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- PDB-6pln: X-ray crystal structure of Pyrococcus furiosus general transcript... -

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Basic information

Entry
Database: PDB / ID: 6pln
TitleX-ray crystal structure of Pyrococcus furiosus general transcription factor TFE-alpha
ComponentsTranscription factor E
KeywordsTRANSCRIPTION / TFE / general transcription factor
Function / homology
Function and homology information


transcription initiation at RNA polymerase II promoter / regulation of DNA-templated transcription / DNA binding
Similarity search - Function
Transcription factor TFE, archaea / Transcription initiation factor IIE subunit alpha, N-terminal / Transcription factor TFE/TFIIEalpha HTH domain / TFIIEalpha/SarR/Rpc3 HTH domain / Transcription factor E / TFIIE alpha subunit / TFE/IIEalpha-type HTH domain profile. / Transcription initiation factor IIE / Winged helix DNA-binding domain superfamily / Winged helix-like DNA-binding domain superfamily
Similarity search - Domain/homology
Transcription factor E
Similarity search - Component
Biological speciesPyrococcus furiosus (archaea)
MethodX-RAY DIFFRACTION / SYNCHROTRON / SAD / Resolution: 2.6 Å
AuthorsMurakami, K.S. / Jun, S.H.
Funding support United States, 1items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)R35 GM131860 United States
CitationJournal: Nat Commun / Year: 2020
Title: Direct binding of TFEα opens DNA binding cleft of RNA polymerase.
Authors: Sung-Hoon Jun / Jaekyung Hyun / Jeong Seok Cha / Hoyoung Kim / Michael S Bartlett / Hyun-Soo Cho / Katsuhiko S Murakami /
Abstract: Opening of the DNA binding cleft of cellular RNA polymerase (RNAP) is necessary for transcription initiation but the underlying molecular mechanism is not known. Here, we report on the cryo-electron ...Opening of the DNA binding cleft of cellular RNA polymerase (RNAP) is necessary for transcription initiation but the underlying molecular mechanism is not known. Here, we report on the cryo-electron microscopy structures of the RNAP, RNAP-TFEα binary, and RNAP-TFEα-promoter DNA ternary complexes from archaea, Thermococcus kodakarensis (Tko). The structures reveal that TFEα bridges the RNAP clamp and stalk domains to open the DNA binding cleft. Positioning of promoter DNA into the cleft closes it while maintaining the TFEα interactions with the RNAP mobile modules. The structures and photo-crosslinking results also suggest that the conserved aromatic residue in the extended winged-helix domain of TFEα interacts with promoter DNA to stabilize the transcription bubble. This study provides a structural basis for the functions of TFEα and elucidates the mechanism by which the DNA binding cleft is opened during transcription initiation in the stalk-containing RNAPs, including archaeal and eukaryotic RNAPs.
History
DepositionJul 1, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jul 8, 2020Provider: repository / Type: Initial release
Revision 1.1Jan 13, 2021Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year
Revision 1.2Mar 13, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Transcription factor E
B: Transcription factor E


Theoretical massNumber of molelcules
Total (without water)45,8572
Polymers45,8572
Non-polymers00
Water32418
1
A: Transcription factor E


Theoretical massNumber of molelcules
Total (without water)22,9281
Polymers22,9281
Non-polymers00
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
2
B: Transcription factor E


Theoretical massNumber of molelcules
Total (without water)22,9281
Polymers22,9281
Non-polymers00
Water181
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
MethodPISA
Unit cell
Length a, b, c (Å)55.689, 55.689, 248.673
Angle α, β, γ (deg.)90.000, 90.000, 90.000
Int Tables number92
Space group name H-MP41212
Space group name HallP4abw2nw
Symmetry operation#1: x,y,z
#2: -y+1/2,x+1/2,z+1/4
#3: y+1/2,-x+1/2,z+3/4
#4: x+1/2,-y+1/2,-z+3/4
#5: -x+1/2,y+1/2,-z+1/4
#6: -x,-y,z+1/2
#7: y,x,-z
#8: -y,-x,-z+1/2

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Components

#1: Protein Transcription factor E / TFE / TFIIE subunit alpha homolog / Transcription initiation factor TFIIE


Mass: 22928.322 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Pyrococcus furiosus (archaea) / Gene: tfe, PF0491 / Production host: Escherichia coli (E. coli) / References: UniProt: Q8U3H5
#2: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 18 / Source method: isolated from a natural source / Formula: H2O

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Experimental details

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Experiment

ExperimentMethod: X-RAY DIFFRACTION / Number of used crystals: 1

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Sample preparation

CrystalDensity Matthews: 2.1 Å3/Da / Density % sol: 41.49 %
Crystal growTemperature: 300 K / Method: vapor diffusion, hanging drop
Details: 10 % PEG 8000, 0.2 M MgCl2, and 0.1 M Tris-HCl (pH 7)

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Data collection

DiffractionMean temperature: 100 K / Serial crystal experiment: N
Diffraction sourceSource: SYNCHROTRON / Site: CHESS / Beamline: A1 / Wavelength: 0.977 Å
DetectorType: ADSC QUANTUM 210 / Detector: CCD / Date: Dec 5, 2013
RadiationProtocol: SINGLE WAVELENGTH / Monochromatic (M) / Laue (L): M / Scattering type: x-ray
Radiation wavelengthWavelength: 0.977 Å / Relative weight: 1
ReflectionResolution: 2.6→40 Å / Num. obs: 13000 / % possible obs: 99.9 % / Redundancy: 13.1 % / Biso Wilson estimate: 66.79 Å2 / CC1/2: 0.842 / Rmerge(I) obs: 0.064 / Χ2: 1.804 / Net I/σ(I): 59
Reflection shellResolution: 2.6→2.64 Å / Redundancy: 12.8 % / Rmerge(I) obs: 0.991 / Mean I/σ(I) obs: 3.55 / Num. unique obs: 645 / CC1/2: 0.997 / Χ2: 1.009 / % possible all: 100

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Processing

Software
NameVersionClassification
PHENIX1.15.2_3472refinement
HKL-2000data reduction
HKL-2000data scaling
PHENIXphasing
Cootmodel building
RefinementMethod to determine structure: SAD / Resolution: 2.6→33.27 Å / SU ML: 0.3422 / Cross valid method: FREE R-VALUE / σ(F): 0 / Phase error: 29.4814
RfactorNum. reflection% reflection
Rfree0.2865 1265 9.95 %
Rwork0.2395 --
obs0.2444 12709 97.89 %
Solvent computationShrinkage radii: 0.9 Å / VDW probe radii: 1.11 Å
Displacement parametersBiso mean: 76.22 Å2
Refinement stepCycle: LAST / Resolution: 2.6→33.27 Å
ProteinNucleic acidLigandSolventTotal
Num. atoms1724 0 0 18 1742
Refine LS restraints
Refine-IDTypeDev idealNumber
X-RAY DIFFRACTIONf_bond_d0.00951746
X-RAY DIFFRACTIONf_angle_d1.12522350
X-RAY DIFFRACTIONf_chiral_restr0.0554266
X-RAY DIFFRACTIONf_plane_restr0.0055296
X-RAY DIFFRACTIONf_dihedral_angle_d8.7321076
LS refinement shell
Resolution (Å)Rfactor RfreeNum. reflection RfreeRfactor RworkNum. reflection RworkRefine-ID% reflection obs (%)
2.6-2.70.35171290.30281156X-RAY DIFFRACTION91.85
2.7-2.820.44921300.35761196X-RAY DIFFRACTION94.38
2.82-2.970.41411340.31951216X-RAY DIFFRACTION97.12
2.97-3.160.32481370.29271252X-RAY DIFFRACTION98.51
3.16-3.40.30831420.27061276X-RAY DIFFRACTION99.58
3.4-3.740.30341440.26261291X-RAY DIFFRACTION100
3.74-4.280.2721460.23851312X-RAY DIFFRACTION99.93
4.28-5.390.23641460.21551307X-RAY DIFFRACTION100
5.39-33.270.26481570.19321438X-RAY DIFFRACTION99.19

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