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- PDB-8cro: Cryo-EM structure of Pyrococcus furiosus transcription elongation... -

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Basic information

Entry
Database: PDB / ID: 8cro
TitleCryo-EM structure of Pyrococcus furiosus transcription elongation complex
Components
  • (DNA-directed RNA polymerase subunit ...Polymerase) x 11
  • DNA Non-Template Strand
  • DNA Template Strand
  • RNA
KeywordsTRANSCRIPTION / RNA / Polymerase / Elongation / Complex / Pyrococcus furiosus / Archaea / DNA
Function / homology
Function and homology information


DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / nucleotide binding / DNA-templated transcription / magnesium ion binding / DNA binding ...DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / nucleotide binding / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / cytoplasm
Similarity search - Function
DNA-directed RNA polymerase subunit Rpo12 / DNA-directed RNA polymerase subunit Rpo1C / DNA-directed RNA polymerase subunit Rpo7 / DNA-directed RNA polymerase subunit Rpo1N / DNA-directed RNA polymerase subunit Rpo2 / DNA-directed RNA polymerase subunit Rpo4 / HRDC domain superfamily / DNA-directed RNA polymerase, subunit E/RPC8 / S1 domain profile. / RNA polymerase subunit Rpb4/RPC9 ...DNA-directed RNA polymerase subunit Rpo12 / DNA-directed RNA polymerase subunit Rpo1C / DNA-directed RNA polymerase subunit Rpo7 / DNA-directed RNA polymerase subunit Rpo1N / DNA-directed RNA polymerase subunit Rpo2 / DNA-directed RNA polymerase subunit Rpo4 / HRDC domain superfamily / DNA-directed RNA polymerase, subunit E/RPC8 / S1 domain profile. / RNA polymerase subunit Rpb4/RPC9 / RNA polymerase Rpb4 / RNA polymerase subunit Rpb7-like / HRDC-like superfamily / RNA polymerase Rpb7-like , N-terminal / RNA polymerase Rpb7-like, N-terminal domain superfamily / SHS2 domain found in N terminus of Rpb7p/Rpc25p/MJ0397 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 4 / RNA polymerase Rpb2, domain 5 / RNA polymerase Rpb2, domain 5 / DNA-directed RNA polymerase subunit RPABC5/Rpb10 / RNA polymerases, subunit N, zinc binding site / RNA polymerase subunit RPB10 / RNA polymerases N / 8 kDa subunit / RNA polymerases N / 8 Kd subunits signature. / DNA-directed RNA polymerase subunit Rpo11 / DNA directed RNA polymerase, 7 kDa subunit / RNA polymerase archaeal subunit P/eukaryotic subunit RPABC4 / RNA polymerase, subunit H/Rpb5, conserved site / RNA polymerases H / 23 Kd subunits signature. / RNA polymerase subunit CX / DNA-directed RNA polymerase Rpb11, 13-16kDa subunit, conserved site / DNA-directed RNA polymerase subunit Rpo5/Rpb5 / RNA polymerases L / 13 to 16 Kd subunits signature. / DNA-directed RNA polymerase, 30-40kDa subunit, conserved site / DNA-directed RNA polymerase subunit Rpo3/Rpb3/RPAC1 / RNA polymerases D / 30 to 40 Kd subunits signature. / DNA-directed RNA polymerase, RBP11-like dimerisation domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerase, subunit H/Rpb5 C-terminal / RNA polymerase subunit RPABC4/transcription elongation factor Spt4 / RPB5-like RNA polymerase subunit superfamily / RNA polymerase Rpb5, C-terminal domain / Archaeal Rpo6/eukaryotic RPB6 RNA polymerase subunit / DNA-directed RNA polymerase, 14-18kDa subunit, conserved site / RNA polymerases K / 14 to 18 Kd subunits signature. / Ribosomal protein S1-like RNA-binding domain / S1 RNA binding domain / S1 domain / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb1, domain 3 superfamily / RNA polymerase Rpb1, clamp domain superfamily / RPB6/omega subunit-like superfamily / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 4 / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 5 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / DNA-directed RNA polymerase, insert domain / DNA-directed RNA polymerase, RpoA/D/Rpb3-type / RNA polymerase Rpb3/RpoA insert domain / RNA polymerase Rpb3/Rpb11 dimerisation domain / RNA polymerases D / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, RBP11-like subunit / RNA polymerase Rpb2, domain 2 / RNA polymerase Rpb2, domain 2 / RNA polymerase, beta subunit, conserved site / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerase Rpb2, OB-fold / RNA polymerase Rpb2, domain 7 / RNA polymerase Rpb2, domain 3 / RNA polymerases beta chain signature. / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain / DNA-directed RNA polymerase, subunit 2 / DNA-directed RNA polymerase, subunit 2, hybrid-binding domain superfamily / RNA polymerase Rpb2, domain 6 / Nucleic acid-binding, OB-fold
Similarity search - Domain/homology
DNA / DNA (> 10) / RNA / DNA-directed RNA polymerase subunit Rpo1N / DNA-directed RNA polymerase subunit Rpo10 / DNA-directed RNA polymerase subunit Rpo12 / DNA-directed RNA polymerase subunit Rpo3 / DNA-directed RNA polymerase subunit Rpo6 / DNA-directed RNA polymerase subunit Rpo5 / DNA-directed RNA polymerase subunit beta ...DNA / DNA (> 10) / RNA / DNA-directed RNA polymerase subunit Rpo1N / DNA-directed RNA polymerase subunit Rpo10 / DNA-directed RNA polymerase subunit Rpo12 / DNA-directed RNA polymerase subunit Rpo3 / DNA-directed RNA polymerase subunit Rpo6 / DNA-directed RNA polymerase subunit Rpo5 / DNA-directed RNA polymerase subunit beta / DNA-directed RNA polymerase subunit Rpo1C / DNA-directed RNA polymerase subunit Rpo4 / DNA-directed RNA polymerase subunit Rpo7 / DNA-directed RNA polymerase subunit Rpo11
Similarity search - Component
Biological speciesPyrococcus furiosus DSM 3638 (archaea)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsTarau, D.M. / Grunberger, F. / Reichelt, R. / Heiss, F.B. / Pilsl, M. / Hausner, W. / Engel, C. / Grohmann, D.
Funding support Germany, 1items
OrganizationGrant numberCountry
German Research Foundation (DFG)SFB960 Germany
CitationJournal: Nucleic Acids Res. / Year: 2024
Title: Structural basis of archaeal RNA polymerase transcription elongation and Spt4/5 recruitment
Authors: Tarau, D.M. / Grunberger, F. / Pilsl, M. / Reichelt, R. / Heiss, F.B. / Koenig, S. / Urlaub, H. / Hausner, W. / Engel, C. / Grohmann, D.
History
DepositionMar 8, 2023Deposition site: PDBE / Processing site: PDBE
Revision 1.0Apr 17, 2024Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: DNA-directed RNA polymerase subunit Rpo1N
B: DNA-directed RNA polymerase subunit beta
C: DNA-directed RNA polymerase subunit Rpo1C
D: DNA-directed RNA polymerase subunit Rpo3
E: DNA-directed RNA polymerase subunit Rpo7
F: DNA-directed RNA polymerase subunit Rpo4
L: DNA-directed RNA polymerase subunit Rpo11
H: DNA-directed RNA polymerase subunit Rpo5
N: DNA-directed RNA polymerase subunit Rpo10
K: DNA-directed RNA polymerase subunit Rpo6
P: DNA-directed RNA polymerase subunit Rpo12
X: DNA Template Strand
Y: DNA Non-Template Strand
Z: RNA
hetero molecules


Theoretical massNumber of molelcules
Total (without water)393,55720
Polymers393,20514
Non-polymers3516
Water0
1


  • Idetical with deposited unit
  • defined by author
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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DNA-directed RNA polymerase subunit ... , 11 types, 11 molecules ABCDEFLHNKP

#1: Protein DNA-directed RNA polymerase subunit Rpo1N / Polymerase / DNA-directed RNA polymerase subunit A'


Mass: 103612.250 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpoA1, rpo1N, PFDSM3638_07875 / Production host: Pyrococcus furiosus DSM 3638 (archaea)
References: UniProt: A0A5C0XPL7, DNA-directed RNA polymerase
#2: Protein DNA-directed RNA polymerase subunit beta / Polymerase


Mass: 127188.898 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: PF1564 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8U0M3
#3: Protein DNA-directed RNA polymerase subunit Rpo1C / Polymerase / DNA-directed RNA polymerase subunit A''


Mass: 44468.156 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo1C, rpoA2, PF1562 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8U0M5, DNA-directed RNA polymerase
#4: Protein DNA-directed RNA polymerase subunit Rpo3 / Polymerase / DNA-directed RNA polymerase subunit D


Mass: 29817.182 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo3, rpoD, PF1647 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8U0E4, DNA-directed RNA polymerase
#5: Protein DNA-directed RNA polymerase subunit Rpo7 / Polymerase


Mass: 21731.455 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo7 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8U439
#6: Protein DNA-directed RNA polymerase subunit Rpo4 / Polymerase / DNA-directed RNA polymerase subunit F


Mass: 14116.311 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo4, rpoF, PF1036 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8U216, DNA-directed RNA polymerase
#7: Protein DNA-directed RNA polymerase subunit Rpo11 / Polymerase / DNA-directed RNA polymerase subunit L


Mass: 11132.764 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo11, rpoL, PF0050 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8U4N1, DNA-directed RNA polymerase
#8: Protein DNA-directed RNA polymerase subunit Rpo5 / Polymerase / DNA-directed RNA polymerase subunit H


Mass: 9260.792 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo5, rpoH, PF1565 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8U0M2, DNA-directed RNA polymerase
#9: Protein DNA-directed RNA polymerase subunit Rpo10 / Polymerase / DNA-directed RNA polymerase subunit N


Mass: 7800.158 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo10, rpoN, PF1643 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: P60292, DNA-directed RNA polymerase
#10: Protein DNA-directed RNA polymerase subunit Rpo6 / Polymerase / DNA-directed RNA polymerase subunit K


Mass: 6243.490 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo6, rpoK, PF1642 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8U0E8, DNA-directed RNA polymerase
#11: Protein/peptide DNA-directed RNA polymerase subunit Rpo12 / Polymerase / DNA-directed RNA polymerase subunit P


Mass: 5770.050 Da / Num. of mol.: 1 / Mutation: 0
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus DSM 3638 (archaea) / Gene: rpo12, rpoP, PF2009 / Production host: Pyrococcus furiosus DSM 3638 (archaea) / References: UniProt: Q8TZI3, DNA-directed RNA polymerase

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DNA chain , 2 types, 2 molecules XY

#12: DNA chain DNA Template Strand


Mass: 6190.001 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Pyrococcus furiosus DSM 3638 (archaea)
#13: DNA chain DNA Non-Template Strand


Mass: 2963.969 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Pyrococcus furiosus DSM 3638 (archaea)

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RNA chain , 1 types, 1 molecules Z

#14: RNA chain RNA /


Mass: 2909.823 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Pyrococcus furiosus DSM 3638 (archaea)

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Non-polymers , 2 types, 6 molecules

#15: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 5 / Source method: obtained synthetically / Formula: Zn
#16: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg / Feature type: SUBJECT OF INVESTIGATION

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Details

Has ligand of interestY

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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
IDNameTypeEntity IDParent-IDSource
1Transcription Elongation Complex (TEC)COMPLEX#1-#140MULTIPLE SOURCES
2Pyrococcus furiosus RNA polymeraseCOMPLEX#1-#111RECOMBINANT
3Nucleotide strandsCOMPLEX#12-#141RECOMBINANT
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Pyrococcus furiosus DSM 3638 (archaea)186497
33Pyrococcus furiosus DSM 3638 (archaea)186497
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-ID
22Pyrococcus furiosus DSM 3638 (archaea)186497
33synthetic construct (others)32630
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid type: UltrAuFoil R1.2/1.3
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 % / Chamber temperature: 277.15 K

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: OTHER
Electron lensMode: OTHER / Nominal defocus max: 2000 nm / Nominal defocus min: 1200 nm
Image recordingElectron dose: 88.73 e/Å2 / Detector mode: INTEGRATING / Film or detector model: FEI FALCON III (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.20.1_4487: / Classification: refinement
EM software
IDNameVersionCategory
7Coot0.9.8.1model fitting
9RELION3.1initial Euler assignment
10RELION4final Euler assignment
11RELION4classification
12RELION3D reconstruction
13PHENIX1.20.1-4487model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 333535 / Symmetry type: POINT
Atomic model buildingProtocol: AB INITIO MODEL / Space: REAL
Atomic model buildingAccession code: 6kf9 / Source name: SwissModel / Type: in silico model
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00327766
ELECTRON MICROSCOPYf_angle_d0.52737660
ELECTRON MICROSCOPYf_dihedral_angle_d10.2374048
ELECTRON MICROSCOPYf_chiral_restr0.0424186
ELECTRON MICROSCOPYf_plane_restr0.0044745

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