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- EMDB-9316: Dihedral oligomeric complex of GyrA N-terminal fragment with DNA,... -

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Basic information

Entry
Database: EMDB / ID: 9316
TitleDihedral oligomeric complex of GyrA N-terminal fragment with DNA, solved by cryoEM in C2 symmetry
Map dataDihedral oligomeric complex of GyrA N-terminal fragment with DNA, solved by cryoEM in C2 symmetry
SampleComplex of DNA Gyrase A subunit with DNA in C2 symmetry:
DNA Gyrase A / DNA / DNA gyrase subunit A / (nucleic-acidNucleic acid) x 2
Function / homologyDNA gyrase, subunit A / DNA gyrase/topoisomerase IV, subunit A, C-terminal repeat / DNA gyrase C-terminal domain, beta-propeller / DNA gyrase/topoisomerase IV, subunit A / DNA gyrase/topoisomerase IV, subunit A, C-terminal / DNA topoisomerase, type IIA-like domain superfamily / DNA topoisomerase, type IIA, subunit A/ C-terminal, alpha-beta / DNA topoisomerase, type IIA, subunit A/C-terminal / Type IIA DNA topoisomerase subunit A, alpha-helical domain superfamily / DNA topoisomerase type II (ATP-hydrolyzing) activity ...DNA gyrase, subunit A / DNA gyrase/topoisomerase IV, subunit A, C-terminal repeat / DNA gyrase C-terminal domain, beta-propeller / DNA gyrase/topoisomerase IV, subunit A / DNA gyrase/topoisomerase IV, subunit A, C-terminal / DNA topoisomerase, type IIA-like domain superfamily / DNA topoisomerase, type IIA, subunit A/ C-terminal, alpha-beta / DNA topoisomerase, type IIA, subunit A/C-terminal / Type IIA DNA topoisomerase subunit A, alpha-helical domain superfamily / DNA topoisomerase type II (ATP-hydrolyzing) activity / DNA topoisomerase (ATP-hydrolysing) / DNA topological change / DNA-dependent DNA replication / chromosome / DNA binding / ATP binding / cytoplasm / DNA gyrase subunit A
Function and homology information
SourceStreptococcus pneumoniae G54 (bacteria) / Cloning vector pBR322 (others)
Methodsingle particle reconstruction / cryo EM / 6.35 Å resolution
AuthorsSoczek KM / Grant T / Rosenthal PB / Mondragon A
CitationJournal: Elife / Year: 2018
Title: CryoEM structures of open dimers of gyrase A in complex with DNA illuminate mechanism of strand passage.
Authors: Katarzyna M Soczek / Tim Grant / Peter B Rosenthal / Alfonso Mondragón
Abstract: Gyrase is a unique type IIA topoisomerase that uses ATP hydrolysis to maintain the negatively supercoiled state of bacterial DNA. In order to perform its function, gyrase undergoes a sequence of ...Gyrase is a unique type IIA topoisomerase that uses ATP hydrolysis to maintain the negatively supercoiled state of bacterial DNA. In order to perform its function, gyrase undergoes a sequence of conformational changes that consist of concerted gate openings, DNA cleavage, and DNA strand passage events. Structures where the transported DNA molecule (T-segment) is trapped by the A subunit have not been observed. Here we present the cryoEM structures of two oligomeric complexes of open gyrase A dimers and DNA. The protein subunits in these complexes were solved to 4 Å and 5.2 Å resolution. One of the complexes traps a linear DNA molecule, a putative T-segment, which interacts with the open gyrase A dimers in two states, representing steps either prior to or after passage through the DNA-gate. The structures locate the T-segment in important intermediate conformations of the catalytic cycle and provide insights into gyrase-DNA interactions and mechanism.
Validation ReportPDB-ID: 6n1p

SummaryFull reportAbout validation report
DateDeposition: Nov 10, 2018 / Header (metadata) release: Nov 21, 2018 / Map release: Dec 5, 2018 / Last update: Dec 5, 2018

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.059
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by radius
  • Surface level: 0.059
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: : PDB-6n1p
  • Surface level: 0.059
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

Fileemd_9316.map.gz (map file in CCP4 format, 32001 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
200 pix
1.24 Å/pix.
= 248. Å
200 pix
1.24 Å/pix.
= 248. Å
200 pix
1.24 Å/pix.
= 248. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.24 Å
Density
Contour Level:0.059 (by author), 0.059 (movie #1):
Minimum - Maximum-0.13098237 - 0.34460974
Average (Standard dev.)0.00288104 (0.018942565)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions200200200
Origin0.00.00.0
Limit199.0199.0199.0
Spacing200200200
CellA=B=C: 248.0 Å
α=β=γ: 90.0 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.241.241.24
M x/y/z200200200
origin x/y/z0.0000.0000.000
length x/y/z248.000248.000248.000
α/β/γ90.00090.00090.000
start NX/NY/NZ000
NX/NY/NZ450450450
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS200200200
D min/max/mean-0.1310.3450.003

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Supplemental data

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Sample components

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Entire Complex of DNA Gyrase A subunit with DNA in C2 symmetry

EntireName: Complex of DNA Gyrase A subunit with DNA in C2 symmetry
Number of components: 6

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Component #1: protein, Complex of DNA Gyrase A subunit with DNA in C2 symmetry

ProteinName: Complex of DNA Gyrase A subunit with DNA in C2 symmetry
Recombinant expression: No
MassTheoretical: 490 kDa

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Component #2: protein, DNA Gyrase A

ProteinName: DNA Gyrase A / Recombinant expression: No
SourceSpecies: Streptococcus pneumoniae G54 (bacteria)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria) / Vector: pMCSG7

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Component #3: protein, DNA

ProteinName: DNA / Recombinant expression: No
SourceSpecies: Cloning vector pBR322 (others)

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Component #4: protein, DNA gyrase subunit A

ProteinName: DNA gyrase subunit A / Number of Copies: 8 / Recombinant expression: No
MassTheoretical: 57.977578 kDa
SourceSpecies: Streptococcus pneumoniae G54 (bacteria)
Source (engineered)Expression System: Escherichia coli BL21(DE3) (bacteria)

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Component #5: nucleic-acid, DNA (44-MER)

Nucleic-acidName: DNA (44-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DG)(DA)(DG)(DA)(DA)(DG)(DA)(DA)(DT)(DC) (DA)(DT)(DA)(DA)(DT)(DG)(DG)(DG)(DG)(DA) (DA)(DG)(DG)(DC)(DC)(DA)(DT)(DC)(DC)(DA) (DG)(DC)(DC)(DT)(DC)(DG)(DC)(DG)(DT)(DC) (DG)(DC)(DG)(DA)
MassTheoretical: 13.641771 kDa
SourceSpecies: Cloning vector pBR322 (others)

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Component #6: nucleic-acid, DNA (44-MER)

Nucleic-acidName: DNA (44-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DT)(DC)(DG)(DC)(DG)(DA)(DC)(DG)(DC)(DG) (DA)(DG)(DG)(DC)(DT)(DG)(DG)(DA)(DT)(DG) (DG)(DC)(DC)(DT)(DT)(DC)(DC)(DC)(DC)(DA) (DT)(DT)(DA)(DT)(DG)(DA)(DT)(DT)(DC)(DT) (DT)(DC)(DT)(DC)
MassTheoretical: 13.4586 kDa
SourceSpecies: Cloning vector pBR322 (others)

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Experimental details

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Sample preparation

SpecimenSpecimen state: particle / Method: cryo EM
Sample solutionpH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 277 K / Humidity: 100 %

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Electron microscopy imaging

ImagingMicroscope: JEOL 3200FS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Electron dose: 62 e/Å2 / Illumination mode: OTHER
LensMagnification: 30000.0 X (nominal), 40323.0 X (calibrated) / Cs: 2 mm / Imaging mode: BRIGHT FIELD / Defocus: 1000.0 - 3000.0 nm / Energy filter: In-column Omega Filter
Specimen HolderModel: GATAN LIQUID NITROGEN / Temperature: K ( 100.0 - 100.0 K)
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 718 / Sampling size: 5 microns

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C2 (2 fold cyclic) / Number of projections: 30637
3D reconstructionAlgorithm: FOURIER SPACE / Software: RELION / Resolution: 6.35 Å / Resolution method: FSC 0.143 CUT-OFF
FSC plot
(resolution estimation)

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Atomic model buiding

Modeling #1Refinement protocol: flexible / Target criteria: correlation coefficient / Refinement space: REAL
Input PDB model: 4Z2C
Chain ID: A
Output model

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