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- EMDB-3590: Cryo-EM structure of RNA polymerase I in complex with Rrn3 and Co... -

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Entry
Database: EMDB / ID: 3590
TitleCryo-EM structure of RNA polymerase I in complex with Rrn3 and Core Factor (Orientation III)
Map data
SampleRNA polymerase I in complex with Rrn3 and Core Factor
  • RNA polymerase I
  • Initiation factor Rrn3
  • Core Factor
  • (DNA-directed RNA polymerase I subunit ...Polymerase) x 7
  • (DNA-directed RNA polymerases I and III subunit ...) x 2
  • (DNA-directed RNA polymerases I, II, and III subunit ...RNA polymerase) x 5
  • (RNA polymerase I-specific transcription initiation factor ...) x 4
  • (ligand) x 3
Function / homologyRNA polymerase subunit RPB10 / RNA polymerase Rpb7-like, N-terminal domain superfamily / Pol I subunit A12, C-terminal zinc ribbon / RPB5-like RNA polymerase subunit superfamily / RPB6/omega subunit-like superfamily / RNA polymerase, RBP11-like subunit / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase Rpb5, N-terminal domain superfamily / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / DNA-directed RNA polymerases I and III subunit AC19 ...RNA polymerase subunit RPB10 / RNA polymerase Rpb7-like, N-terminal domain superfamily / Pol I subunit A12, C-terminal zinc ribbon / RPB5-like RNA polymerase subunit superfamily / RPB6/omega subunit-like superfamily / RNA polymerase, RBP11-like subunit / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase Rpb5, N-terminal domain superfamily / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / DNA-directed RNA polymerases I and III subunit AC19 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, funnel domain superfamily / DNA-directed RNA polymerase subunit Rpb5-like / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / RNA polymerase Rpb2, domain 6 / RNA polymerase Rpb1, domain 2 / DNA-directed RNA polymerases I and III subunit AC40 / RNA polymerase I transcription initiation factor TAF1B/Rrn7 / Transcription factor S-II (TFIIS) / RNA polymerase I-specific transcription initiation factor RRN6-like / DNA-directed RNA polymerase RPB5 subunit, eukaryote/virus / RNA polymerase Rpb2, OB-fold / DNA-directed RNA pol I, largest subunit / DNA-directed RNA polymerase, subunit 2 / RNA polymerase I-specific transcription initiation factor Rrn6 / Transcription initiation factor Rrn11, budding yeast / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / 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 / Transcription initiation factor Rrn7, Zinc-finger / Dual incision in TC-NER / DNA-directed RNA polymerase, subunit RPB6 / RNA polymerase Rpb3/RpoA insert domain / RNA polymerase Rpb5, C-terminal domain / RNA polymerase I, subunit Rpa14, fungi / Zinc finger TFIIS-type signature. / Yeast RNA polymerase I subunit RPA14 / DNA-directed RNA polymerase I subunit RPA34.5 / RNA polymerase I-specific transcription-initiation factor / Zinc-finger of RNA-polymerase I-specific TFIIB, Rrn7 / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerases D / 30 to 40 Kd subunits signature. / RNA polymerases M / 15 Kd subunits signature. / A49-like RNA polymerase I associated factor / 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. / Formation of the Early Elongation Complex / RNA polymerase I, Rpa2 specific domain / RNA polymerase I specific transcription initiation factor RRN3 / RNA polymerase Rpb6 / SHS2 domain found in N terminus of Rpb7p/Rpc25p/MJ0397 / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerases N / 8 kDa subunit / RNA polymerases M/15 Kd subunit / DNA directed RNA polymerase, 7 kDa subunit / RNA polymerase Rpb8 / RNA polymerase Rpb5, N-terminal domain / RNA polymerase I specific initiation factor / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 2 / RNA polymerase beta subunit / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 5 / DNA-directed RNA polymerase I, subunit RPA34.5 / Nucleic acid-binding, OB-fold / Formation of TC-NER Pre-Incision Complex / TP53 Regulates Transcription of DNA Repair Genes / RNA polymerase, Rpb8 / DNA-directed RNA polymerase, M/15kDa subunit / DNA-directed RNA polymerase, 30-40kDa subunit, conserved site / Zinc finger, TFIIS-type / RNA polymerase, subunit H/Rpb5 C-terminal / RNA polymerase, alpha subunit / DNA-directed RNA polymerase, subunit N/Rpb10 / RNA polymerase subunit, RPB6/omega / Gap-filling DNA repair synthesis and ligation in TC-NER / RNA polymerase II transcribes snRNA genes / RNA polymerase Rpb7, N-terminal / 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 / RNA polymerase, Rpb5, N-terminal / RNA Polymerase II Pre-transcription Events / RNA polymerase, subunit omega/K/RPB6
Function and homology information
SourceSaccharomyces cerevisiae (baker's yeast) / Baker's yeast (baker's yeast)
Methodsingle particle reconstruction / cryo EM / 7.7 Å resolution
AuthorsEngel C / Gubbey T / Neyer S / Sainsbury S / Oberthuer C / Baejen C / Bernecky C / Cramer P
CitationJournal: Cell / Year: 2017
Title: Structural Basis of RNA Polymerase I Transcription Initiation.
Authors: Christoph Engel / Tobias Gubbey / Simon Neyer / Sarah Sainsbury / Christiane Oberthuer / Carlo Baejen / Carrie Bernecky / Patrick Cramer
Abstract: Transcription initiation at the ribosomal RNA promoter requires RNA polymerase (Pol) I and the initiation factors Rrn3 and core factor (CF). Here, we combine X-ray crystallography and cryo-electron ...Transcription initiation at the ribosomal RNA promoter requires RNA polymerase (Pol) I and the initiation factors Rrn3 and core factor (CF). Here, we combine X-ray crystallography and cryo-electron microscopy (cryo-EM) to obtain a molecular model for basal Pol I initiation. The three-subunit CF binds upstream promoter DNA, docks to the Pol I-Rrn3 complex, and loads DNA into the expanded active center cleft of the polymerase. DNA unwinding between the Pol I protrusion and clamp domains enables cleft contraction, resulting in an active Pol I conformation and RNA synthesis. Comparison with the Pol II system suggests that promoter specificity relies on a distinct "bendability" and "meltability" of the promoter sequence that enables contacts between initiation factors, DNA, and polymerase.
Validation ReportPDB-ID: 5n5y

SummaryFull reportAbout validation report
DateDeposition: Feb 14, 2017 / Header (metadata) release: Mar 22, 2017 / Map release: Apr 5, 2017 / Last update: Oct 3, 2018

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0205
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.0205
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-5n5y
  • Surface level: 0.0205
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_3590.map.gz (map file in CCP4 format, 55297 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
240 pix
1.35 Å/pix.
= 324. Å
240 pix
1.35 Å/pix.
= 324. Å
240 pix
1.35 Å/pix.
= 324. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.35 Å
Density
Contour Level:0.0205 (by author), 0.0205 (movie #1):
Minimum - Maximum-0.029996127 - 0.06755038
Average (Standard dev.)-0.000064416134 (0.004714947)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions240240240
Origin0.00.00.0
Limit239.0239.0239.0
Spacing240240240
CellA=B=C: 324.0 Å
α=β=γ: 90.0 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.351.351.35
M x/y/z240240240
origin x/y/z0.0000.0000.000
length x/y/z324.000324.000324.000
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.0300.068-0.000

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Supplemental data

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Sample components

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Entire RNA polymerase I in complex with Rrn3 and Core Factor

EntireName: RNA polymerase I in complex with Rrn3 and Core Factor / Number of components: 25

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Component #1: protein, RNA polymerase I in complex with Rrn3 and Core Factor

ProteinName: RNA polymerase I in complex with Rrn3 and Core Factor / Recombinant expression: No

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Component #2: protein, RNA polymerase I

ProteinName: RNA polymerase I / Recombinant expression: No
MassTheoretical: 590 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)

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Component #3: protein, Initiation factor Rrn3

ProteinName: Initiation factor Rrn3 / Recombinant expression: No
MassTheoretical: 70 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #4: protein, Core Factor

ProteinName: Core Factor / Recombinant expression: No
MassTheoretical: 220 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #5: protein, DNA-directed RNA polymerase I subunit RPA190

ProteinName: DNA-directed RNA polymerase I subunit RPA190Polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 186.676969 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #6: protein, DNA-directed RNA polymerase I subunit RPA135

ProteinName: DNA-directed RNA polymerase I subunit RPA135Polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 135.910328 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #7: protein, DNA-directed RNA polymerases I and III subunit RPAC1

ProteinName: DNA-directed RNA polymerases I and III subunit RPAC1 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 37.732613 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #8: protein, DNA-directed RNA polymerase I subunit RPA14

ProteinName: DNA-directed RNA polymerase I subunit RPA14Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 14.599128 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #9: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC1

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC1RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 25.117094 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #10: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC2

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC2RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 17.931834 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #11: protein, DNA-directed RNA polymerase I subunit RPA43

ProteinName: DNA-directed RNA polymerase I subunit RPA43Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 36.264852 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #12: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC3

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC3RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 16.525363 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #13: protein, DNA-directed RNA polymerase I subunit RPA12

ProteinName: DNA-directed RNA polymerase I subunit RPA12Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 13.676566 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #14: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC5

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC5RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 8.290732 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #15: protein, DNA-directed RNA polymerases I and III subunit RPAC2

ProteinName: DNA-directed RNA polymerases I and III subunit RPAC2 / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 16.16786 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #16: protein, DNA-directed RNA polymerases I, II, and III subunit RPABC4

ProteinName: DNA-directed RNA polymerases I, II, and III subunit RPABC4RNA polymerase
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 7.729969 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #17: protein, DNA-directed RNA polymerase I subunit RPA49

ProteinName: DNA-directed RNA polymerase I subunit RPA49Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 46.721707 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #18: protein, DNA-directed RNA polymerase I subunit RPA34

ProteinName: DNA-directed RNA polymerase I subunit RPA34Polymerase / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 26.933518 kDa
SourceSpecies: Baker's yeast (baker's yeast)

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Component #19: protein, RNA polymerase I-specific transcription initiation facto...

ProteinName: RNA polymerase I-specific transcription initiation factor RRN3
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 72.458258 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #20: protein, RNA polymerase I-specific transcription initiation facto...

ProteinName: RNA polymerase I-specific transcription initiation factor RRN6
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 102.163227 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #21: protein, RNA polymerase I-specific transcription initiation facto...

ProteinName: RNA polymerase I-specific transcription initiation factor RRN7
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 60.435195 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #22: protein, RNA polymerase I-specific transcription initiation facto...

ProteinName: RNA polymerase I-specific transcription initiation factor RRN11
Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 59.334262 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #23: ligand, ZINC ION

LigandName: ZINC ION / Number of Copies: 8 / Recombinant expression: No
MassTheoretical: 6.540905 MDa

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Component #24: ligand, SULFATE ION

LigandName: SULFATE IONSulfate / Number of Copies: 5 / Recombinant expression: No
MassTheoretical: 9.606305 MDa

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Component #25: ligand, MAGNESIUM ION

LigandName: MAGNESIUM ION / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 2.430505 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.1 mg/ml / pH: 7.8
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 298 K / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 4 e/Å2 / Illumination mode: FLOOD BEAM
LensImaging mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of projections: 8317
3D reconstructionResolution: 7.7 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Modeling #1Refinement protocol: rigid body
Output model

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