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- PDB-5oqj: STRUCTURE OF YEAST TRANSCRIPTION PRE-INITIATION COMPLEX WITH TFIIH -

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Entry
Database: PDB / ID: 5oqj
TitleSTRUCTURE OF YEAST TRANSCRIPTION PRE-INITIATION COMPLEX WITH TFIIH
Components
  • (DNA repair helicase ...) x 2
  • (DNA-directed RNA polymerase II subunit ...Polymerase) x 7
  • (DNA-directed RNA polymerases I, II, and III subunit ...RNA polymerase) x 5
  • (RNA polymerase II transcription factor B subunit ...) x 5
  • (Transcription initiation factor IIA ...) x 2
  • (Transcription initiation factor IIE subunit ...) x 2
  • (Transcription initiation factor IIF subunit ...) x 2
  • NONTEMPLATE DNA
  • Suppressor of stem-loop protein 1
  • TATA-box-binding protein
  • TEMPLATE DNA
  • Transcription initiation factor IIB
  • Unknown protein
KeywordsTRANSCRIPTION / transcription initiation / macromolecular complex
Function / homology
Function and homology information


: / regulation of mitotic recombination / RNA polymerase II promoter clearance / RNA polymerase II complex recruiting activity / TFIIA-class transcription factor complex binding / transcription factor TFIIIB complex / RNA polymerase III preinitiation complex assembly / RNA polymerase III transcription regulatory region sequence-specific DNA binding / phosphatidylinositol-5-phosphate binding / positive regulation of mitotic recombination ...: / regulation of mitotic recombination / RNA polymerase II promoter clearance / RNA polymerase II complex recruiting activity / TFIIA-class transcription factor complex binding / transcription factor TFIIIB complex / RNA polymerase III preinitiation complex assembly / RNA polymerase III transcription regulatory region sequence-specific DNA binding / phosphatidylinositol-5-phosphate binding / positive regulation of mitotic recombination / regulation of transcription by RNA polymerase III / RNA polymerase I general transcription initiation factor binding / DNA translocase activity / nucleotide-excision repair factor 3 complex / transcription factor TFIIE complex / nucleotide-excision repair, preincision complex assembly / transcription factor TFIIK complex / transcription open complex formation at RNA polymerase II promoter / TFIIF-class transcription factor complex binding / transcriptional start site selection at RNA polymerase II promoter / RPB4-RPB7 complex / positive regulation of transcription regulatory region DNA binding / transcription factor TFIIF complex / phosphatidylinositol-3-phosphate binding / 5'-3' DNA helicase activity / transcription factor TFIIA complex / : / RNA polymerase I preinitiation complex assembly / transcription factor TFIIH holo complex / transcription factor TFIIH core complex / cyclin-dependent protein serine/threonine kinase activator activity / nuclear-transcribed mRNA catabolic process, deadenylation-dependent decay / 3'-5' DNA helicase activity / transcription preinitiation complex / poly(A)+ mRNA export from nucleus / DNA binding, bending / RNA Polymerase I Transcription Initiation / DNA duplex unwinding / : / positive regulation of nuclear-transcribed mRNA poly(A) tail shortening / Processing of Capped Intron-Containing Pre-mRNA / : / RNA Polymerase III Transcription Initiation From Type 2 Promoter / RNA Pol II CTD phosphorylation and interaction with CE / Formation of the Early Elongation Complex / mRNA Capping / termination of RNA polymerase II transcription / Formation of TC-NER Pre-Incision Complex / RNA polymerase II transcribes snRNA genes / RNA Polymerase I Promoter Escape / TP53 Regulates Transcription of DNA Repair Genes / Estrogen-dependent gene expression / RNA Polymerase II Promoter Escape / RNA Polymerase II Transcription Pre-Initiation And Promoter Opening / RNA Polymerase II Transcription Initiation / RNA Polymerase II Transcription Initiation And Promoter Clearance / RNA polymerase II general transcription initiation factor activity / transcription factor TFIID complex / RNA-templated transcription / termination of RNA polymerase III transcription / RNA Polymerase II Pre-transcription Events / maintenance of transcriptional fidelity during transcription elongation by RNA polymerase II / Dual incision in TC-NER / RNA polymerase II complex binding / RNA polymerase I activity / termination of RNA polymerase I transcription / transcription initiation at RNA polymerase III promoter / ATPase activator activity / protein phosphatase activator activity / tRNA transcription by RNA polymerase III / nucleolar large rRNA transcription by RNA polymerase I / Gap-filling DNA repair synthesis and ligation in TC-NER / transcription initiation at RNA polymerase I promoter / transcription elongation by RNA polymerase I / RNA polymerase II activity / positive regulation of translational initiation / transcription-coupled nucleotide-excision repair / positive regulation of transcription initiation by RNA polymerase II / translesion synthesis / RNA polymerase II core promoter sequence-specific DNA binding / RNA polymerase I complex / transcription by RNA polymerase I / RNA polymerase III complex / transcription by RNA polymerase III / ATP-dependent activity, acting on DNA / RNA polymerase II, core complex / translation initiation factor binding / RNA polymerase II preinitiation complex assembly / DNA helicase activity / TBP-class protein binding / transcription elongation by RNA polymerase II / P-body / DNA-templated transcription initiation / transcription initiation at RNA polymerase II promoter / nucleotide-excision repair / positive regulation of transcription elongation by RNA polymerase II / transcription coregulator activity / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase
Similarity search - Function
Bacterial type XPD DNA helicase, FeS cluster domain / TFIIH subunit Tfb4/GTF2H3 / Transcription factor Tfb4 / TFIIH C1-like domain / Ssl1-like / TFIIH subunit Ssl1/p44 / Ssl1-like / TFIIH C1-like domain / TFIIH C1-like domain / TFIIH p62 subunit, N-terminal ...Bacterial type XPD DNA helicase, FeS cluster domain / TFIIH subunit Tfb4/GTF2H3 / Transcription factor Tfb4 / TFIIH C1-like domain / Ssl1-like / TFIIH subunit Ssl1/p44 / Ssl1-like / TFIIH C1-like domain / TFIIH C1-like domain / TFIIH p62 subunit, N-terminal / TFIIH subunit Tfb1/GTF2H1 / TFIIH p62 subunit, N-terminal domain / BSD domain / BSD domain superfamily / BSD domain / BSD domain profile. / domain in transcription factors and synapse-associated proteins / RAD3/XPD family / Helicase XPB/Ssl2 / Helical and beta-bridge domain / ERCC3/RAD25/XPB helicase, C-terminal domain / Helicase XPB/Ssl2, N-terminal domain / Helical and beta-bridge domain / Helicase conserved C-terminal domain / ERCC3/RAD25/XPB C-terminal helicase / Transcription factor TFIIH subunit p52/Tfb2 / ATP-dependent helicase Rad3/Chl1-like / Transcription factor Tfb2, C-terminal domain / Transcription factor Tfb2 / Transcription factor Tfb2 (p52) C-terminal domain / Transcription factor TFIIE beta subunit, DNA-binding domain / Transcription initiation factor TFIIE, beta subunit / TFA2, Winged helix domain 2 / TFIIE beta subunit core domain / TFA2 Winged helix domain 2 / TFIIE beta central core DNA-binding domain profile. / TFIIH subunit TTDA/Tfb5 / TFB5-like superfamily / Transcription factor TFIIH complex subunit Tfb5 / Transcription factor TFIIH complex subunit Tfb5 / Cdk-activating kinase assembly factor MAT1/Tfb3 / Cdk-activating kinase assembly factor MAT1, centre / CDK-activating kinase assembly factor MAT1 / Zinc finger, C3HC4 type (RING finger) / Helicase superfamily 1/2, DinG/Rad3-like / Helicase-like, DEXD box c2 type / ATP-dependent helicase, C-terminal / DEAD2 / Helicase superfamily 1/2, ATP-binding domain, DinG/Rad3-type / DEAD_2 / Helicase C-terminal domain / Superfamilies 1 and 2 helicase ATP-binding type-2 domain profile. / DEXDc2 / HELICc2 / Transcription initiation factor IIE subunit alpha, N-terminal / Transcription factor TFE/TFIIEalpha HTH domain / TFIIEalpha/SarR/Rpc3 HTH domain / Transcription factor E / TFIIE alpha subunit / TFE/IIEalpha-type HTH domain profile. / Transcription initiation factor IIE / Transcription factor IIA, alpha/beta subunit / Transcription factor IIA, alpha/beta subunit / Transcription factor IIA, alpha/beta subunit / Transcription initiation factor IIA, gamma subunit / Transcription factor IIA, alpha-helical domain / Transcription factor IIA, beta-barrel / Transcription initiation factor IIA, gamma subunit, C-terminal / Transcription initiation factor IIA, gamma subunit, N-terminal / Transcription initiation factor IIA, gamma subunit, helical domain / Transcription initiation factor IIA, gamma subunit / Transcription initiation factor IIF, beta subunit / TFIIF beta subunit, HTH domain / TFIIF, beta subunit, N-terminal / TFIIF, beta subunit HTH domain / TFIIF, beta subunit N-terminus / Transcription initiation factor IIF, alpha subunit / Transcription initiation factor IIF, alpha subunit (TFIIF-alpha) / Transcription Factor IIF, Rap30/Rap74, interaction / Transcription factor TFIIB, cyclin-like domain / Transcription factor TFIIB, conserved site / Transcription factor TFIIB repeat / Transcription factor TFIIB repeat signature. / Transcription factor TFIIB / Zinc finger TFIIB-type profile. / Zinc finger, TFIIB-type / TFIIB zinc-binding / TATA-box binding protein, eukaryotic / TATA-box binding protein / TATA-box binding protein, conserved site / Transcription factor TFIID (or TATA-binding protein, TBP) / Transcription factor TFIID repeat signature. / Helicase/UvrB, N-terminal / Type III restriction enzyme, res subunit / DNA/RNA helicase, ATP-dependent, DEAH-box type, conserved site / DEAH-box subfamily ATP-dependent helicases signature. / TBP domain superfamily / DNA-directed RNA polymerase II subunit Rpb4-like / RNA polymerase Rpb4/RPC9, core / DNA-directed RNA-polymerase II subunit
Similarity search - Domain/homology
IRON/SULFUR CLUSTER / DNA / DNA (> 10) / DNA (> 100) / DNA-directed RNA polymerase II subunit RPB1 / General transcription and DNA repair factor IIH helicase subunit XPD / DNA-directed RNA polymerase II subunit RPB2 / TATA-box-binding protein / DNA-directed RNA polymerase II subunit RPB3 / DNA-directed RNA polymerase II subunit RPB4 ...IRON/SULFUR CLUSTER / DNA / DNA (> 10) / DNA (> 100) / DNA-directed RNA polymerase II subunit RPB1 / General transcription and DNA repair factor IIH helicase subunit XPD / DNA-directed RNA polymerase II subunit RPB2 / TATA-box-binding protein / DNA-directed RNA polymerase II subunit RPB3 / DNA-directed RNA polymerase II subunit RPB4 / DNA-directed RNA polymerases I, II, and III subunit RPABC1 / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC3 / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerase II subunit RPB9 / Transcription initiation factor IIB / Transcription initiation factor IIA large subunit / Transcription initiation factor IIA subunit 2 / General transcription and DNA repair factor IIH subunit TFB1 / DNA-directed RNA polymerase II subunit RPB7 / Transcription initiation factor IIE subunit alpha / Transcription initiation factor IIE subunit beta / DNA-directed RNA polymerase II subunit RPB11 / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / Transcription initiation factor IIF subunit alpha / Transcription initiation factor IIF subunit beta / General transcription and DNA repair factor IIH helicase subunit XPB / General transcription and DNA repair factor IIH subunit TFB2 / RNA polymerase II transcription factor B subunit 3 / General transcription and DNA repair factor IIH subunit SSL1 / General transcription and DNA repair factor IIH subunit TFB4 / General transcription and DNA repair factor IIH subunit TFB5
Similarity search - Component
Biological speciesSaccharomyces cerevisiae (brewer's yeast)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 4.7 Å
AuthorsSchilbach, S. / Hantsche, M. / Tegunov, D. / Dienemann, C. / Wigge, C. / Henning, U. / Cramer, P.
Funding support Germany, 4items
OrganizationGrant numberCountry
German Research FoundationSFB860 Germany
German Research FoundationSPP1935 Germany
European Research Council693023 Germany
Volkswagen Foundation Germany
CitationJournal: Nature / Year: 2017
Title: Structures of transcription pre-initiation complex with TFIIH and Mediator.
Authors: S Schilbach / M Hantsche / D Tegunov / C Dienemann / C Wigge / H Urlaub / P Cramer /
Abstract: For the initiation of transcription, RNA polymerase II (Pol II) assembles with general transcription factors on promoter DNA to form the pre-initiation complex (PIC). Here we report cryo-electron ...For the initiation of transcription, RNA polymerase II (Pol II) assembles with general transcription factors on promoter DNA to form the pre-initiation complex (PIC). Here we report cryo-electron microscopy structures of the Saccharomyces cerevisiae PIC and PIC-core Mediator complex at nominal resolutions of 4.7 Å and 5.8 Å, respectively. The structures reveal transcription factor IIH (TFIIH), and suggest how the core and kinase TFIIH modules function in the opening of promoter DNA and the phosphorylation of Pol II, respectively. The TFIIH core subunit Ssl2 (a homologue of human XPB) is positioned on downstream DNA by the 'E-bridge' helix in TFIIE, consistent with TFIIE-stimulated DNA opening. The TFIIH kinase module subunit Tfb3 (MAT1 in human) anchors the kinase Kin28 (CDK7), which is mobile in the PIC but preferentially located between the Mediator hook and shoulder in the PIC-core Mediator complex. Open spaces between the Mediator head and middle modules may allow access of the kinase to its substrate, the C-terminal domain of Pol II.
History
DepositionAug 11, 2017Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 25, 2018Provider: repository / Type: Initial release

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Assembly

Deposited unit
A: DNA-directed RNA polymerase II subunit RPB1
B: DNA-directed RNA polymerase II subunit RPB2
C: DNA-directed RNA polymerase II subunit RPB3
D: DNA-directed RNA polymerase II subunit RPB4
E: DNA-directed RNA polymerases I, II, and III subunit RPABC1
F: DNA-directed RNA polymerases I, II, and III subunit RPABC2
G: DNA-directed RNA polymerase II subunit RPB7
H: DNA-directed RNA polymerases I, II, and III subunit RPABC3
I: DNA-directed RNA polymerase II subunit RPB9
J: DNA-directed RNA polymerases I, II, and III subunit RPABC5
K: DNA-directed RNA polymerase II subunit RPB11
L: DNA-directed RNA polymerases I, II, and III subunit RPABC4
M: Transcription initiation factor IIB
N: NONTEMPLATE DNA
O: TATA-box-binding protein
Q: Transcription initiation factor IIF subunit alpha
R: Transcription initiation factor IIF subunit beta
T: TEMPLATE DNA
U: Transcription initiation factor IIA large subunit
V: Transcription initiation factor IIA subunit 2
W: Transcription initiation factor IIE subunit alpha
X: Transcription initiation factor IIE subunit beta
0: DNA repair helicase RAD3
1: RNA polymerase II transcription factor B subunit 1
2: RNA polymerase II transcription factor B subunit 2
3: RNA polymerase II transcription factor B subunit 3
4: RNA polymerase II transcription factor B subunit 4
5: RNA polymerase II transcription factor B subunit 5
6: Suppressor of stem-loop protein 1
7: DNA repair helicase RAD25
Z: Unknown protein
hetero molecules


Theoretical massNumber of molelcules
Total (without water)1,350,17349
Polymers1,348,75131
Non-polymers1,42218
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: cross-linking
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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DNA-directed RNA polymerase II subunit ... , 7 types, 7 molecules ABCDGIK

#1: Protein DNA-directed RNA polymerase II subunit RPB1 / Polymerase / RNA polymerase II subunit B1 / DNA-directed RNA polymerase III largest subunit / RNA polymerase II ...RNA polymerase II subunit B1 / DNA-directed RNA polymerase III largest subunit / RNA polymerase II subunit B220


Mass: 191821.578 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P04050, DNA-directed RNA polymerase
#2: Protein DNA-directed RNA polymerase II subunit RPB2 / Polymerase / RNA polymerase II subunit 2 / B150 / DNA-directed RNA polymerase II 140 kDa polypeptide


Mass: 138937.297 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P08518, DNA-directed RNA polymerase
#3: Protein DNA-directed RNA polymerase II subunit RPB3 / Polymerase / RNA polymerase II subunit B3 / B44.5 / DNA-directed RNA polymerase II 45 kDa polypeptide


Mass: 35330.457 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P16370
#4: Protein DNA-directed RNA polymerase II subunit RPB4 / Polymerase / RNA polymerase II subunit B4 / B32 / DNA-directed RNA polymerase II 32 kDa polypeptide


Mass: 25451.191 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P20433
#7: Protein DNA-directed RNA polymerase II subunit RPB7 / Polymerase / RNA polymerase II subunit B7 / B16


Mass: 19081.053 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P34087
#9: Protein DNA-directed RNA polymerase II subunit RPB9 / Polymerase / RNA polymerase II subunit B9 / B12.6 / DNA-directed RNA polymerase II 14.2 kDa polypeptide / DNA- ...RNA polymerase II subunit B9 / B12.6 / DNA-directed RNA polymerase II 14.2 kDa polypeptide / DNA-directed RNA polymerase II subunit 9


Mass: 14308.161 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P27999
#11: Protein DNA-directed RNA polymerase II subunit RPB11 / Polymerase / RNA polymerase II subunit B11 / B13.6 / DNA-directed RNA polymerase II 13.6 kDa polypeptide


Mass: 13633.493 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P38902

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DNA-directed RNA polymerases I, II, and III subunit ... , 5 types, 5 molecules EFHJL

#5: Protein DNA-directed RNA polymerases I, II, and III subunit RPABC1 / RNA polymerase / RNA polymerases I / II / and III subunit ABC1 / ABC27 / DNA-directed RNA polymerases I / and III 27 ...RNA polymerases I / II / and III subunit ABC1 / ABC27 / DNA-directed RNA polymerases I / and III 27 kDa polypeptide


Mass: 25117.094 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P20434
#6: Protein DNA-directed RNA polymerases I, II, and III subunit RPABC2 / RNA polymerase / RNA polymerases I / II / and III subunit ABC2 / ABC23 / DNA-directed RNA polymerases I / and III 23 ...RNA polymerases I / II / and III subunit ABC2 / ABC23 / DNA-directed RNA polymerases I / and III 23 kDa polypeptide


Mass: 17931.834 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P20435
#8: Protein DNA-directed RNA polymerases I, II, and III subunit RPABC3 / RNA polymerase / RNA polymerases I / II / and III subunit ABC3 / ABC14.4 / ABC14.5 / DNA-directed RNA polymerases I ...RNA polymerases I / II / and III subunit ABC3 / ABC14.4 / ABC14.5 / DNA-directed RNA polymerases I / and III 14.5 kDa polypeptide


Mass: 16525.363 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P20436
#10: Protein DNA-directed RNA polymerases I, II, and III subunit RPABC5 / RNA polymerase / RNA polymerases I / II / and III subunit ABC5 / ABC10-beta / ABC8 / DNA-directed RNA polymerases I ...RNA polymerases I / II / and III subunit ABC5 / ABC10-beta / ABC8 / DNA-directed RNA polymerases I / and III 8.3 kDa polypeptide


Mass: 8290.732 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P22139
#12: Protein DNA-directed RNA polymerases I, II, and III subunit RPABC4 / RNA polymerase / RNA polymerases I / II / and III subunit ABC4 / ABC10-alpha


Mass: 7729.969 Da / Num. of mol.: 1 / Source method: isolated from a natural source
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P40422

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Protein , 3 types, 3 molecules MO6

#13: Protein Transcription initiation factor IIB / General transcription factor TFIIB / Transcription factor E


Mass: 38257.340 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: SUA7, YPR086W, P9513.4 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P29055
#15: Protein TATA-box-binding protein / TATA sequence-binding protein / TBP / TATA-binding factor / TATA-box factor / Transcription factor ...TATA sequence-binding protein / TBP / TATA-binding factor / TATA-box factor / Transcription factor D / Transcription initiation factor TFIID TBP subunit


Mass: 27042.275 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: SPT15, BTF1, TBP1, YER148W / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P13393
#29: Protein Suppressor of stem-loop protein 1 / General transcription and DNA repair factor IIH subunit SSL1 / TFIIH subunit SSL1


Mass: 51122.695 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: SSL1, YLR005W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q04673

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DNA chain , 2 types, 2 molecules NT

#14: DNA chain NONTEMPLATE DNA


Mass: 32716.977 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#18: DNA chain TEMPLATE DNA


Mass: 32680.920 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)

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Transcription initiation factor IIF subunit ... , 2 types, 2 molecules QR

#16: Protein Transcription initiation factor IIF subunit alpha / TFIIF-alpha / TFIIF large subunit / Transcription factor G 105 kDa subunit / P105


Mass: 82320.570 Da / Num. of mol.: 1
Mutation: Residues indicated as UNK are of uncertain amino acid register and may deviate from the given register by few amino acids. For details about modelling of the subunits refer to the ...Mutation: Residues indicated as UNK are of uncertain amino acid register and may deviate from the given register by few amino acids. For details about modelling of the subunits refer to the associated publication. Residues indicated as UNK with the chain identifier Z are part of an assembly between the clutch domains of Tfb2 and Ssl2 but could not be assigned to any of the two proteins with confidence since domain swapping/beta-strand exchange may occur. To account for this uncertainty, a theoretical chain Z was introduced for this residue range but not assigned to any of the PIC components. Submission based on experimental data from EMDB EMD-3846.
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFG1, SSU71, YGR186W, G7526 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P41895
#17: Protein Transcription initiation factor IIF subunit beta / ATP-dependent helicase TFG2 / TFIIF medium subunit / TFIIF-beta / Transcription factor G 54 kDa subunit


Mass: 46684.492 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFG2, YGR005C / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P41896, DNA helicase

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Transcription initiation factor IIA ... , 2 types, 2 molecules UV

#19: Protein Transcription initiation factor IIA large subunit / TFIIA large subunit / TFIIA 32 kDa subunit


Mass: 32230.805 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TOA1, YOR194C / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P32773
#20: Protein Transcription initiation factor IIA subunit 2 / General transcription factor IIA subunit 2 / TFIIA 13.5 kDa subunit / Transcription initiation ...General transcription factor IIA subunit 2 / TFIIA 13.5 kDa subunit / Transcription initiation factor IIA small chain / Transcription initiation factor IIA small subunit


Mass: 13473.070 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TOA2, YKL058W / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P32774

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Transcription initiation factor IIE subunit ... , 2 types, 2 molecules WX

#21: Protein Transcription initiation factor IIE subunit alpha / TFIIE-alpha / Factor A 66 kDa subunit / Transcription factor A large subunit


Mass: 57538.461 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFA1, YKL028W / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P36100
#22: Protein Transcription initiation factor IIE subunit beta / TFIIE-beta / Factor A 43 kDa subunit / Transcription factor A small subunit


Mass: 37050.434 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFA2, YKR062W / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P36145

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DNA repair helicase ... , 2 types, 2 molecules 07

#23: Protein DNA repair helicase RAD3 / General transcription and DNA repair factor IIH subunit RAD3 / TFIIH subunit RAD3


Mass: 89064.555 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: RAD3, REM1, YER171W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P06839, DNA helicase
#30: Protein DNA repair helicase RAD25 / General transcription and DNA repair factor IIH subunit RAD25 / TFIIH subunit RAD25 / Suppressor of ...General transcription and DNA repair factor IIH subunit RAD25 / TFIIH subunit RAD25 / Suppressor of stem-loop mutation 2


Mass: 94049.695 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: SSL2, LOM3, RAD25, UVS112, YIL143C / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q00578, DNA helicase

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RNA polymerase II transcription factor B subunit ... , 5 types, 5 molecules 12345

#24: Protein RNA polymerase II transcription factor B subunit 1 / General transcription and DNA repair factor IIH subunit TFB1 / TFIIH subunit TFB1 / RNA polymerase ...General transcription and DNA repair factor IIH subunit TFB1 / TFIIH subunit TFB1 / RNA polymerase II transcription factor B 73 kDa subunit / RNA polymerase II transcription factor B p73 subunit


Mass: 65440.922 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFB1, YDR311W, D9740.3 / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: P32776
#25: Protein RNA polymerase II transcription factor B subunit 2 / General transcription and DNA repair factor IIH subunit TFB2 / TFIIH subunit TFB2 / RNA polymerase ...General transcription and DNA repair factor IIH subunit TFB2 / TFIIH subunit TFB2 / RNA polymerase II transcription factor B 52 kDa subunit / RNA polymerase II transcription factor B p52 subunit


Mass: 48406.277 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFB2, YPL122C / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q02939
#26: Protein RNA polymerase II transcription factor B subunit 3 / RNA polymerase II transcription factor B 38 kDa subunit / RNA polymerase II transcription factor B p38 subunit


Mass: 38188.438 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFB3, RIG2, YDR460W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q03290
#27: Protein RNA polymerase II transcription factor B subunit 4 / General transcription and DNA repair factor IIH subunit TFB4 / TFIIH subunit TFB4 / RNA polymerase ...General transcription and DNA repair factor IIH subunit TFB4 / TFIIH subunit TFB4 / RNA polymerase II transcription factor B 34 kDa subunit / RNA polymerase II transcription factor B p34 subunit


Mass: 36403.598 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFB4, YPR056W / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q12004
#28: Protein RNA polymerase II transcription factor B subunit 5 / General transcription and DNA repair factor IIH subunit TFB5 / TFIIH subunit TFB5


Mass: 8243.490 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFB5, YDR079C-A / Production host: Trichoplusia ni (cabbage looper) / References: UniProt: Q3E7C1

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Protein/peptide , 1 types, 1 molecules Z

#31: Protein/peptide Unknown protein


Mass: 3677.524 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Details: Residues indicated as UNK with the chain identifier Z are part of an assembly between the clutch domains of Tfb2 and Ssl2 but could not be assigned to any of the two proteins with confidence ...Details: Residues indicated as UNK with the chain identifier Z are part of an assembly between the clutch domains of Tfb2 and Ssl2 but could not be assigned to any of the two proteins with confidence since domain swapping/beta-strand exchange may occur. To account for this uncertainty, a theoretical chain Z was introduced for this residue range but not assigned to any of the PIC components.
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Production host: Trichoplusia ni (cabbage looper)

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Non-polymers , 3 types, 18 molecules

#32: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 16 / Source method: obtained synthetically / Formula: Zn
#33: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#34: Chemical ChemComp-SF4 / IRON/SULFUR CLUSTER / Iron–sulfur cluster


Mass: 351.640 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Fe4S4

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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
IDNameTypeDetailsEntity IDParent-IDSource
1Pre-Initiation Complex (PIC)COMPLEXPRE-INITIATION COMPLEX WITH TFIIH#1-#310MULTIPLE SOURCES
2PolymeraseCOMPLEXnatural source entities#1-#121NATURAL
3initiation factorsInitiation factorCOMPLEXentitites expressed in coli#13, #15-#17, #19-#221RECOMBINANT
4Transcription factorsTranscription factorCOMPLEXentitites expressed in T.ni#23-#311RECOMBINANT
5nucleic acidCOMPLEXSynthetic nucleic acid#14, #181RECOMBINANT
Molecular weightValue: 1.43 MDa / Experimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
12Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)559292
23Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)559292
34Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)559292
45synthetic construct (others)32630
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
13Escherichia coli BL21(DE3) (bacteria)469008
24Trichoplusia ni (cabbage looper)7111
35synthetic construct (others)32630
Buffer solutionpH: 7.6
SpecimenConc.: 0.7 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportDetails: QUANTIFOIL HOLEY GOLD GRIDS WERE GLOW-DISCHARGED BEFORE DEPOSITION OF SAMPLE.
Grid material: GOLD / Grid type: Quantifoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE
Details: GRIDS WERE BLOTTED FOR 8 S AND PLUNGE-FROZEN IN LIQUID 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 FIELDBright-field microscopy / Nominal magnification: 105000 X / Nominal defocus max: 5000 nm / Nominal defocus min: 500 nm / Cs: 2.7 mm / Alignment procedure: COMA FREE
Specimen holderCryogen: NITROGEN / Specimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER
Image recordingAverage exposure time: 10 sec. / Electron dose: 42 e/Å2 / Detector mode: COUNTING / Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Num. of grids imaged: 3 / Num. of real images: 13000
Image scansMovie frames/image: 40

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Processing

EM software
IDNameVersionCategory
1EPUimage acquisition
3GctfCTF correction
8RELION2.0.4initial Euler assignment
9RELION2.0.4final Euler assignment
10RELION2.0.4classification
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 4.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 58000 / Symmetry type: POINT
Atomic model buildingProtocol: OTHER / Space: REAL

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