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- PDB-7ye1: The cryo-EM structure of C. crescentus GcrA-TACup -

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Basic information

Entry
Database: PDB / ID: 7ye1
TitleThe cryo-EM structure of C. crescentus GcrA-TACup
Components
  • (DNA (57-MER)- ...) x 2
  • (DNA-directed RNA polymerase subunit ...) x 4
  • Cell cycle regulatory protein GcrA
  • RNA polymerase sigma factor RpoD
KeywordsTRANSCRIPTION / transcription activator
Function / homology
Function and homology information


sigma factor activity / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed RNA polymerase / DNA-directed RNA polymerase activity / protein dimerization activity / DNA-templated transcription / magnesium ion binding / DNA binding ...sigma factor activity / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / DNA-directed RNA polymerase / DNA-directed RNA polymerase activity / protein dimerization activity / DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / metal ion binding / cytoplasm
Similarity search - Function
GcrA cell cycle regulator / GcrA cell cycle regulator / RNA polymerase sigma factor 70, non-essential domain / Sigma-70, non-essential region / RNA polymerase sigma factor 70, region 1.1 / Sigma-70 factor, region 1.1 superfamily / Sigma-70 factor, region 1.1 / RNA polymerase II, Rpb2 subunit, wall domain / RNA polymerase subunit, RPB6/omega / Eukaryotic RPB6 RNA polymerase subunit ...GcrA cell cycle regulator / GcrA cell cycle regulator / RNA polymerase sigma factor 70, non-essential domain / Sigma-70, non-essential region / RNA polymerase sigma factor 70, region 1.1 / Sigma-70 factor, region 1.1 superfamily / Sigma-70 factor, region 1.1 / RNA polymerase II, Rpb2 subunit, wall domain / RNA polymerase subunit, RPB6/omega / Eukaryotic RPB6 RNA polymerase subunit / RNA polymerase Rpb1 funnel domain / Sigma-70 factors family signature 1. / Topoisomerase I; Chain A, domain 4 / DNA-directed RNA polymerase, insert domain / RNA polymerase sigma factor RpoD, C-terminal / RNA polymerase sigma factor RpoD / RNA Polymerase Alpha Subunit; Chain A, domain 2 / : / RNA polymerase sigma-70 region 1.2 / Sigma-70 factor, region 1.2 / RNA polymerase sigma-70 region 3 / Sigma-70 region 3 / Sigma-70 factors family signature 2. / RNA polymerase sigma-70 / RNA polymerase sigma-70 region 4 / Sigma-70, region 4 / RNA polymerase sigma-70 region 2 / RNA polymerase sigma-70 like domain / Sigma-70 region 2 / RNA polymerase sigma factor, region 2 / RNA polymerase sigma factor, region 3/4-like / DNA-directed RNA polymerase, omega subunit / DNA-directed RNA polymerase, subunit beta-prime, bacterial type / DNA-directed RNA polymerase, beta subunit, external 1 domain superfamily / DNA-directed RNA polymerase, beta subunit, external 1 domain / RNA polymerase beta subunit external 1 domain / RNA polymerase, alpha subunit, C-terminal / Bacterial RNA polymerase, alpha chain C terminal domain / DNA-directed RNA polymerase beta subunit, bacterial-type / DNA-directed RNA polymerase, alpha subunit / Beta Complex / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 3 superfamily / RPB6/omega subunit-like superfamily / 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 / RNA polymerase Rpb1, clamp domain superfamily / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / RNA polymerase, alpha subunit / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / RNA polymerase Rpb1, domain 2 / RNA polymerase Rpb1, domain 5 / RNA polymerase Rpb1, domain 4 / DNA-directed RNA polymerase, insert domain superfamily / RNA polymerase, N-terminal / RNA polymerase Rpb2, domain 2 / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase Rpb2, domain 2 / RNA polymerase I subunit A N-terminus / RNA polymerase, RBP11-like subunit / 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 / OB fold (Dihydrolipoamide Acetyltransferase, E2P) / Winged helix-like DNA-binding domain superfamily / Alpha-Beta Complex / Beta Barrel / Orthogonal Bundle / Mainly Beta / Mainly Alpha / Alpha Beta
Similarity search - Domain/homology
DNA / DNA (> 10) / DNA-directed RNA polymerase subunit beta' / Cell cycle regulatory protein GcrA / RNA polymerase sigma factor RpoD / DNA-directed RNA polymerase subunit beta / DNA-directed RNA polymerase subunit alpha / DNA-directed RNA polymerase subunit omega
Similarity search - Component
Biological speciesCaulobacter vibrioides NA1000 (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.7 Å
AuthorsWu, X.X. / Zhang, Y.
Funding support China, 2items
OrganizationGrant numberCountry
Other governmentXDB29020000
National Natural Science Foundation of China (NSFC)31870047 China
CitationJournal: Nucleic Acids Res. / Year: 2023
Title: Cryo-EM structures of Caulobacter crescentus transcription activation complex with an essential cell cycle regulator GcrA
Authors: Wu, X.X. / Zhang, Y.
History
DepositionJul 5, 2022Deposition site: PDBJ / Processing site: PDBJ
Revision 1.0Jan 18, 2023Provider: repository / Type: Initial release
Revision 1.1Jul 3, 2024Group: Data collection / Category: chem_comp_atom / chem_comp_bond / em_admin / Item: _em_admin.last_update

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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 alpha
B: DNA-directed RNA polymerase subunit alpha
C: DNA-directed RNA polymerase subunit beta
D: DNA-directed RNA polymerase subunit beta'
E: DNA-directed RNA polymerase subunit omega
F: RNA polymerase sigma factor RpoD
G: Cell cycle regulatory protein GcrA
H: DNA (57-MER)-non template
I: DNA (57-MER)-template
hetero molecules


Theoretical massNumber of molelcules
Total (without water)520,30713
Polymers520,0879
Non-polymers2214
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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

#1: Protein DNA-directed RNA polymerase subunit alpha / RNAP subunit alpha / RNA polymerase subunit alpha / Transcriptase subunit alpha


Mass: 37315.523 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Caulobacter vibrioides NA1000 (bacteria)
Strain: NA1000 / Gene: rpoA / Production host: Escherichia coli DH5[alpha] (bacteria) / References: UniProt: B8H4F8, DNA-directed RNA polymerase
#2: Protein DNA-directed RNA polymerase subunit beta / RNAP subunit beta / RNA polymerase subunit beta / Transcriptase subunit beta


Mass: 151085.609 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Caulobacter vibrioides NA1000 (bacteria)
Strain: NA1000 / Gene: rpoB / Production host: Escherichia coli DH5[alpha] (bacteria) / References: UniProt: B8GZW7, DNA-directed RNA polymerase
#3: Protein DNA-directed RNA polymerase subunit beta' / RNAP subunit beta' / RNA polymerase subunit beta' / Transcriptase subunit beta'


Mass: 154467.000 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Caulobacter vibrioides NA1000 (bacteria)
Strain: NA1000 / Gene: rpoC / Production host: Escherichia coli DH5[alpha] (bacteria)
References: UniProt: A0A0H3C4W8, DNA-directed RNA polymerase
#4: Protein DNA-directed RNA polymerase subunit omega / RNAP omega subunit / RNA polymerase omega subunit / Transcriptase subunit omega


Mass: 13289.844 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Caulobacter vibrioides NA1000 (bacteria)
Strain: NA1000 / Gene: rpoZ / Production host: Escherichia coli DH5[alpha] (bacteria) / References: UniProt: B8H618, DNA-directed RNA polymerase

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Protein , 2 types, 2 molecules FG

#5: Protein RNA polymerase sigma factor RpoD / Sigma-70


Mass: 72770.172 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Caulobacter vibrioides NA1000 (bacteria)
Strain: NA1000 / Gene: rpoD / Production host: Escherichia coli DH5[alpha] (bacteria) / References: UniProt: A0A0H3CAV3
#6: Protein Cell cycle regulatory protein GcrA


Mass: 18515.287 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Caulobacter vibrioides NA1000 (bacteria)
Strain: NA1000 / Gene: gcrA / Production host: Escherichia coli DH5[alpha] (bacteria) / References: UniProt: A0A0H3C9J4

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DNA (57-MER)- ... , 2 types, 2 molecules HI

#7: DNA chain DNA (57-MER)-non template


Mass: 17675.373 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Caulobacter vibrioides NA1000 (bacteria)
#8: DNA chain DNA (57-MER)-template


Mass: 17652.293 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Caulobacter vibrioides NA1000 (bacteria)

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Non-polymers , 2 types, 4 molecules

#9: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: Mg
#10: Chemical ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 3 / Source method: obtained synthetically / Formula: Zn

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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
1The cryo-EM structure of C. crescentus GcrA-TACupCOMPLEX#1-#80MULTIPLE SOURCES
2DNA-directed RNA polymerase/GcrA/TACupCOMPLEX#1-#61RECOMBINANT
3DNACOMPLEX#7-#81SYNTHETIC
Molecular weightExperimental value: NO
Source (natural)Organism: Caulobacter vibrioides NA1000 (bacteria)
Source (recombinant)Organism: Escherichia coli DH5[alpha] (bacteria)
Buffer solutionpH: 8
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: COPPER / Grid mesh size: 400 divisions/in. / Grid type: C-flat-1.2/1.3
VitrificationCryogen name: NITROGEN

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 2200 nm / Nominal defocus min: 1200 nm
Image recordingElectron dose: 62.4 e/Å2 / Film or detector model: GATAN K2 SUMMIT (4k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING ONLY
3D reconstructionResolution: 3.7 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 287227 / Symmetry type: POINT

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