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- EMDB-3439: RNA polymerase I-Rrn3 complex at 4.8 A resolution -

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Basic information

Entry
Database: EMDB / ID: EMD-3439
TitleRNA polymerase I-Rrn3 complex at 4.8 A resolution
Map data
SampleS. cerevisiae RNA polymerase I bound to the initiation factor Rrn3:
(RNA polymerase ...) x 2
KeywordsRNA polymerase / transciption
Function / homology
Function and homology information


RNA polymerase I general transcription initiation factor activity / RNA polymerase I preinitiation complex assembly / RNA polymerase I core binding / RNA polymerase I general transcription initiation factor binding / transcription initiation from RNA polymerase I promoter / rDNA binding / RNA polymerase II-specific DNA-binding transcription factor binding / nucleolar large rRNA transcription by RNA polymerase I / transposon integration / transcription elongation from RNA polymerase I promoter ...RNA polymerase I general transcription initiation factor activity / RNA polymerase I preinitiation complex assembly / RNA polymerase I core binding / RNA polymerase I general transcription initiation factor binding / transcription initiation from RNA polymerase I promoter / rDNA binding / RNA polymerase II-specific DNA-binding transcription factor binding / nucleolar large rRNA transcription by RNA polymerase I / transposon integration / transcription elongation from RNA polymerase I promoter / termination of RNA polymerase I transcription / termination of RNA polymerase III transcription / RNA polymerase I activity / regulation of cell size / tRNA transcription by RNA polymerase III / RNA polymerase I complex / RNA polymerase III complex / mRNA cleavage / RNA polymerase II, core complex / transcription by RNA polymerase I / transcription by RNA polymerase III / transcription, RNA-templated / ribonucleoside binding / DNA-directed RNA polymerase / promoter-specific chromatin binding / ribosome biogenesis / transcription by RNA polymerase II / nucleic acid binding / protein dimerization activity / nucleolus / negative regulation of transcription by RNA polymerase II / DNA binding / zinc ion binding / nucleoplasm / metal ion binding / nucleus / cytoplasm
RNA polymerase Rpb2, domain 2 / RNA polymerase, beta subunit, conserved site / RNA polymerase I associated factor, A49-like / DNA-directed RNA polymerase, RBP11-like dimerisation domain / DNA-directed RNA polymerase Rpb11, 13-16kDa subunit, conserved site / RNA polymerase I specific transcription initiation factor RRN3 / RNA polymerase Rpb2, domain 3 / RNA polymerase, beta subunit, protrusion / RPB6/omega subunit-like superfamily / RNA polymerase Rpb2, domain 7 ...RNA polymerase Rpb2, domain 2 / RNA polymerase, beta subunit, conserved site / RNA polymerase I associated factor, A49-like / DNA-directed RNA polymerase, RBP11-like dimerisation domain / DNA-directed RNA polymerase Rpb11, 13-16kDa subunit, conserved site / RNA polymerase I specific transcription initiation factor RRN3 / RNA polymerase Rpb2, domain 3 / RNA polymerase, beta subunit, protrusion / RPB6/omega subunit-like superfamily / RNA polymerase Rpb2, domain 7 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain / DNA-directed RNA polymerase, insert domain / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 3 / RNA polymerase, N-terminal / RNA polymerase archaeal subunit P/eukaryotic subunit RPABC4 / Archaeal RpoK/eukaryotic RPB6 RNA polymerase subunit / RNA polymerase, subunit omega/K/RPB6 / DNA-directed RNA polymerase I subunit RPA2, domain 4 / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / RNA polymerase, Rpb5, N-terminal / RNA polymerase, subunit H/Rpb5, conserved site / DNA-directed RNA polymerases I and III subunit AC40 / DNA-directed RNA polymerases I and III subunit AC19 / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / DNA-directed RNA polymerase, subunit RPB6 / RNA polymerase subunit RPB10 / RNA polymerases, subunit N, zinc binding site / DNA-directed RNA polymerase, 14-18kDa subunit, conserved site / Archaeal RpoH /eukaryotic RPB5 RNA polymerase subunit / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / DNA-directed RNA polymerase subunit/transcription factor S / DNA-directed RNA polymerase, subunit 2 / DNA-directed RNA pol I, largest subunit / RNA polymerase Rpb2, OB-fold / DNA-directed RNA polymerase RPB5 subunit, eukaryote/virus / DNA-directed RNA polymerase I, subunit RPA34.5 / RNA polymerase I, subunit Rpa14, fungi / Nucleic acid-binding, OB-fold / RNA polymerase subunit, RPB6/omega / RNA polymerase Rpb7-like , N-terminal / RNA polymerase, Rpb8 / RPB5-like RNA polymerase subunit superfamily / RNA polymerase Rpb5, N-terminal domain superfamily / RNA polymerase Rpb1, domain 3 superfamily / Rpa43, N-terminal ribonucleoprotein (RNP) domain / RPA43, OB domain / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / DNA-directed RNA polymerase subunit Rpb5-like / RNA polymerase Rpb1, funnel domain superfamily / DNA-directed RNA polymerase, M/15kDa subunit / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb7-like, N-terminal domain superfamily / RNA polymerase Rpb2, domain 2 superfamily / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, RBP11-like 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 / Pol I subunit A12, C-terminal zinc ribbon / RNA polymerase I specific transcription initiation factor RRN3
DNA-directed RNA polymerase I subunit RPA12 / DNA-directed RNA polymerase I subunit RPA14 / DNA-directed RNA polymerase I subunit RPA34 / DNA-directed RNA polymerase I subunit RPA43 / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / RNA polymerase I-specific transcription initiation factor RRN3 / DNA-directed RNA polymerase I subunit RPA190 / DNA-directed RNA polymerases I and III subunit RPAC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerase I subunit RPA135 ...DNA-directed RNA polymerase I subunit RPA12 / DNA-directed RNA polymerase I subunit RPA14 / DNA-directed RNA polymerase I subunit RPA34 / DNA-directed RNA polymerase I subunit RPA43 / DNA-directed RNA polymerases I, II, and III subunit RPABC4 / RNA polymerase I-specific transcription initiation factor RRN3 / DNA-directed RNA polymerase I subunit RPA190 / DNA-directed RNA polymerases I and III subunit RPAC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC5 / DNA-directed RNA polymerase I subunit RPA135 / DNA-directed RNA polymerases I, II, and III subunit RPABC3 / DNA-directed RNA polymerases I, II, and III subunit RPABC2 / DNA-directed RNA polymerases I, II, and III subunit RPABC1 / DNA-directed RNA polymerases I and III subunit RPAC1 / DNA-directed RNA polymerase I subunit RPA49
Biological speciesSaccharomyces cerevisiae (baker's yeast)
Methodsingle particle reconstruction / cryo EM / negative staining / Resolution: 4.8 Å
AuthorsEngel C / Plitzko J / Cramer P
CitationJournal: Nat Commun / Year: 2016
Title: RNA polymerase I-Rrn3 complex at 4.8 Å resolution.
Authors: Christoph Engel / Jürgen Plitzko / Patrick Cramer /
Abstract: Transcription of ribosomal DNA by RNA polymerase I (Pol I) requires the initiation factor Rrn3. Here we report the cryo-EM structure of the Pol I-Rrn3 complex at 4.8 Å resolution. The structure ...Transcription of ribosomal DNA by RNA polymerase I (Pol I) requires the initiation factor Rrn3. Here we report the cryo-EM structure of the Pol I-Rrn3 complex at 4.8 Å resolution. The structure reveals how Rrn3 binding converts an inactive Pol I dimer into an initiation-competent monomeric complex and provides insights into the mechanisms of Pol I-specific initiation and regulation.
Validation ReportPDB-ID: 5g5l

SummaryFull reportAbout validation report
History
Current status-Processing site: PDBe / Status: Released
DepositionMay 25, 2016-
Header (metadata) releaseJul 27, 2016-
Map releaseJul 27, 2016-
UpdateJul 27, 2016-

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.023
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.023
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-5g5l
  • Surface level: 0.023
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_3439.map.gz / Format: CCP4 / Size: 51.5 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.35 Å/pix.
x 240 pix.
= 324. Å
1.35 Å/pix.
x 240 pix.
= 324. Å
1.35 Å/pix.
x 240 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 LevelBy AUTHOR: 0.023 / Movie #1: 0.023
Minimum - Maximum-0.03415515 - 0.0790224
Average (Standard dev.)-0.00010222 (±0.00440425)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions240240240
Spacing240240240
CellA=B=C: 324.0 Å
α=β=γ: 90.0 °

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
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS240240240
D min/max/mean-0.0340.079-0.000

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Supplemental data

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Sample components

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Entire S. cerevisiae RNA polymerase I bound to the initiation factor Rrn3

EntireName: S. cerevisiae RNA polymerase I bound to the initiation factor Rrn3
Number of components: 2 / Oligomeric State: monomeric
MassTheoretical: 663 kDa / Experimental: 663 kDa / Measured by: Size exclusion chromatography

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Component #1: protein, RNA polymerase I

ProteinName: RNA polymerase I / a.k.a: Pol IDNA polymerase I / Oligomeric Details: monomer / Number of Copies: 1 / Recombinant expression: No
MassTheoretical: 590 kDa / Experimental: 590 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast) / Strain: CB010
Source (natural)Location in cell: Nucleus
External referencesGene Ontology: RNA polymerase I complex

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Component #2: protein, RNA polymerase I-specific transcription initiation facto...

ProteinName: RNA polymerase I-specific transcription initiation factor RRN3
a.k.a: Rrn3 / Oligomeric Details: 1 / Recombinant expression: Yes / Number of Copies: 1
MassTheoretical: 73 kDa / Experimental: 73 kDa
SourceSpecies: Saccharomyces cerevisiae (baker's yeast) / Strain: CB010
Source (engineered)Expression System: Escherichia coli (E. coli) / Vector: pET28 / Strain: BL21(DE3)RIL
Source (natural)Location in cell: Nucleus
External referencesInterPro: RNA polymerase I specific transcription initiation factor RRN3
UniProt: RNA polymerase I-specific transcription initiation factor RRN3

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Experimental details

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Sample preparation

SpecimenSpecimen state: Particle / Method: negative staining, cryo EM
Sample solutionSpecimen conc.: 0.1 mg/mL
Buffer solution: 150mM NaCl, 5mM Hepes, 1mM MgCl2, 0.01 mM ZnCl2, 5 mM DTT
pH: 7.8
Support filmR2/1 holey carbon grids (Quantifoil)
Stainingno staining
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 %
Method: 4.5 microliters of sample was applied to glow-discharged Quantifoil R 2/1 holey carbon grids, which were then blotted for 8.5s and plunge-frozen in liquid ethane

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS / Date: Mar 27, 2015
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 40 e/Å2 / Electron beam tilt params: 0 / Illumination mode: FLOOD BEAM
LensMagnification: 37000 X (nominal), 37037 X (calibrated) / Cs: 2.7 mm / Imaging mode: BRIGHT FIELD / Defocus: 800 - 3600 nm / Energy filter: GIF Quantum / Energy window: 0-20 eV
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 1174
Details: Movies with 33 frames were collected over 9.9 seconds

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 63445
3D reconstructionAlgorithm: RELION / Software: RELION / CTF correction: each particle / Resolution: 4.8 Å / Resolution method: FSC 0.143, semi-independent

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Atomic model buiding

Modeling #1Software: Chimera, COOT / Refinement protocol: rigid body / Refinement space: REAL
Details: Initial placement was performed in Chimera. Domains as specified in the PDB file were fitted as rigid bodies an hinges regularized in COOT.
Input PDB model: 4C2M
Chain ID: A, B, C, D, E, F, G, H, I, J, K, L, M, N

Overall bvalue: 165
Modeling #2Software: Chimera, COOT / Refinement protocol: rigid body / Refinement space: REAL
Details: After initial placement in Chimera, secondary structure elements were rigid body fittet in COOT and hinges regularized.
Input PDB model: 3TJ1
Chain ID: A

Overall bvalue: 165
Output model

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