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- PDB-6gyl: Structure of a yeast closed complex with distorted DNA (core CCdist) -

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Entry
Database: PDB / ID: 6gyl
TitleStructure of a yeast closed complex with distorted DNA (core CCdist)
Components
  • (DNA-directed RNA polymerase II subunit ...Polymerase) x 7
  • (DNA-directed RNA polymerases I, II, and III subunit ...RNA polymerase) x 5
  • (GAT1 promoter ...) x 2
  • (Transcription initiation factor IIA ...) x 2
  • (Transcription initiation factor IIE subunit ...) x 2
  • (Transcription initiation factor IIF subunit ...) x 2
  • TATA-box-binding protein
  • Transcription initiation factor IIB
KeywordsTRANSCRIPTION / rna polymerase II / transcription initiation / promoter dna opening
Function / homologyTFA2 Winged helix domain 2 / RNA polymerase RBP11 / Pol II subunit B9, C-terminal zinc ribbon / RPB5-like RNA polymerase subunit superfamily / RPB6/omega subunit-like superfamily / Winged helix-like DNA-binding domain superfamily / Winged helix DNA-binding domain superfamily / RNA polymerase, RBP11-like subunit / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase Rpb5, N-terminal domain superfamily ...TFA2 Winged helix domain 2 / RNA polymerase RBP11 / Pol II subunit B9, C-terminal zinc ribbon / RPB5-like RNA polymerase subunit superfamily / RPB6/omega subunit-like superfamily / Winged helix-like DNA-binding domain superfamily / Winged helix DNA-binding domain superfamily / RNA polymerase, RBP11-like subunit / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase Rpb5, N-terminal domain superfamily / RNA polymerase Rpb7-like, N-terminal domain superfamily / Cyclin-like superfamily / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, funnel domain superfamily / TATA-box binding protein, conserved site / Rpb4/RPC9 superfamily / RNA polymerase Rpb1, domain 7 superfamily / DNA-directed RNA polymerase subunit Rpb5-like / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / Transcription factor E / Transcription factor TFIID (or TATA-binding protein, TBP) / Transcription factor TFIIB repeat / RNA polymerase Rpb2, domain 6 / S1 RNA binding domain / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb3/RpoA insert domain / Transcription factor S-II (TFIIS) / RNA polymerase Rpb5, C-terminal domain / TATA-box binding protein, eukaryotic / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerase Rpb2, OB-fold / Transcription factor IIA, beta-barrel / HRDC-like superfamily / Transcription Factor IIF, Rap30/Rap74, interaction / DNA-directed RNA polymerase, insert domain / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / DNA-directed RNA polymerase subunit/transcription factor S / RNA polymerase subunit, RPB6/omega / Nucleic acid-binding, OB-fold / Zinc finger, RING/FYVE/PHD-type / Zinc finger, TFIIB-type / Transcription factor TFIIB, cyclin-like domain / Cyclin-like / DNA-directed RNA polymerase RPB5 subunit, eukaryote/virus / DNA-directed RNA polymerase, subunit 2 / DNA-directed RNA polymerase, subunit RPB6 / Transcription initiation factor IIA, gamma subunit, C-terminal / Transcription initiation factor IIA, gamma subunit, N-terminal / Transcription initiation factor TFIIE, beta subunit / Transcription factor TFE/TFIIEalpha HTH domain / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / RNA polymerase, subunit H/Rpb5, conserved site / Archaeal RpoH /eukaryotic RPB5 RNA polymerase subunit / DNA-directed RNA polymerase, 14-18kDa subunit, conserved site / RNA polymerases, subunit N, zinc binding site / RNA-binding domain, S1 / Transcription factor TFIIB, conserved site / RNA polymerase subunit RPB10 / TFIIEalpha/SarR/Rpc3 HTH domain / RNA polymerase Rpb6 / RNA polymerases N / 8 kDa subunit / DNA-directed RNA polymerase, RBP11-like dimerisation domain / Formation of TC-NER Pre-Incision Complex / RNA polymerases M / 15 Kd subunits signature. / RNA polymerases H / 23 Kd subunits signature. / RNA polymerases K / 14 to 18 Kd subunits signature. / RNA polymerases N / 8 Kd subunits signature. / RNA polymerases L / 13 to 16 Kd subunits signature. / RNA polymerases beta chain signature. / Zinc finger TFIIS-type profile. / Zinc finger TFIIB-type profile. / TFE/IIEalpha-type HTH domain profile. / TFIIE beta central core DNA-binding domain profile. / Formation of the Early Elongation Complex / RNA Polymerase II Pre-transcription Events / Dual incision in TC-NER / Zinc finger TFIIS-type signature. / Gap-filling DNA repair synthesis and ligation in TC-NER / TP53 Regulates Transcription of DNA Repair Genes / RNA polymerase II transcribes snRNA genes / mRNA Capping / mRNA Splicing - Minor Pathway / RNA Polymerase I Transcription Initiation / 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 Pol II CTD phosphorylation and interaction with CE / Estrogen-dependent gene expression / TFIIF, beta subunit N-terminus / Transcription factor TFIIB repeat signature. / RNA polymerases D / 30 to 40 Kd subunits signature. / TFIIE alpha subunit / RNA polymerase beta subunit / RNA polymerases M/15 Kd subunit / TFIIE beta subunit core domain / Transcription initiation factor IIA, gamma subunit, helical domain / TFIIF, beta subunit HTH domain
Function and homology information
Specimen sourceSaccharomyces cerevisiae (baker's yeast)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / 4.8 Å resolution
AuthorsDienemann, C. / Schwalb, B. / Schilbach, S. / Cramer, P.
CitationJournal: Mol. Cell / Year: 2018
Title: Promoter Distortion and Opening in the RNA Polymerase II Cleft.
Authors: Christian Dienemann / Björn Schwalb / Sandra Schilbach / Patrick Cramer
Abstract: Transcription initiation requires opening of promoter DNA in the RNA polymerase II (Pol II) pre-initiation complex (PIC), but it remains unclear how this is achieved. Here we report the cryo-electron ...Transcription initiation requires opening of promoter DNA in the RNA polymerase II (Pol II) pre-initiation complex (PIC), but it remains unclear how this is achieved. Here we report the cryo-electron microscopic (cryo-EM) structure of a yeast PIC that contains underwound, distorted promoter DNA in the closed Pol II cleft. The DNA duplex axis is offset at the upstream edge of the initially melted DNA region (IMR) where DNA opening begins. Unstable IMRs are found in a subset of yeast promoters that we show can still initiate transcription after depletion of the transcription factor (TF) IIH (TFIIH) translocase Ssl2 (XPB in human) from the nucleus in vivo. PIC-induced DNA distortions may thus prime the IMR for melting and may explain how unstable IMRs that are predicted in promoters of Pol I and Pol III can open spontaneously. These results suggest that DNA distortion in the polymerase cleft is a general mechanism that contributes to promoter opening.
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Jun 30, 2018 / Release: Dec 5, 2018

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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: GAT1 promoter DNA
O: TATA-box-binding protein
Q: Transcription initiation factor IIF subunit alpha
R: Transcription initiation factor IIF subunit beta
T: GAT1 promoter DNA
U: Transcription initiation factor IIA large subunit,Transcription initiation factor IIA large subunit
V: Transcription initiation factor IIA subunit 2
W: Transcription initiation factor IIE subunit alpha,Tfa1,Tfa1
X: Transcription initiation factor IIE subunit beta
hetero molecules


Theoretical massNumber of molelcules
Total (without water)872,11133
Polyers871,43322
Non-polymers67811
Water0
1


TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

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

#1: Protein/peptide DNA-directed RNA polymerase II subunit RPB1 / Polymerase / 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: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P04050, DNA-directed RNA polymerase
#2: Protein/peptide 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: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P08518, DNA-directed RNA polymerase
#3: Protein/peptide 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: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P16370
#4: Protein/peptide 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: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P20433
#7: Protein/peptide DNA-directed RNA polymerase II subunit RPB7 / Polymerase / RNA polymerase II subunit B7 / B16


Mass: 19081.053 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P34087
#9: Protein/peptide 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-directed RNA polymerase II subunit 9


Mass: 14308.161 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P27999
#11: Protein/peptide 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: (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/peptide 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 kDa polypeptide


Mass: 25117.094 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P20434
#6: Protein/peptide 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 kDa polypeptide


Mass: 17931.834 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P20435
#8: Protein/peptide 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 / and III 14.5 kDa polypeptide


Mass: 16525.363 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P20436
#10: Protein/peptide 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 / and III 8.3 kDa polypeptide


Mass: 8290.732 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P22139
#12: Protein/peptide 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: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
References: UniProt: P40422

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Protein/peptide , 2 types, 2 molecules MO

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


Mass: 38257.340 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: SUA7, YPR086W, P9513.4 / Production host: Escherichia coli (E. coli) / References: UniProt: P29055
#15: Protein/peptide TATA-box-binding protein / 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: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: SPT15, BTF1, TBP1, YER148W / Production host: Escherichia coli (E. coli) / References: UniProt: P13393

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GAT1 promoter ... , 2 types, 2 molecules NT

#14: DNA chain GAT1 promoter DNA


Mass: 17129.941 Da / Num. of mol.: 1
Source: (synth.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
#18: DNA chain GAT1 promoter DNA


Mass: 17388.111 Da / Num. of mol.: 1
Source: (synth.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)

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

#16: Protein/peptide 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
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFG1, SSU71, YGR186W, G7526 / Production host: Escherichia coli (E. coli) / References: UniProt: P41895
#17: Protein/peptide 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: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFG2, YGR005C / Production host: Escherichia coli (E. coli) / References: UniProt: P41896, DNA helicase

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

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


Mass: 19432.029 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TOA1, YOR194C / Production host: Escherichia coli (E. coli) / References: UniProt: P32773
#20: Protein/peptide Transcription initiation factor IIA subunit 2 / General transcription factor IIA subunit 2 / TFIIA 13.5 kDa subunit / Transcription initiation factor IIA small chain / Transcription initiation factor IIA small subunit


Mass: 14431.131 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TOA2, YKL058W / Production host: Escherichia coli (E. coli) / References: UniProt: P32774

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

#21: Protein/peptide Transcription initiation factor IIE subunit alpha,Tfa1,Tfa1 / TFIIE-alpha / Factor A 66 kDa subunit / Transcription factor A large subunit


Mass: 57538.461 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFA1, YKL028W / Production host: Escherichia coli (E. coli) / References: UniProt: P36100
#22: Protein/peptide 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: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Gene: TFA2, YKR062W / Production host: Escherichia coli (E. coli) / References: UniProt: P36145

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Non-polymers , 2 types, 11 molecules

#23: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 10 / Formula: Zn / Zinc
#24: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Formula: Mg / Magnesium

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Experimental details

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Experiment

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

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Sample preparation

Component
IDNameTypeEntity IDParent IDSource
1Pre-initiation complexCOMPLEX1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 220MULTIPLE SOURCES
2PolymeraseCOMPLEX1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 121NATURAL
3TIFsCOMPLEX13, 15, 16, 17, 19, 20, 21, 221RECOMBINANT
4nucleic acidsCOMPLEX14, 181RECOMBINANT
Source (natural)
IDEntity assembly IDNcbi tax IDOrganism
12559292Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
23559292Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
34559292Saccharomyces cerevisiae (strain ATCC 204508 / S288c) (yeast)
Source (recombinant)
IDEntity assembly IDNcbi tax IDOrganism
13562Escherichia coli (E. coli)
2432630synthetic construct (others)
Buffer solutionpH: 7
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyMicroscope model: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: BRIGHT FIELDBright-field microscopy
Image recordingElectron dose: 37 e/Å2 / Film or detector model: GATAN K2 QUANTUM (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.13_2998: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1
3D reconstructionResolution: 4.8 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 38000 / Symmetry type: POINT

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