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- EMDB-0603: Cryo-EM structure of the human TFIIH core complex: Multibody-refi... -

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Basic information

Entry
Database: EMDB / ID: EMD-0603
TitleCryo-EM structure of the human TFIIH core complex: Multibody-refined map, body 2
Map dataMultibody-refined cryo-EM map, body 2 (post-processed).
Sample
  • Complex: Transcription factor IIH (TFIIH)
Biological speciesHomo sapiens (human)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.6 Å
AuthorsGreber BJ / Toso D / Fang J / Nogales E
Funding support United States, Switzerland, 4 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical SciencesR01-GM63072 United States
Swiss National Science FoundationP300PA_160983 Switzerland
National Institutes of Health/National Institute of General Medical SciencesP01-GM063210 United States
Swiss National Science FoundationP300PA_174355 Switzerland
CitationJournal: Elife / Year: 2019
Title: The complete structure of the human TFIIH core complex.
Authors: Basil J Greber / Daniel B Toso / Jie Fang / Eva Nogales /
Abstract: Transcription factor IIH (TFIIH) is a heterodecameric protein complex critical for transcription initiation by RNA polymerase II and nucleotide excision DNA repair. The TFIIH core complex is ...Transcription factor IIH (TFIIH) is a heterodecameric protein complex critical for transcription initiation by RNA polymerase II and nucleotide excision DNA repair. The TFIIH core complex is sufficient for its repair functions and harbors the XPB and XPD DNA-dependent ATPase/helicase subunits, which are affected by human disease mutations. Transcription initiation additionally requires the CdK activating kinase subcomplex. Previous structural work has provided only partial insight into the architecture of TFIIH and its interactions within transcription pre-initiation complexes. Here, we present the complete structure of the human TFIIH core complex, determined by phase-plate cryo-electron microscopy at 3.7 Å resolution. The structure uncovers the molecular basis of TFIIH assembly, revealing how the recruitment of XPB by p52 depends on a pseudo-symmetric dimer of homologous domains in these two proteins. The structure also suggests a function for p62 in the regulation of XPD, and allows the mapping of previously unresolved human disease mutations.
History
Header (metadata) releaseFeb 13, 2019-
DepositionFeb 21, 2019-
Map releaseMar 13, 2019-
UpdateDec 2, 2020-
Current statusDec 2, 2020Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0243
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.0243
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_0603.png

Downloads & links

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Map

FileDownload / File: emd_0603.map.gz / Format: CCP4 / Size: 64 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationMultibody-refined cryo-EM map, body 2 (post-processed).
Voxel sizeX=Y=Z: 1.15 Å
Density
Contour LevelBy AUTHOR: 0.0243 / Movie #1: 0.0243
Minimum - Maximum-0.10053686 - 0.20372707
Average (Standard dev.)-0.0000213628 (±0.0031070653)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions256256256
Spacing256256256
CellA=B=C: 294.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.151.151.15
M x/y/z256256256
origin x/y/z0.0000.0000.000
length x/y/z294.400294.400294.400
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS256256256
D min/max/mean-0.1010.204-0.000

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Supplemental data

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Mask #1

Fileemd_0603_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Unfiltered half-map.

Fileemd_0603_half_map_1.map
AnnotationUnfiltered half-map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Half map: Unfiltered half-map.

Fileemd_0603_half_map_2.map
AnnotationUnfiltered half-map.
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : Transcription factor IIH (TFIIH)

EntireName: Transcription factor IIH (TFIIH)
Components
  • Complex: Transcription factor IIH (TFIIH)

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Supramolecule #1: Transcription factor IIH (TFIIH)

SupramoleculeName: Transcription factor IIH (TFIIH) / type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#8
Source (natural)Organism: Homo sapiens (human)
Molecular weightTheoretical: 500 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

Concentration0.0049 mg/mL
BufferpH: 7.9
Component:
ConcentrationName
20.0 mMHepes-KOH
150.0 mMSodium chloride
5.0 mMMagnesium chloride
0.015 %NP40 substitute
1.5 %Glycerol
2.0 %Trehalose
VitrificationCryogen name: ETHANE / Chamber humidity: 100 % / Chamber temperature: 277 K / Instrument: FEI VITROBOT MARK IV
Details: A thin film of continuous carbon was floated onto Protochips C-flat CF-4/2 holey carbon grids and glow discharged or plasma cleaned before application of 4 uL of sample solution..

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsCalibrated magnification: 43478 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELDBright-field microscopy / Cs: 2.7 mm / Nominal defocus max: 0.7 µm / Nominal defocus min: 0.5 µm
Specialist opticsPhase plate: VOLTA PHASE PLATE / Energy filter - Name: GIF Quantum LS / Energy filter - Slit width: 20 eV
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
DetailsResidual beam tilt corrected in RELION 3.
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Number grids imaged: 7 / Number real images: 21437 / Average exposure time: 8.25 sec. / Average electron dose: 50.0 e/Å2
Details: Images were collected as dose-fractionated movie frames (33 or 50 frames per exposure).
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

CTF correctionSoftware:
Namedetails
GctfCTF determination
RELION (ver. 3)CTF correction

Details: CTF correction during 3D reconstruction in RELION 3.
Startup modelType of model: EMDB MAP
EMDB ID:

3802

Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3)
Final 3D classificationNumber classes: 8 / Avg.num./class: 100000 / Software - Name: RELION (ver. 3)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 3)
Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: FOURIER SPACE / Resolution.type: BY AUTHOR / Resolution: 3.6 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 3) / Details: Multibody-refined in RELION 3. / Number images used: 138659

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Atomic model buiding 1

Initial modelPDB ID:

6nmi

DetailsThis map supported model re-building in O and COOT.
RefinementSpace: REAL / Protocol: RIGID BODY FIT

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