relion.
Startup modelType of model: EMDB MAP
EMDB ID:

3178

Initial angle assignmentType: RANDOM ASSIGNMENT / Software - Name: RELION (ver. 2.1)
Final angle assignmentType: PROJECTION MATCHING / Software - Name: RELION (ver. 2.1)
Final reconstructionResolution.type: BY AUTHOR / Resolution: 8.5 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number images used: 34991
FSC plot (resolution estimation)

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