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- PDB-8k60: Cryo-EM structure of Streptomyces coelicolor transcription initia... -

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Basic information

Entry
Database: PDB / ID: 8k60
TitleCryo-EM structure of Streptomyces coelicolor transcription initiation complex with the global transcription factor AfsR
Components
  • (DNA-directed RNA polymerase subunit ...) x 4
  • Non-template strand DNA for AfsS promoter
  • RNA polymerase principal sigma factor HrdB
  • Regulatory protein AfsR
  • Template strand DNA for AfsS promoter
KeywordsGENE REGULATION / transcription factor / TRANSCRIPTION-DNA complex
Function / homology
Function and homology information


pigment binding / bacterial-type RNA polymerase holo enzyme binding / nucleoid / phosphorelay signal transduction system / sigma factor activity / antibiotic biosynthetic process / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / ADP binding ...pigment binding / bacterial-type RNA polymerase holo enzyme binding / nucleoid / phosphorelay signal transduction system / sigma factor activity / antibiotic biosynthetic process / DNA-directed RNA polymerase complex / DNA-templated transcription initiation / ribonucleoside binding / ADP binding / DNA-directed 5'-3' RNA polymerase activity / DNA-directed RNA polymerase / protein dimerization activity / DNA-templated transcription / regulation of DNA-templated transcription / magnesium ion binding / DNA binding / zinc ion binding / ATP binding / cytoplasm
Similarity search - Function
Bacterial transcriptional activator domain / Bacterial transcriptional activator domain / Bacterial transcriptional activator domain / Tetratricopeptide repeat / NB-ARC / NB-ARC domain / OmpR/PhoB-type DNA-binding domain profile. / OmpR/PhoB-type DNA-binding domain / Transcriptional regulatory protein, C terminal / Transcriptional regulatory protein, C terminal ...Bacterial transcriptional activator domain / Bacterial transcriptional activator domain / Bacterial transcriptional activator domain / Tetratricopeptide repeat / NB-ARC / NB-ARC domain / OmpR/PhoB-type DNA-binding domain profile. / OmpR/PhoB-type DNA-binding domain / Transcriptional regulatory protein, C terminal / Transcriptional regulatory protein, C terminal / Tetratricopeptide repeat / Signal transduction response regulator, C-terminal effector / Sigma-70 factors family signature 1. / RNA polymerase sigma factor RpoD, C-terminal / RNA polymerase sigma factor RpoD / 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 / TPR repeat region circular profile. / DNA-directed RNA polymerase, alpha subunit / DNA-directed RNA polymerase beta subunit, bacterial-type / TPR repeat profile. / Tetratricopeptide repeats / Tetratricopeptide repeat / RNA polymerase Rpb6 / RNA polymerase, subunit omega/Rpo6/RPB6 / RNA polymerase Rpb6 / RNA polymerase Rpb1, domain 3 superfamily / RPB6/omega subunit-like superfamily / RNA polymerase Rpb1, clamp domain superfamily / DNA-directed RNA polymerase, subunit beta-prime / RNA polymerase Rpb2, domain 2 superfamily / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 3 / RNA polymerase Rpb1, domain 1 / RNA polymerase Rpb1, domain 1 / 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 / RNA polymerase, beta subunit, protrusion / RNA polymerase beta subunit / RNA polymerase, N-terminal / RNA polymerase Rpb1, funnel domain superfamily / RNA polymerase I subunit A N-terminus / 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 / Tetratricopeptide-like helical domain superfamily / Winged helix-like DNA-binding domain superfamily / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
DNA / DNA (> 10) / RNA polymerase principal sigma factor HrdB / Regulatory protein AfsR / DNA-directed RNA polymerase subunit alpha / DNA-directed RNA polymerase subunit beta' / DNA-directed RNA polymerase subunit omega / DNA-directed RNA polymerase subunit beta
Similarity search - Component
Biological speciesStreptomyces coelicolor (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.4 Å
AuthorsLin, W. / Shi, J.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)81903526 China
CitationJournal: Iscience / Year: 2024
Title: Structural insights into transcription activation of the Streptomyces antibiotic regulatory protein, AfsR
Authors: Shi, J. / Ye, Z. / Feng, Z. / Wen, A. / Wang, L. / Zhang, Z. / Xu, L. / Song, Q. / Wang, F. / Liu, T. / Wang, S. / Feng, Y. / Lin, W.
History
DepositionJul 24, 2023Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Jul 31, 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 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 principal sigma factor HrdB
G: Non-template strand DNA for AfsS promoter
H: Template strand DNA for AfsS promoter
I: Regulatory protein AfsR
J: Regulatory protein AfsR
K: DNA-directed RNA polymerase subunit alpha
hetero molecules


Theoretical massNumber of molelcules
Total (without water)697,76914
Polymers697,61411
Non-polymers1553
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration, not applicable
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

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DNA-directed RNA polymerase subunit ... , 4 types, 6 molecules ABKCDE

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


Mass: 36734.641 Da / Num. of mol.: 3
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)
Gene: rpoA, SCO4729, SC6G4.07 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P60312, DNA-directed RNA polymerase
#2: Protein DNA-directed RNA polymerase subunit beta / RNAP subunit beta / RNA polymerase subunit beta / Transcriptase subunit beta


Mass: 128644.945 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)
Gene: rpoB, SCO4654, SCD82.26 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9L0L0, DNA-directed RNA polymerase
#3: Protein DNA-directed RNA polymerase subunit beta' / RNAP subunit beta' / RNA polymerase subunit beta' / Transcriptase subunit beta'


Mass: 144807.078 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)
Gene: rpoC, SCO4655, SCD40A.01, SCD82.27 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q8CJT1, DNA-directed RNA polymerase
#4: Protein DNA-directed RNA polymerase subunit omega / RNAP omega subunit / RNA polymerase omega subunit / Transcriptase subunit omega


Mass: 9716.941 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)
Gene: rpoZ, SCO1478, SC9C5.02c / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: Q9KXS1, DNA-directed RNA polymerase

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Protein , 2 types, 3 molecules FIJ

#5: Protein RNA polymerase principal sigma factor HrdB / RNA polymerase sigma factor SigA


Mass: 56017.578 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)
Gene: hrdB, sigA, SCO5820, SC5B8.10 / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P18183
#8: Protein Regulatory protein AfsR


Mass: 105842.578 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)
Gene: afsR, afsB, SCO4426, SCD6.04c / Production host: Escherichia coli BL21(DE3) (bacteria) / References: UniProt: P25941

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DNA chain , 2 types, 2 molecules GH

#6: DNA chain Non-template strand DNA for AfsS promoter


Mass: 18177.602 Da / Num. of mol.: 1 / Source method: obtained synthetically
Source: (synth.) Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)
#7: DNA chain Template strand DNA for AfsS promoter


Mass: 18360.770 Da / Num. of mol.: 1 / Source method: obtained synthetically
Source: (synth.) Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)

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Non-polymers , 2 types, 3 molecules

#9: Chemical ChemComp-MG / MAGNESIUM ION


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


Mass: 65.409 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Zn / 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

ComponentName: Streptomyces coelicolor transcription initiation complex with the global transcription factor AfsR
Type: COMPLEX / Entity ID: #1-#8 / Source: MULTIPLE SOURCES
Source (natural)Organism: Streptomyces coelicolor (strain ATCC BAA-471 / A3(2) / M145) (bacteria)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.9
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
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: 2000 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 52 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: PHENIX / Version: 1.20.1_4487: / Category: model refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.4 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 231221 / Symmetry type: POINT

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