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- PDB-5vt0: Escherichia coli 6S RNA derivative in complex with Escherichia co... -

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Basic information

Entry
Database: PDB / ID: 5vt0
TitleEscherichia coli 6S RNA derivative in complex with Escherichia coli RNA polymerase sigma70-holoenzyme
DescriptorEscherichia coli sigma 70 (rpoD)/RNA Complex
(Escherichia coli RNA polymerase ...) x 4
Keywordstranscription/RNA / RNAP / 6S RNA / ncRNA / transcription regulation / transcription-RNA complex
Specimen sourceEscherichia coli (strain k12) / bacteria /
Escherichia coli / bacteria / エシェリキア・コリ, 大腸菌 /
MethodElectron microscopy (3.78 Å resolution / Particle / Single particle)
AuthorsChen, J. / Darst, S.A.
CitationMol. Cell, 2017, 68, 388-397.e6

Mol. Cell, 2017, 68, 388-397.e6 Yorodumi Papers
6S RNA Mimics B-Form DNA to Regulate Escherichia coli RNA Polymerase.
James Chen / Karen M Wassarman / Shili Feng / Katherine Leon / Andrey Feklistov / Jared T Winkelman / Zongli Li / Thomas Walz / Elizabeth A Campbell / Seth A Darst

Validation Report
SummaryFull reportAbout validation report
DateDeposition: May 15, 2017 / Release: Nov 1, 2017

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Assembly

Deposited unit
G: DNA-directed RNA polymerase subunit alpha
H: DNA-directed RNA polymerase subunit alpha
I: DNA-directed RNA polymerase subunit beta
J: DNA-directed RNA polymerase subunit beta'
K: DNA-directed RNA polymerase subunit omega
L: RNA polymerase sigma factor RpoD
R: Escherichia coli 6S RNA derivative (112-MER)
hetero molecules


Theoretical massNumber of molelcules
Total (without water)476,02710
Polyers475,8727
Non-polymers1553
Water0
#1


TypeNameSymmetry operationNumber
identity operation1_555x,y,z1
Buried area (Å2)47140
ΔGint (kcal/M)-246
Surface area (Å2)174630
MethodPISA

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Components

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DNA-directed RNA polymerase subunit ... , 4 types, 5 molecules GHIJK

#1: Polypeptide(L)DNA-directed RNA polymerase subunit alpha / RNAP subunit alpha / RNA polymerase subunit alpha / Transcriptase subunit alpha


Mass: 26330.996 Da / Num. of mol.: 2 / Source: (gene. exp.) Escherichia coli (strain K12) / References: UniProt: P0A7Z4, EC: 2.7.7.6
#2: Polypeptide(L)DNA-directed RNA polymerase subunit beta / RNAP subunit beta / RNA polymerase subunit beta / Transcriptase subunit beta


Mass: 150820.875 Da / Num. of mol.: 1 / Source: (gene. exp.) Escherichia coli (strain K12) / References: UniProt: P0A8V2, EC: 2.7.7.6
#3: Polypeptide(L)DNA-directed RNA polymerase subunit beta' / RNAP subunit beta' / RNA polymerase subunit beta' / Transcriptase subunit beta'


Mass: 155366.781 Da / Num. of mol.: 1 / Source: (gene. exp.) Escherichia coli (strain K12) / References: UniProt: P0A8T7, EC: 2.7.7.6
#4: Polypeptide(L)DNA-directed RNA polymerase subunit omega / RNAP omega subunit / RNA polymerase omega subunit / Transcriptase subunit omega


Mass: 10249.547 Da / Num. of mol.: 1 / Source: (gene. exp.) Escherichia coli (strain K12) / References: UniProt: P0A800, EC: 2.7.7.6

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Polypeptide(L) / RNA chain , 2 types, 2 molecules LR

#5: Polypeptide(L)RNA polymerase sigma factor RpoD / Sigma-70


Mass: 60464.371 Da / Num. of mol.: 1 / Source: (gene. exp.) Escherichia coli (strain K12) / References: UniProt: P00579
#6: RNA chainEscherichia coli 6S RNA derivative (112-MER)


Mass: 46308.500 Da / Num. of mol.: 1 / Details: prepared by in vitro transcription / Source: (synth.) Escherichia coli

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Non-polymers , 2 types, 3 molecules

#7: ChemicalChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Formula: Mg
#8: ChemicalChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 2 / Formula: Zn

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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

ComponentName: Escherichia coli 6S RNA derivative in complex with Escherichia coli RNA polymerase sigma70-holoenzyme
Type: COMPLEX / Entity ID: 1, 2, 3, 4, 5, 6 / Source: RECOMBINANT
Molecular weightValue: 0.475 deg. / Units: MEGADALTONS / Experimental value: YES
Source (natural)Organism: Escherichia coli
Source (recombinant)Organism: Escherichia coli BL21(DE3)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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: 1.8 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.12_2829: / Classification: refinement
EM software
IDNameVersionCategoryDetailsFitting ID
7Coot1.10.10-2155MODEL FITTING0.81
9PHENIX1.10.1-2155MODEL REFINEMENT1
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
SymmetryPoint symmetry: C1
3D reconstructionResolution: 3.78 Å / Resolution method: FSC 0.143 CUT-OFF / Number of particles: 362926 / Symmetry type: POINT
Atomic model buildingRef protocol: OTHER / Ref space: REAL / Target criteria: Correlation coefficient
Least-squares processHighest resolution: 3.78 Å
Refine LS restraints
Refine IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00731911
ELECTRON MICROSCOPYf_angle_d1.02743624
ELECTRON MICROSCOPYf_dihedral_angle_d6.41419425
ELECTRON MICROSCOPYf_chiral_restr0.0615063
ELECTRON MICROSCOPYf_plane_restr0.0075286

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