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- PDB-9gck: yeast TFIIIC TauA subcomplex bound to a tRNA gene -

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Basic information

Entry
Database: PDB / ID: 9gck
Titleyeast TFIIIC TauA subcomplex bound to a tRNA gene
Components
  • (DNA (45-MER)) x 2
  • (Transcription factor tau ...) x 4
KeywordsDNA BINDING PROTEIN / A-box / tRNA gene
Function / homology
Function and homology information


5S class rRNA transcription by RNA polymerase III / transcription factor TFIIIC complex / RNA Polymerase III Transcription Initiation From Type 2 Promoter / transcription initiation at RNA polymerase III promoter / phosphatase activity / transcription by RNA polymerase III / protein localization to chromatin / mitochondrion / DNA binding / nucleoplasm ...5S class rRNA transcription by RNA polymerase III / transcription factor TFIIIC complex / RNA Polymerase III Transcription Initiation From Type 2 Promoter / transcription initiation at RNA polymerase III promoter / phosphatase activity / transcription by RNA polymerase III / protein localization to chromatin / mitochondrion / DNA binding / nucleoplasm / nucleus / cytoplasm
Similarity search - Function
Transcription factor tau subunit sfc3/Tfc3, C-terminal / : / Family of unknown function (DUF6581) / Transcription factor tau 138 kDa subunit, extended winged helix / Transcription factor TFIIIC subunit Tfc7/tau55 / Transcription factor IIIC subunit 5, HTH domain / Transcription factor TFIIIC, triple barrel domain / Transcription factor Tfc4/TFIIIC-102/Sfc4 / Transcription factor IIIC subunit Tfc1/Sfc1 / Transcription factor IIIC subunit Tfc1/Sfc1, triple barrel domain ...Transcription factor tau subunit sfc3/Tfc3, C-terminal / : / Family of unknown function (DUF6581) / Transcription factor tau 138 kDa subunit, extended winged helix / Transcription factor TFIIIC subunit Tfc7/tau55 / Transcription factor IIIC subunit 5, HTH domain / Transcription factor TFIIIC, triple barrel domain / Transcription factor Tfc4/TFIIIC-102/Sfc4 / Transcription factor IIIC subunit Tfc1/Sfc1 / Transcription factor IIIC subunit Tfc1/Sfc1, triple barrel domain / TFIIIC, subcomplex tauA subunit Sfc1, triple barrel domain superfamily / RNA polymerase III transcription factor (TF)IIIC subunit HTH domain / TFIIIC subunit triple barrel domain / Tau95 Triple barrel domain / B-block binding subunit of TFIIIC / Tfc3, extended winged-helix domain / Transcription facto Tfc3-like / B-block binding subunit of TFIIIC / : / Phosphoglycerate mutase family / Histidine phosphatase superfamily, clade-1 / Histidine phosphatase superfamily (branch 1) / Histidine phosphatase superfamily / Tetratricopeptide repeat / Tetratricopeptide repeat / TPR repeat region circular profile. / TPR repeat profile. / Tetratricopeptide repeats / Tetratricopeptide repeat / Tetratricopeptide-like helical domain superfamily
Similarity search - Domain/homology
DNA / DNA (> 10) / Transcription factor tau 95 kDa subunit / Transcription factor tau 131 kDa subunit / Transcription factor tau 138 kDa subunit / Transcription factor tau 55 kDa subunit
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsSeifert-Davila, W. / Girbig, M. / Hauptmann, L. / Hoffmann, T. / Eustermann, S. / Mueller, C. / Chaban, A. / Duss, O. / Baudin, F.
Funding support1items
OrganizationGrant numberCountry
Not funded
CitationJournal: Nucleic Acids Res / Year: 2024
Title: Structural and kinetic insights into tRNA promoter engagement by yeast general transcription factor TFIIIC.
Authors: Wolfram Seifert-Dávila / Anastasiia Chaban / Florence Baudin / Mathias Girbig / Luis Hauptmann / Thomas Hoffmann / Olivier Duss / Sebastian Eustermann / Christoph W Müller /
Abstract: Transcription of transfer RNA (tRNA) genes by RNA polymerase (Pol) III requires the general transcription factor IIIC (TFIIIC), which recognizes intragenic A-box and B-box DNA motifs of type II ...Transcription of transfer RNA (tRNA) genes by RNA polymerase (Pol) III requires the general transcription factor IIIC (TFIIIC), which recognizes intragenic A-box and B-box DNA motifs of type II gene promoters. However, the underlying mechanism has remained elusive, in part due to missing structural information for A-box recognition. In this study, we use single-particle cryogenic electron microscopy (cryo-EM) and single-molecule fluorescence resonance energy transfer (smFRET) to reveal structural and real-time kinetic insights into how the 520-kDa yeast TFIIIC complex engages A-box and B-box DNA motifs in the context of a tRNA gene promoter. Cryo-EM structures of τA and τB subcomplexes bound to the A-box and B-box were obtained at 3.7 and 2.5 Å resolution, respectively, while cryo-EM single-particle mapping determined the specific distance and relative orientation of the τA and τB subcomplexes revealing a fully engaged state of TFIIIC. smFRET experiments show that overall recruitment and residence times of TFIIIC on a tRNA gene are primarily governed by B-box recognition, while footprinting experiments suggest a key role of τA and the A-box in TFIIIB and Pol III recruitment following TFIIIC recognition of type II promoters.
History
DepositionAug 2, 2024Deposition site: PDBE / Processing site: PDBE
Revision 1.0Nov 27, 2024Provider: repository / Type: Initial release
Revision 1.1Dec 25, 2024Group: Data collection / Database references / Category: citation / citation_author / em_admin
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_ASTM / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year / _citation_author.identifier_ORCID / _citation_author.name / _em_admin.last_update

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Transcription factor tau 138 kDa subunit
B: Transcription factor tau 131 kDa subunit
C: Transcription factor tau 95 kDa subunit
D: Transcription factor tau 55 kDa subunit
E: DNA (45-MER)
F: DNA (45-MER)


Theoretical massNumber of molelcules
Total (without water)403,3086
Polymers403,3086
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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Transcription factor tau ... , 4 types, 4 molecules ABCD

#1: Protein Transcription factor tau 138 kDa subunit / TFIIIC 138 kDa subunit / Transcription factor C subunit 3


Mass: 136602.500 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: TFC3, TSV115, YAL001C, FUN24 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P34111
#2: Protein Transcription factor tau 131 kDa subunit / TFIIIC 131 kDa subunit / Transcription factor C subunit 4


Mass: 120746.969 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: TFC4, PCF1, YGR047C / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P33339
#3: Protein Transcription factor tau 95 kDa subunit / TFIIIC 95 kDa subunit / Transcription factor C subunit 1


Mass: 69066.438 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: TFC1, YBR123C, YBR0919 / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: P32367
#4: Protein Transcription factor tau 55 kDa subunit / TFIIIC 55 kDa subunit / Transcription factor C subunit 7


Mass: 49198.898 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (brewer's yeast)
Gene: TFC7, YOR110W, O3234, YOR3234w / Production host: Spodoptera frugiperda (fall armyworm) / References: UniProt: Q12415

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DNA chain , 2 types, 2 molecules EF

#5: DNA chain DNA (45-MER)


Mass: 13643.719 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Saccharomyces cerevisiae (brewer's yeast)
#6: DNA chain DNA (45-MER)


Mass: 14049.344 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Saccharomyces cerevisiae (brewer's yeast)

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Details

Has protein modificationN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

Component
IDNameTypeEntity IDParent-IDSource
1Yeast TFIIIC TauA monomer bound to a tRNA geneCOMPLEXall0MULTIPLE SOURCES
2Transcription factor tauCOMPLEX#1-#41RECOMBINANT
3DNACOMPLEX#5-#61RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Saccharomyces cerevisiae (brewer's yeast)4932
33Saccharomyces cerevisiae (brewer's yeast)4932
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
22Spodoptera frugiperda (fall armyworm)7108
33synthetic construct (others)32630
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1700 nm / Nominal defocus min: 700 nm
Image recordingElectron dose: 39.6 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)
Details: dataset1: 39.6 dataset2: 43.6 dataset3: 43.2 dataset4: 43.2 dataset5: 44.4

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 114621 / Symmetry type: POINT
RefinementResolution: 3.7→3.7 Å / Cor.coef. Fo:Fc: 0.926 / SU B: 57.24 / SU ML: 0.786 / ESU R: 0.803
Stereochemistry target values: MAXIMUM LIKELIHOOD WITH PHASES
Details: HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS
RfactorNum. reflection% reflection
Rwork0.46105 --
obs0.46105 98925 100 %
Solvent computationSolvent model: PARAMETERS FOR MASK CACLULATION
Displacement parametersBiso mean: 247.984 Å2
Refinement stepCycle: 1 / Total: 12301
Refine LS restraints
Refine-IDTypeDev idealDev ideal targetNumber
ELECTRON MICROSCOPYr_bond_refined_d0.0080.01312752
ELECTRON MICROSCOPYr_bond_other_d0.0030.01611267
ELECTRON MICROSCOPYr_angle_refined_deg1.8281.65817638
ELECTRON MICROSCOPYr_angle_other_deg1.3011.57926046
ELECTRON MICROSCOPYr_dihedral_angle_1_deg9.6251280
ELECTRON MICROSCOPYr_dihedral_angle_2_deg33.40122.434567
ELECTRON MICROSCOPYr_dihedral_angle_3_deg13.521151966
ELECTRON MICROSCOPYr_dihedral_angle_4_deg17.2761569
ELECTRON MICROSCOPYr_chiral_restr0.1130.21738
ELECTRON MICROSCOPYr_gen_planes_refined0.0090.0212936
ELECTRON MICROSCOPYr_gen_planes_other0.0030.022720
ELECTRON MICROSCOPYr_nbd_refined
ELECTRON MICROSCOPYr_nbd_other
ELECTRON MICROSCOPYr_nbtor_refined
ELECTRON MICROSCOPYr_nbtor_other
ELECTRON MICROSCOPYr_xyhbond_nbd_refined
ELECTRON MICROSCOPYr_xyhbond_nbd_other
ELECTRON MICROSCOPYr_metal_ion_refined
ELECTRON MICROSCOPYr_metal_ion_other
ELECTRON MICROSCOPYr_symmetry_vdw_refined
ELECTRON MICROSCOPYr_symmetry_vdw_other
ELECTRON MICROSCOPYr_symmetry_hbond_refined
ELECTRON MICROSCOPYr_symmetry_hbond_other
ELECTRON MICROSCOPYr_symmetry_metal_ion_refined
ELECTRON MICROSCOPYr_symmetry_metal_ion_other
ELECTRON MICROSCOPYr_mcbond_it24.7223.6055141
ELECTRON MICROSCOPYr_mcbond_other24.71523.6035140
ELECTRON MICROSCOPYr_mcangle_it40.04435.256414
ELECTRON MICROSCOPYr_mcangle_other40.04535.2516415
ELECTRON MICROSCOPYr_scbond_it20.0729.3777611
ELECTRON MICROSCOPYr_scbond_other20.0729.3777611
ELECTRON MICROSCOPYr_scangle_it
ELECTRON MICROSCOPYr_scangle_other34.14143.89711225
ELECTRON MICROSCOPYr_long_range_B_refined62.87451008
ELECTRON MICROSCOPYr_long_range_B_other62.87351009
ELECTRON MICROSCOPYr_rigid_bond_restr
ELECTRON MICROSCOPYr_sphericity_free
ELECTRON MICROSCOPYr_sphericity_bonded
LS refinement shellResolution: 3.5→3.591 Å / Total num. of bins used: 20
RfactorNum. reflection% reflection
Rfree0 0 -
Rwork1.435 7355 -
obs--100 %

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