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- PDB-7z9g: E.coli gyrase holocomplex with 217 bp DNA and Albi-2 -

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Basic information

Entry
Database: PDB / ID: 7z9g
TitleE.coli gyrase holocomplex with 217 bp DNA and Albi-2
Components
  • (DNA gyrase subunit ...) x 2
  • DNA (5'-D(*AP*AP*TP*CP*AP*CP*CP*CP*GP*CP*AP*CP*AP*GP*AP*TP*TP*T)-3')
  • DNA (5'-D(*GP*AP*TP*TP*TP*TP*AP*TP*GP*CP*CP*TP*GP*AP*TP*TP*CP*T)-3')
  • DNA (5'-D(P*AP*AP*AP*TP*CP*TP*GP*TP*GP*CP*GP*GP*GP*T)-3')
  • DNA (5'-D(P*AP*GP*AP*AP*TP*CP*AP*GP*GP*CP*AP*TP*AP*A)-3')
KeywordsISOMERASE / type II topoisomerase / antibiotic / albicidin / DNA gyrase
Function / homology
Function and homology information


negative regulation of DNA-templated DNA replication / DNA topoisomerase type II (double strand cut, ATP-hydrolyzing) complex / DNA negative supercoiling activity / DNA topoisomerase type II (double strand cut, ATP-hydrolyzing) activity / DNA topoisomerase (ATP-hydrolysing) / DNA topological change / ATP-dependent activity, acting on DNA / DNA-templated DNA replication / chromosome / response to xenobiotic stimulus ...negative regulation of DNA-templated DNA replication / DNA topoisomerase type II (double strand cut, ATP-hydrolyzing) complex / DNA negative supercoiling activity / DNA topoisomerase type II (double strand cut, ATP-hydrolyzing) activity / DNA topoisomerase (ATP-hydrolysing) / DNA topological change / ATP-dependent activity, acting on DNA / DNA-templated DNA replication / chromosome / response to xenobiotic stimulus / response to antibiotic / DNA-templated transcription / DNA binding / ATP binding / membrane / identical protein binding / metal ion binding / cytosol / cytoplasm
Similarity search - Function
: / GyrB, hook / DNA gyrase subunit B insert domain / DNA gyrase B subunit insert domain / DNA gyrase, subunit A / DNA gyrase/topoisomerase IV, subunit A, C-terminal repeat / DNA gyrase/topoisomerase IV, subunit A, C-terminal / DNA gyrase C-terminal domain, beta-propeller / Topoisomerase (Topo) IIA-type catalytic domain profile. / DNA gyrase subunit B, TOPRIM domain ...: / GyrB, hook / DNA gyrase subunit B insert domain / DNA gyrase B subunit insert domain / DNA gyrase, subunit A / DNA gyrase/topoisomerase IV, subunit A, C-terminal repeat / DNA gyrase/topoisomerase IV, subunit A, C-terminal / DNA gyrase C-terminal domain, beta-propeller / Topoisomerase (Topo) IIA-type catalytic domain profile. / DNA gyrase subunit B, TOPRIM domain / DNA gyrase, subunit B / DNA topoisomerase, type IIA, subunit B / DNA topoisomerase, type IIA, alpha-helical domain superfamily / DNA topoisomerase, type IIA, domain A / DNA topoisomerase, type IIA, domain A, alpha-beta / DNA gyrase/topoisomerase IV, subunit A / DNA Topoisomerase IV / DNA gyrase B subunit, C-terminal / DNA gyrase B subunit, carboxyl terminus / DNA topoisomerase, type IIA, subunit B, domain 2 / DNA gyrase B / DNA topoisomerase, type IIA / DNA topoisomerase, type IIA, conserved site / DNA topoisomerase II signature. / TopoisomeraseII / DNA topoisomerase, type IIA, subunit B, C-terminal / Toprim domain / DNA topoisomerase, type IIA-like domain superfamily / Toprim domain profile. / TOPRIM domain / Histidine kinase-, DNA gyrase B-, and HSP90-like ATPase / Histidine kinase-like ATPases / Histidine kinase/HSP90-like ATPase / Histidine kinase/HSP90-like ATPase superfamily / Ribosomal protein S5 domain 2-type fold, subgroup / Ribosomal protein S5 domain 2-type fold
Similarity search - Domain/homology
Chem-IM0 / DNA / DNA (> 10) / DNA gyrase subunit A / DNA gyrase subunit B
Similarity search - Component
Biological speciesEscherichia coli str. K-12 substr. MG1655 (bacteria)
Escherichia phage Mu (virus)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.25 Å
AuthorsGhilarov, D. / Heddle, J.G.H.
Funding support United Kingdom, Poland, 2items
OrganizationGrant numberCountry
Wellcome Trust221868/Z/20/Z United Kingdom
Polish National Science Centre2020/39/B/NZ1/02898 Poland
CitationJournal: Nat Catal / Year: 2023
Title: Molecular mechanism of topoisomerase poisoning by the peptide antibiotic albicidin.
Authors: Elizabeth Michalczyk / Kay Hommernick / Iraj Behroz / Marcel Kulike / Zuzanna Pakosz-Stępień / Lukasz Mazurek / Maria Seidel / Maria Kunert / Karine Santos / Holger von Moeller / Bernhard ...Authors: Elizabeth Michalczyk / Kay Hommernick / Iraj Behroz / Marcel Kulike / Zuzanna Pakosz-Stępień / Lukasz Mazurek / Maria Seidel / Maria Kunert / Karine Santos / Holger von Moeller / Bernhard Loll / John B Weston / Andi Mainz / Jonathan G Heddle / Roderich D Süssmuth / Dmitry Ghilarov /
Abstract: The peptide antibiotic albicidin is a DNA topoisomerase inhibitor with low-nanomolar bactericidal activity towards fluoroquinolone-resistant Gram-negative pathogens. However, its mode of action is ...The peptide antibiotic albicidin is a DNA topoisomerase inhibitor with low-nanomolar bactericidal activity towards fluoroquinolone-resistant Gram-negative pathogens. However, its mode of action is poorly understood. We determined a 2.6 Å resolution cryoelectron microscopy structure of a ternary complex between topoisomerase DNA gyrase, a 217 bp double-stranded DNA fragment and albicidin. Albicidin employs a dual binding mechanism where one end of the molecule obstructs the crucial gyrase dimer interface, while the other intercalates between the fragments of cleaved DNA substrate. Thus, albicidin efficiently locks DNA gyrase, preventing it from religating DNA and completing its catalytic cycle. Two additional structures of this trapped state were determined using synthetic albicidin analogues that demonstrate improved solubility, and activity against a range of gyrase variants and topoisomerase IV. The extraordinary promiscuity of the DNA-intercalating region of albicidins and their excellent performance against fluoroquinolone-resistant bacteria holds great promise for the development of last-resort antibiotics.
History
DepositionMar 21, 2022Deposition site: PDBE / Processing site: PDBE
Revision 1.0Feb 15, 2023Provider: repository / Type: Initial release
Revision 1.1Jul 19, 2023Group: Author supporting evidence / Category: pdbx_audit_support

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: DNA gyrase subunit A
B: DNA gyrase subunit B
C: DNA gyrase subunit A
D: DNA gyrase subunit B
E: DNA (5'-D(P*AP*AP*AP*TP*CP*TP*GP*TP*GP*CP*GP*GP*GP*T)-3')
F: DNA (5'-D(P*AP*GP*AP*AP*TP*CP*AP*GP*GP*CP*AP*TP*AP*A)-3')
G: DNA (5'-D(*AP*AP*TP*CP*AP*CP*CP*CP*GP*CP*AP*CP*AP*GP*AP*TP*TP*T)-3')
H: DNA (5'-D(*GP*AP*TP*TP*TP*TP*AP*TP*GP*CP*CP*TP*GP*AP*TP*TP*CP*T)-3')
hetero molecules


Theoretical massNumber of molelcules
Total (without water)398,05511
Polymers397,0828
Non-polymers9733
Water18010
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: electron microscopy, gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area21970 Å2
ΔGint-136 kcal/mol
Surface area100110 Å2

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Components

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DNA gyrase subunit ... , 2 types, 4 molecules ACBD

#1: Protein DNA gyrase subunit A /


Mass: 97854.305 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli str. K-12 substr. MG1655 (bacteria)
Strain: K12 / Gene: gyrA, hisW, nalA, parD, b2231, JW2225 / Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): Star
References: UniProt: P0AES4, DNA topoisomerase (ATP-hydrolysing)
#2: Protein DNA gyrase subunit B / / Type IIA topoisomerase subunit GyrB


Mass: 90891.734 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia coli str. K-12 substr. MG1655 (bacteria)
Strain: K12
Gene: gyrB, acrB, cou, himB, hisU, nalC, parA, pcbA, b3699, JW5625
Production host: Escherichia coli BL21(DE3) (bacteria) / Strain (production host): Star
References: UniProt: P0AES6, DNA topoisomerase (ATP-hydrolysing)

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DNA chain , 4 types, 4 molecules EFGH

#3: DNA chain DNA (5'-D(P*AP*AP*AP*TP*CP*TP*GP*TP*GP*CP*GP*GP*GP*T)-3')


Mass: 4335.827 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia phage Mu (virus) / Production host: Escherichia coli (E. coli)
#4: DNA chain DNA (5'-D(P*AP*GP*AP*AP*TP*CP*AP*GP*GP*CP*AP*TP*AP*A)-3')


Mass: 4321.856 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia phage Mu (virus) / Production host: Escherichia coli (E. coli)
#5: DNA chain DNA (5'-D(*AP*AP*TP*CP*AP*CP*CP*CP*GP*CP*AP*CP*AP*GP*AP*TP*TP*T)-3')


Mass: 5444.558 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia phage Mu (virus) / Production host: Escherichia coli (E. coli)
#6: DNA chain DNA (5'-D(*GP*AP*TP*TP*TP*TP*AP*TP*GP*CP*CP*TP*GP*AP*TP*TP*CP*T)-3')


Mass: 5487.561 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Escherichia phage Mu (virus) / Production host: Escherichia coli (E. coli)

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Non-polymers , 3 types, 13 molecules

#7: Chemical ChemComp-MG / MAGNESIUM ION


Mass: 24.305 Da / Num. of mol.: 2 / Source method: obtained synthetically / Formula: Mg
#8: Chemical ChemComp-IM0 / 4-[[3-(2-azanylethoxy)-2-oxidanyl-4-[[5-[[(2~{S})-2-[[4-[(6-oxidanylnaphthalen-2-yl)carbonylamino]phenyl]carbonylamino]-3-(1~{H}-1,2,3-triazol-4-yl)propanoyl]amino]pyridin-2-yl]carbonylamino]phenyl]carbonylamino]-3-methoxy-2-oxidanyl-benzoic acid


Mass: 924.870 Da / Num. of mol.: 1 / Source method: obtained synthetically / Formula: C46H40N10O12 / Feature type: SUBJECT OF INVESTIGATION
#9: Water ChemComp-HOH / water / Water


Mass: 18.015 Da / Num. of mol.: 10 / Source method: isolated from a natural source / Formula: H2O

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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
1Escherichia coli gyrase holocomplex with 217 bp phage Mu SGS DNA and albicidin stabilised by ADPNPCOMPLEX#1-#60MULTIPLE SOURCES
2Escherichia coli gyrase holocomplexCOMPLEX#1-#21RECOMBINANT
3217 bp phage Mu SGS DNACOMPLEX#3-#61RECOMBINANT
Molecular weightExperimental value: NO
Source (natural)
IDEntity assembly-IDOrganismNcbi tax-ID
22Escherichia coli str. K-12 substr. MG1655 (bacteria)511145
33Escherichia phage Mu (virus)2681603
Source (recombinant)
IDEntity assembly-IDOrganismNcbi tax-IDStrain
22Escherichia coli BL21 (DE3) (bacteria)469008Star
33Escherichia coli (E. coli)562
Buffer solutionpH: 8
Buffer componentConc.: 20 mM / Name: HEPES
SpecimenConc.: 12 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 95 % / Chamber temperature: 283 K

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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 FIELDBright-field microscopy / Nominal defocus max: 3000 nm / Nominal defocus min: 900 nm
Image recordingElectron dose: 40 e/Å2 / Film or detector model: GATAN K3 BIOQUANTUM (6k x 4k)

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Processing

SoftwareName: PHENIX / Version: 1.19.2_4158: / Classification: refinement
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
Particle selectionNum. of particles selected: 1222762
SymmetryPoint symmetry: C1 (asymmetric)
3D reconstructionResolution: 3.25 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 49921 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.00316360
ELECTRON MICROSCOPYf_angle_d0.55122390
ELECTRON MICROSCOPYf_dihedral_angle_d12.7736380
ELECTRON MICROSCOPYf_chiral_restr0.0412503
ELECTRON MICROSCOPYf_plane_restr0.0032764

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