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- EMDB-4909: E. coli DNA Gyrase - DNA binding and cleavage domain in State 1 w... -

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Entry
Database: EMDB / ID: EMD-4909
TitleE. coli DNA Gyrase - DNA binding and cleavage domain in State 1 without TOPRIM insertion
Map dataE. coli DNA Gyrase DNA binding and cleavage domain. The TOPRIM insertion was masked away during refinement.
Sample
  • Complex: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
    • Complex: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
      • Protein or peptide: DNA gyrase subunit A
    • Complex: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
      • Protein or peptide: DNA gyrase subunit B
    • Complex: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
      • DNA: DNA Strand 1
      • DNA: DNA Strand 2
  • Ligand: (3~{R})-3-[[4-(3,4-dihydro-2~{H}-pyrano[2,3-c]pyridin-6-ylmethylamino)piperidin-1-yl]methyl]-1,4,7-triazatricyclo[6.3.1.0^{4,12}]dodeca-6,8(12),9-triene-5,11-dione
KeywordsIsomerase / Complex / DNA Gyrase / Inhibitor / DNA BINDING PROTEIN
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 / identical protein binding / membrane / 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 / 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 / DNA gyrase subunit B, TOPRIM domain / Topoisomerase (Topo) IIA-type catalytic domain profile. / DNA gyrase, subunit B / DNA topoisomerase, type IIA, subunit B / DNA gyrase B subunit, C-terminal / DNA gyrase B subunit, carboxyl terminus / 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 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
DNA gyrase subunit A / DNA gyrase subunit B
Similarity search - Component
Biological speciesEscherichia coli K-12 (bacteria) / Escherichia coli O157:H7 str. SS52 (bacteria) / Escherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 4.0 Å
AuthorsVanden Broeck A / Lamour V
Funding support France, 3 items
OrganizationGrant numberCountry
French National Research AgencyANR-10-LABX-0030-INRT France
French National Research AgencyANR-10-IDEX-0002-02 France
French Infrastructure for Integrated Structural BiologyFRISBI ANR-10-INBS-05 France
CitationJournal: Nat Commun / Year: 2019
Title: Cryo-EM structure of the complete E. coli DNA gyrase nucleoprotein complex.
Authors: Arnaud Vanden Broeck / Christophe Lotz / Julio Ortiz / Valérie Lamour /
Abstract: DNA gyrase is an essential enzyme involved in the homeostatic control of DNA supercoiling and the target of successful antibacterial compounds. Despite extensive studies, a detailed architecture of ...DNA gyrase is an essential enzyme involved in the homeostatic control of DNA supercoiling and the target of successful antibacterial compounds. Despite extensive studies, a detailed architecture of the full-length DNA gyrase from the model organism E. coli is still missing. Herein, we report the complete structure of the E. coli DNA gyrase nucleoprotein complex trapped by the antibiotic gepotidacin, using phase-plate single-particle cryo-electron microscopy. Our data unveil the structural and spatial organization of the functional domains, their connections and the position of the conserved GyrA-box motif. The deconvolution of two states of the DNA-binding/cleavage domain provides a better understanding of the allosteric movements of the enzyme complex. The local atomic resolution in the DNA-bound area reaching up to 3.0 Å enables the identification of the antibiotic density. Altogether, this study paves the way for the cryo-EM determination of gyrase complexes with antibiotics and opens perspectives for targeting conformational intermediates.
History
DepositionApr 30, 2019-
Header (metadata) releaseMay 15, 2019-
Map releaseNov 6, 2019-
UpdateMay 22, 2024-
Current statusMay 22, 2024Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.0263
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.0263
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-6rks
  • Surface level: 0.0263
  • Imaged by UCSF Chimera
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Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_4909.png

Downloads & links

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Map

FileDownload / File: emd_4909.map.gz / Format: CCP4 / Size: 178 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
AnnotationE. coli DNA Gyrase DNA binding and cleavage domain. The TOPRIM insertion was masked away during refinement.
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
0.88 Å/pix.
x 360 pix.
= 316.8 Å		0.88 Å/pix.
x 360 pix.
= 316.8 Å		0.88 Å/pix.
x 360 pix.
= 316.8 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 0.88 Å
Density

Histogram

Histogram (log scale)
Contour LevelBy AUTHOR: 0.0263 / Movie #1: 0.0263
Minimum - Maximum-0.05388446 - 0.105341084
Average (Standard dev.)0.00012960639 (±0.0024498145)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions360360360
Spacing360360360
CellA=B=C: 316.8 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z0.880.880.88
M x/y/z360360360
origin x/y/z0.0000.0000.000
length x/y/z316.800316.800316.800
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS360360360
D min/max/mean-0.0540.1050.000

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Supplemental data

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Mask #1

Fileemd_4909_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Sample components

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Entire : DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1

EntireName: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
Components
  • Complex: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
    • Complex: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
      • Protein or peptide: DNA gyrase subunit A
    • Complex: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
      • Protein or peptide: DNA gyrase subunit B
    • Complex: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
      • DNA: DNA Strand 1
      • DNA: DNA Strand 2
  • Ligand: (3~{R})-3-[[4-(3,4-dihydro-2~{H}-pyrano[2,3-c]pyridin-6-ylmethylamino)piperidin-1-yl]methyl]-1,4,7-triazatricyclo[6.3.1.0^{4,12}]dodeca-6,8(12),9-triene-5,11-dione

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Supramolecule #1: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1

SupramoleculeName: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
type: complex / ID: 1 / Parent: 0 / Macromolecule list: #1-#4
Details: The TOPRIM insertion was masked out during refinement.

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Supramolecule #2: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1

SupramoleculeName: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
type: complex / ID: 2 / Parent: 1 / Macromolecule list: #1
Details: The TOPRIM insertion was masked out during refinement.
Source (natural)Organism: Escherichia coli K-12 (bacteria)

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Supramolecule #3: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1

SupramoleculeName: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
type: complex / ID: 3 / Parent: 1 / Macromolecule list: #2
Details: The TOPRIM insertion was masked out during refinement.
Source (natural)Organism: Escherichia coli O157:H7 str. SS52 (bacteria)

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Supramolecule #4: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1

SupramoleculeName: DNA binding and cleavage domain of the E. coli DNA Gyrase in State 1
type: complex / ID: 4 / Parent: 1 / Macromolecule list: #3-#4
Details: The TOPRIM insertion was masked out during refinement.
Source (natural)Organism: Escherichia coli (E. coli)

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Macromolecule #1: DNA gyrase subunit A

MacromoleculeName: DNA gyrase subunit A / type: protein_or_peptide / ID: 1 / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA topoisomerase (ATP-hydrolysing)
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Molecular weightTheoretical: 97.088586 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MSDLAREITP VNIEEELKSS YLDYAMSVIV GRALPDVRDG LKPVHRRVLY AMNVLGNDWN KAYKKSARVV GDVIGKYHPH GDSAVYDTI VRMAQPFSLR YMLVDGQGNF GSIDGDSAAA MRYTEIRLAK IAHELMADLE KETVDFVDNY DGTEKIPDVM P TKIPNLLV ...String:
MSDLAREITP VNIEEELKSS YLDYAMSVIV GRALPDVRDG LKPVHRRVLY AMNVLGNDWN KAYKKSARVV GDVIGKYHPH GDSAVYDTI VRMAQPFSLR YMLVDGQGNF GSIDGDSAAA MRYTEIRLAK IAHELMADLE KETVDFVDNY DGTEKIPDVM P TKIPNLLV NGSSGIAVGM ATNIPPHNLT EVINGCLAYI DDEDISIEGL MEHIPGPDFP TAAIINGRRG IEEAYRTGRG KV YIRARAE VEVDAKTGRE TIIVHEIPYQ VNKARLIEKI AELVKEKRVE GISALRDESD KDGMRIVIEV KRDAVGEVVL NNL YSQTQL QVSFGINMVA LHHGQPKIMN LKDIIAAFVR HRREVVTRRT IFELRKARDR AHILEALAVA LANIDPIIEL IRHA PTPAE AKTALVANPW QLGNVAAMLE RAGDDAARPE WLEPEFGVRD GLYYLTEQQA QAILDLRLQK LTGLEHEKLL DEYKE LLDQ IAELLRILGS ADRLMEVIRE ELELVREQFG DKRRTEITAN SADINLEDLI TQEDVVVTLS HQGYVKYQPL SEYEAQ RRG GKGKSAARIK EEDFIDRLLV ANTHDHILCF SSRGRVYSMK VYQLPEATRG ARGRPIVNLL PLEQDERITA ILPVTEF EE GVKVFMATAN GTVKKTVLTE FNRLRTAGKV AIKLVDGDEL IGVDLTSGED EVMLFSAEGK VVRFKESSVR AMGCNTTG V RGIRLGEGDK VVSLIVPRGD GAILTATQNG YGKRTAVAEY PTKSRATKGV ISIKVTERNG LVVGAVQVDD CDQIMMITD AGTLVRTRVS EISIVGRNTQ GVILIRTAED ENVVGLQRVA EPVDEEDLDT IDGSAAEGDD EIAPEVDVDD EPEEE

UniProtKB: DNA gyrase subunit A

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Macromolecule #2: DNA gyrase subunit B

MacromoleculeName: DNA gyrase subunit B / type: protein_or_peptide / ID: 2 / Details: GyrB subunit DNA Binding and cleavage domain / Number of copies: 2 / Enantiomer: LEVO / EC number: DNA topoisomerase (ATP-hydrolysing)
Source (natural)Organism: Escherichia coli K-12 (bacteria)
Molecular weightTheoretical: 90.073922 KDa
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria)
SequenceString: MSNSYDSSSI KVLKGLDAVR KRPGMYIGDT DDGTGLHHMV FEVVDNAIDE ALAGHCKEII VTIHADNSVS VQDDGRGIPT GIHPEEGVS AAEVIMTVLH AGGKFDDNSY KVSGGLHGVG VSVVNALSQK LELVIQREGK IHRQIYEHGV PQAPLAVTGE T EKTGTMVR ...String:
MSNSYDSSSI KVLKGLDAVR KRPGMYIGDT DDGTGLHHMV FEVVDNAIDE ALAGHCKEII VTIHADNSVS VQDDGRGIPT GIHPEEGVS AAEVIMTVLH AGGKFDDNSY KVSGGLHGVG VSVVNALSQK LELVIQREGK IHRQIYEHGV PQAPLAVTGE T EKTGTMVR FWPSLETFTN VTEFEYEILA KRLRELSFLN SGVSIRLRDK RDGKEDHFHY EGGIKAFVEY LNKNKTPIHP NI FYFSTEK DGIGVEVALQ WNDGFQENIY CFTNNIPQRD GGTHLAGFRA AMTRTLNAYM DKEGYSKKAK VSATGDDARE GLI AVVSVK VPDPKFSSQT KDKLVSSEVK SAVEQQMNEL LAEYLLENPT DAKIVVGKII DAARAREAAR RAREMTRRKG ALDL AGLPG KLADCQERDP ALSELYLVEG DSAGGSAKQG RNRKNQAILP LKGKILNVEK ARFDKMLSSQ EVATLITALG CGIGR DEYN PDKLRYHSII IMTDADVDGS HIRTLLLTFF YRQMPEIVER GHVYIAQPPL YKVKKGKQEQ YIKDDEAMDQ YQISIA LDG ATLHTNASAP ALAGEALEKL VSEYNATQKM INRMERRYPK AMLKELIYQP TLTEADLSDE QTVTRWVNAL VSELNDK EQ HGSQWKFDVH TNAEQNLFEP IVRVRTHGVD TDYPLDHEFI TGGEYRRICT LGEKLRGLLE EDAFIERGER RQPVASFE Q ALDWLVKESR RGLSIQRYKG LGEMNPEQLW ETTMDPESRR MLRVTVKDAI AADQLFTTLM GDAVEPRRAF IEENALKAA NIDI

UniProtKB: DNA gyrase subunit B

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Macromolecule #3: DNA Strand 1

MacromoleculeName: DNA Strand 1 / type: dna / ID: 3 / Number of copies: 2 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 4.288818 KDa
SequenceString:
(DA)(DT)(DC)(DA)(DT)(DA)(DA)(DG)(DC)(DC) (DG)(DT)(DA)(DG)

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Macromolecule #4: DNA Strand 2

MacromoleculeName: DNA Strand 2 / type: dna / ID: 4 / Number of copies: 2 / Classification: DNA
Source (natural)Organism: Escherichia coli (E. coli)
Molecular weightTheoretical: 5.506578 KDa
SequenceString:
(DG)(DT)(DA)(DC)(DC)(DT)(DA)(DC)(DG)(DG) (DC)(DT)(DT)(DA)(DT)(DG)(DA)(DT)

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Macromolecule #5: (3~{R})-3-[[4-(3,4-dihydro-2~{H}-pyrano[2,3-c]pyridin-6-ylmethyla...

MacromoleculeName: (3~{R})-3-[[4-(3,4-dihydro-2~{H}-pyrano[2,3-c]pyridin-6-ylmethylamino)piperidin-1-yl]methyl]-1,4,7-triazatricyclo[6.3.1.0^{4,12}]dodeca-6,8(12),9-triene-5,11-dione
type: ligand / ID: 5 / Number of copies: 1 / Formula: JHN
Molecular weightTheoretical: 448.518 Da
Chemical component information

ChemComp-JHN:
(3~{R})-3-[[4-(3,4-dihydro-2~{H}-pyrano[2,3-c]pyridin-6-ylmethylamino)piperidin-1-yl]methyl]-1,4,7-triazatricyclo[6.3.1.0^{4,12}]dodeca-6,8(12),9-triene-5,11-dione / antibiotic, inhibitor*YM

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Experimental details

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Structure determination

Methodcryo EM
Processingsingle particle reconstruction
Aggregation stateparticle

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Sample preparation

BufferpH: 8
VitrificationCryogen name: ETHANE

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsPhase plate: VOLTA PHASE PLATE / Energy filter - Slit width: 20 eV
Image recordingFilm or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: SUPER-RESOLUTION / Digitization - Frames/image: 3-40 / Average electron dose: 50.0 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsIllumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.01 mm / Nominal defocus max: 0.8 µm / Nominal defocus min: 0.3 µm / Nominal magnification: 130000
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Particle selectionNumber selected: 1572962
Startup modelType of model: NONE / Details: Ab initio model was generated with CryoSPARC V1
Final reconstructionApplied symmetry - Point group: C2 (2 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 4.0 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.0.3) / Number images used: 60548
Initial angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: cryoSPARC (ver. V1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial model
PDB IDChain

3nuh

chain_id: A, source_name: PDB, initial_model_type: experimental model

3nuh

chain_id: B, source_name: PDB, initial_model_type: experimental model

3nuh

chain_id: C, source_name: PDB, initial_model_type: experimental model

3nuh

chain_id: D, source_name: PDB, initial_model_type: experimental model
RefinementSpace: REAL
Output model

PDB-6rks:
E. coli DNA Gyrase - DNA binding and cleavage domain in State 1 without TOPRIM insertion

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