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- PDB-5n60: Cryo-EM structure of RNA polymerase I in complex with Rrn3 and Co... -

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Entry
Database: PDB / ID: 5n60
TitleCryo-EM structure of RNA polymerase I in complex with Rrn3 and Core Factor (Orientation I)
Components
  • (DNA-directed RNA polymerase I subunit ...) x 7
  • (DNA-directed RNA polymerases I and III subunit ...) x 2
  • (DNA-directed RNA polymerases I, II, and III subunit ...) x 5
  • (RNA polymerase I-specific transcription initiation factor ...) x 4
KeywordsTRANSFERASE / Transferase / Transcription / RNA polymerase I initiation
Function/homologytranscription factor activity, RNA polymerase I transcription factor binding / go:0001180: / RNA polymerase I specific transcription initiation factor RRN3 / transcription factor activity, core RNA polymerase I binding / RNA polymerase I specific transcription initiation factor RRN3 / RNA polymerase I-specific transcription initiation factor Rrn6 / RNA polymerase I transcription initiation factor TAF1B/Rrn7 / Transcription initiation factor Rrn11 / go:0001187: / RNA polymerase I core factor complex ...transcription factor activity, RNA polymerase I transcription factor binding / go:0001180: / RNA polymerase I specific transcription initiation factor RRN3 / transcription factor activity, core RNA polymerase I binding / RNA polymerase I specific transcription initiation factor RRN3 / RNA polymerase I-specific transcription initiation factor Rrn6 / RNA polymerase I transcription initiation factor TAF1B/Rrn7 / Transcription initiation factor Rrn11 / go:0001187: / RNA polymerase I core factor complex / Transcription initiation factor Rrn7, Zinc-finger / RNA polymerase I-specific transcription initiation factor RRN6-like / Transcription initiation factor Rrn11, budding yeast / RNA polymerase transcription factor SL1 complex / RNA polymerase I regulatory region DNA binding / RNA polymerase I-specific transcription-initiation factor / RNA polymerase I specific initiation factor / Zinc-finger of RNA-polymerase I-specific TFIIB, Rrn7 / DNA-directed RNA polymerase I subunit RPA2, domain 4 / Pol I subunit A12, C-terminal zinc ribbon / DNA-directed RNA pol I, largest subunit / RNA polymerase II sequence-specific DNA binding transcription factor binding / RNA polymerase I, subunit Rpa14, fungi / DNA-directed RNA polymerase I, subunit RPA34.5 / RNA polymerase I associated factor, A49-like / RNA polymerase I core binding / RNA polymerase I transcription factor binding / RNA polymerase I CORE element sequence-specific DNA binding / RNA polymerase I, Rpa2 specific domain / DNA-directed RNA polymerases I and III subunit AC19 / DNA-directed RNA polymerases I and III subunit AC40 / Yeast RNA polymerase I subunit RPA14 / DNA-directed RNA polymerase I subunit RPA34.5 / A49-like RNA polymerase I associated factor / transcription initiation from RNA polymerase I promoter / rDNA binding / nucleolar large rRNA transcription by RNA polymerase I / transposon integration / transcription elongation from RNA polymerase I promoter / termination of RNA polymerase I transcription / RNA polymerase Rpb7-like, N-terminal domain superfamily / RNA polymerase Rpb7, N-terminal / termination of RNA polymerase III transcription / RNA Polymerase I Transcription Initiation / Zinc finger TFIIS-type signature. / tRNA transcription by RNA polymerase III / RNA polymerase I activity / Formation of the Early Elongation Complex / RNA Pol II CTD phosphorylation and interaction with CE / mRNA Capping / 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 Promoter Escape / TP53 Regulates Transcription of DNA Repair Genes / DNA-directed RNA polymerase M, 15kDa subunit, conserved site / RNA polymerases M / 15 Kd subunits signature. / DNA-directed RNA polymerase, M/15kDa subunit / DNA-directed RNA polymerase RPB5 subunit, eukaryote/virus / DNA-directed RNA polymerase subunit/transcription factor S / DNA-directed RNA polymerase, subunit RPB6 / RNA polymerase, Rpb5, N-terminal / RNA polymerase Rpb5, N-terminal domain superfamily / RNA polymerase, Rpb8 / RNA polymerase archaeal subunit P/eukaryotic subunit RPABC4 / RNA polymerases N / 8 Kd subunits signature. / RNA polymerases, subunit N, zinc binding site / RNA Polymerase II Pre-transcription Events / DNA-directed RNA polymerase, subunit N/Rpb10 / Zinc finger, TFIIS-type / Zinc finger TFIIS-type profile. / DNA-directed RNA polymerase, 14-18kDa subunit, conserved site / RNA polymerases K / 14 to 18 Kd subunits signature. / RNA polymerase subunit RPB10 / Archaeal RpoK/eukaryotic RPB6 RNA polymerase subunit / promoter-specific chromatin binding / DNA-directed RNA polymerase, 30-40kDa subunit, conserved site / RNA polymerases D / 30 to 40 Kd subunits signature. / transcription by RNA polymerase I / RNA polymerase, subunit H/Rpb5, conserved site / RPB5-like RNA polymerase subunit / RNA polymerases L / 13 to 16 Kd subunits signature. / RNA polymerases H / 23 Kd subunits signature. / Archaeal RpoH /eukaryotic RPB5 RNA polymerase subunit / DNA-directed RNA polymerase Rpb11, 13-16kDa subunit, conserved site / DNA-directed RNA polymerase, RBP11-like dimerisation domain / RNA polymerase, subunit H/Rpb5 C-terminal / DNA-directed RNA polymerase III complex / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / DNA-directed RNA polymerase I complex / transcription by RNA polymerase III / DNA-directed RNA polymerase II, core complex / mRNA cleavage / transcription, RNA-templated / SHS2 domain found in N terminus of Rpb7p/Rpc25p/MJ0397 / mRNA Splicing - Minor Pathway / Formation of TC-NER Pre-Incision Complex / RNA polymerase subunit, RPB6/omega / RNA polymerase, subunit omega/K/RPB6 / RPB6/omega subunit-like superfamily
Function and homology information
Specimen sourceSaccharomyces cerevisiae / yeast / /
MethodElectron microscopy (7.7 Å resolution / Particle / Single particle) / Transmission electron microscopy
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.
Copyright: 2017 Elsevier Inc. All rights reserved.
Validation Report
SummaryFull reportAbout validation report
DateDeposition: Feb 14, 2017 / Release: Apr 5, 2017
RevisionDateData content typeGroupCategoryItemProviderType
1.0Apr 5, 2017Structure modelrepositoryInitial release
1.1Apr 12, 2017Structure modelStructure summary
1.2Sep 6, 2017Structure modelAuthor supporting evidence / Data collectionem_image_scans / pdbx_audit_support_pdbx_audit_support.funding_organization

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Assembly

Deposited unit
A: DNA-directed RNA polymerase I subunit RPA190
B: DNA-directed RNA polymerase I subunit RPA135
C: DNA-directed RNA polymerases I and III subunit RPAC1
D: DNA-directed RNA polymerase I subunit RPA14
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 I subunit RPA43
H: DNA-directed RNA polymerases I, II, and III subunit RPABC3
I: DNA-directed RNA polymerase I subunit RPA12
J: DNA-directed RNA polymerases I, II, and III subunit RPABC5
K: DNA-directed RNA polymerases I and III subunit RPAC2
L: DNA-directed RNA polymerases I, II, and III subunit RPABC4
M: DNA-directed RNA polymerase I subunit RPA49
N: DNA-directed RNA polymerase I subunit RPA34
O: RNA polymerase I-specific transcription initiation factor RRN3
P: RNA polymerase I-specific transcription initiation factor RRN6
Q: RNA polymerase I-specific transcription initiation factor RRN7
R: RNA polymerase I-specific transcription initiation factor RRN11
hetero molecules


Theoretical massNumber of molelcules
Total (without water)885,19326
Polyers884,66918
Non-polymers5238
Water0
1


TypeNameSymmetry operationNumber
identity operation1_5551
Buried area (Å2)97520
ΔGint (kcal/M)-484
Surface area (Å2)253250
MethodPISA

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Components

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DNA-directed RNA polymerase I subunit ... , 7 types, 7 molecules ABDGIMN

#1: Protein/peptide DNA-directed RNA polymerase I subunit RPA190 / DNA-directed RNA polymerase I 190 kDa polypeptide / A190 / DNA-directed RNA polymerase I largest subunit


Mass: 186676.969 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:P10964, EC:2.7.7.6 (DNA-directed RNA polymerase)
#2: Protein/peptide DNA-directed RNA polymerase I subunit RPA135 / DNA-directed RNA polymerase I 135 kDa polypeptide / A135 / DNA-directed RNA polymerase I polypeptide 2 / RNA polymerase I subunit 2


Mass: 135910.328 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:P22138, EC:2.7.7.6 (DNA-directed RNA polymerase)
#4: Protein/peptide DNA-directed RNA polymerase I subunit RPA14 / A14 / DNA-directed RNA polymerase I 14 kDa polypeptide


Mass: 14599.128 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:P50106
#7: Protein/peptide DNA-directed RNA polymerase I subunit RPA43 / A43 / DNA-directed DNA-dependent RNA polymerase 36 kDa polypeptide


Mass: 36264.852 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:P46669
#9: Protein/peptide DNA-directed RNA polymerase I subunit RPA12 / A12 / A12.2 / DNA-directed RNA polymerase I 13.7 kDa polypeptide


Mass: 13676.566 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:P32529
#13: Protein/peptide DNA-directed RNA polymerase I subunit RPA49 / A49 / DNA-directed RNA polymerase I 49 kDa polypeptide


Mass: 46721.707 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:Q01080
#14: Protein/peptide DNA-directed RNA polymerase I subunit RPA34 / A34 / DNA-directed DNA-dependent RNA polymerase 34.5 kDa polypeptide / A34.5


Mass: 26933.518 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:P47006

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DNA-directed RNA polymerases I and III subunit ... , 2 types, 2 molecules CK

#3: Protein/peptide DNA-directed RNA polymerases I and III subunit RPAC1 / RNA polymerases I and III subunit AC1 / C37 / DNA-directed RNA polymerases I and III 40 kDa polypeptide / C40


Mass: 37732.613 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:P07703
#11: Protein/peptide DNA-directed RNA polymerases I and III subunit RPAC2 / RNA polymerases I and III subunit AC2 / AC19 / DNA-directed RNA polymerases I and III 16 kDa polypeptide / RPA19


Mass: 16167.860 Da / Num. of mol.: 1
Source: (natural) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
References: UniProt:P28000

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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 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)
References: UniProt:P20434
#6: Protein/peptide DNA-directed RNA polymerases I, II, and III subunit RPABC2 / 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)
References: UniProt:P20435
#8: Protein/peptide DNA-directed RNA polymerases I, II, and III subunit RPABC3 / 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)
References: UniProt:P20436
#10: Protein/peptide DNA-directed RNA polymerases I, II, and III subunit RPABC5 / 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)
References: UniProt:P22139
#12: Protein/peptide DNA-directed RNA polymerases I, II, and III subunit RPABC4 / 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)
References: UniProt:P40422

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RNA polymerase I-specific transcription initiation factor ... , 4 types, 4 molecules OPQR

#15: Protein/peptide RNA polymerase I-specific transcription initiation factor RRN3


Mass: 72458.258 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
Gene: RRN3, YKL125W / Production host: Escherichia coli BL21(DE3) / References: UniProt:P36070
#16: Protein/peptide RNA polymerase I-specific transcription initiation factor RRN6


Mass: 102163.227 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
Gene: RRN6, YBL014C, YBL0311, YBL0312 / Production host: Escherichia coli BL21(DE3) / References: UniProt:P32786
#17: Protein/peptide RNA polymerase I-specific transcription initiation factor RRN7


Mass: 60435.195 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
Gene: RRN7, YJL025W, J1273 / Production host: Escherichia coli BL21(DE3) / References: UniProt:P40992
#18: Protein/peptide RNA polymerase I-specific transcription initiation factor RRN11


Mass: 59334.262 Da / Num. of mol.: 1
Source: (gene. exp.) Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
Gene: RRN11, YML043C, YM9827.09C / Production host: Escherichia coli BL21(DE3) / References: UniProt:Q04712

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Non-polymers , 1 types, 8 molecules

#19: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 8 / Formula: Zn / : Zinc

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / Reconstruction method: SINGLE PARTICLE

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Sample preparation

Component
IDNameTypeEntity IDParent IDSource
1RNA polymerase I in complex with Rrn3 and Core FactorCOMPLEX1,2,3,4,5,6,7,8,9,10,11,12,13,14,15,16,17,180NATURAL
2RNA polymerase ICOMPLEX1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 141NATURAL
3Initiation factor Rrn3COMPLEX151RECOMBINANT
4Core FactorCOMPLEX16, 17, 181RECOMBINANT
Molecular weight
IDValueUnitsEntity assembly IDExperimental value
10.59MEGADALTONS2NO
20.07MEGADALTONS3NO
30.22MEGADALTONS4NO
Source (natural)
IDEntity assembly IDNcbi tax IDOrganism
12559292Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
23559292Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
34559292Saccharomyces cerevisiae (strain ATCC 204508 / S288c)
Source (recombinant)
IDEntity assembly IDNcbi tax IDOrganism
13469008Escherichia coli BL21(DE3)
24469008Escherichia coli BL21(DE3)
Buffer solutionpH: 7.8
SpecimenConc.: 0.1 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 298 kelvins

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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: FLOOD BEAM
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 40 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 7.7 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 8317 / Symmetry type: POINT
Atomic model buildingRef protocol: RIGID BODY FIT

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