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- EMDB-10286: Cubic Dps-DNA crystal structure in vitro -

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Basic information

Entry
Database: EMDB / ID: EMD-10286
TitleCubic Dps-DNA crystal structure in vitro
Map data
Sample
  • Complex: Dps-DNA co-crystal
Biological speciesEscherichia coli BL21(DE3) (bacteria)
Methodsubtomogram averaging / cryo EM / Resolution: 13.7 Å
AuthorsChesnokov YM / Kamyshinsky RA / Vasiliev AL
Funding support Russian Federation, 1 items
OrganizationGrant numberCountry
Russian Science Foundation18-74-10071 Russian Federation
CitationJournal: Biomolecules / Year: 2019
Title: Polymorphic Protective Dps-DNA Co-Crystals by Cryo Electron Tomography and Small Angle X-Ray Scattering.
Authors: Roman Kamyshinsky / Yury Chesnokov / Liubov Dadinova / Andrey Mozhaev / Ivan Orlov / Maxim Petoukhov / Anton Orekhov / Eleonora Shtykova / Alexander Vasiliev /
Abstract: Rapid increase of intracellular synthesis of specific histone-like Dps protein that binds DNA to protect the genome against deleterious factors leads to in cellulo crystallization-one of the most ...Rapid increase of intracellular synthesis of specific histone-like Dps protein that binds DNA to protect the genome against deleterious factors leads to in cellulo crystallization-one of the most curious processes in the area of life science at the moment. However, the actual structure of the Dps-DNA co-crystals remained uncertain in the details for more than two decades. Cryo-electron tomography and small-angle X-ray scattering revealed polymorphous modifications of the co-crystals depending on the buffer parameters. Two different types of the Dps-DNA co-crystals are formed in vitro: triclinic and cubic. Three-dimensional reconstruction revealed DNA and Dps molecules in cubic co-crystals, and the unit cell parameters of cubic lattice were determined consistently by both methods.
History
DepositionSep 2, 2019-
Header (metadata) releaseJan 8, 2020-
Map releaseJan 8, 2020-
UpdateNov 25, 2020-
Current statusNov 25, 2020Processing site: PDBe / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.4
  • Imaged by UCSF Chimera
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  • Surface view colored by radius
  • Surface level: 0.4
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

emd_10286.png

Downloads & links

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Map

FileDownload / File: emd_10286.map.gz / Format: CCP4 / Size: 2.8 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Voxel sizeX=Y=Z: 3.7 Å
Density
Contour LevelBy AUTHOR: 0.4 / Movie #1: 0.4
Minimum - Maximum-1.2981853 - 1.8784196
Average (Standard dev.)0.02984969 (±0.4618922)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions909090
Spacing909090
CellA=B=C: 333.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z3.73.73.7
M x/y/z909090
origin x/y/z0.0000.0000.000
length x/y/z333.000333.000333.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS909090
D min/max/mean-1.2981.8780.030

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Supplemental data

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Mask #1

Fileemd_10286_msk_1.map
Projections & Slices
AxesZYX

Projections

Slices (1/2)
Density Histograms

Histogram

Histogram (log scale)

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Additional map: Map before any post-processing

Fileemd_10286_additional.map
AnnotationMap before any post-processing
Projections & Slices
AxesZYX

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Additional map: Map before any post-processing

Fileemd_10286_additional_1.map
AnnotationMap before any post-processing
Projections & Slices
AxesZYX

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Half map: #2

Fileemd_10286_half_map_1.map
Projections & Slices
AxesZYX

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Density Histograms

Histogram

Histogram (log scale)

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Half map: #1

Fileemd_10286_half_map_2.map
Projections & Slices
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Sample components

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Entire : Dps-DNA co-crystal

EntireName: Dps-DNA co-crystal
Components
  • Complex: Dps-DNA co-crystal

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Supramolecule #1: Dps-DNA co-crystal

SupramoleculeName: Dps-DNA co-crystal / type: complex / ID: 1 / Parent: 0
Details: Dps protein and DNA were mixed in the concentrations corresponding to the Dps/DNA ratio 1Dps dodecamer/60 bp of DNA. A ring vector pcDNA-hIRR-GFP 9900 bp was used as DNA sample.
Source (natural)Organism: Escherichia coli BL21(DE3) (bacteria)
Recombinant expressionOrganism: Escherichia coli BL21(DE3) (bacteria) / Recombinant plasmid: pET-dps
Molecular weightTheoretical: 224 KDa

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Experimental details

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Structure determination

Methodcryo EM
Processingsubtomogram averaging
Aggregation state3D array

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Sample preparation

Concentration3.1 mg/mL
BufferpH: 7.5 / Component:
ConcentrationFormula
20.0 mMTris-HCl
20.0 mMNaCl
VitrificationCryogen name: ETHANE / Chamber humidity: 95 % / Chamber temperature: 283 K / Instrument: FEI VITROBOT MARK IV / Details: blotting for 1.5 seconds.
Detailsweight Dps/DNA ratio = 3/1

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Electron microscopy

MicroscopeFEI TITAN KRIOS
Specialist opticsSpherical aberration corrector: Cs image corrector (CEOS, Germany)
Image recordingFilm or detector model: FEI FALCON II (4k x 4k) / Detector mode: INTEGRATING / Digitization - Dimensions - Width: 4096 pixel / Digitization - Dimensions - Height: 4096 pixel / Digitization - Sampling interval: 14.0 µm / Number grids imaged: 1 / Average exposure time: 1.0 sec. / Average electron dose: 1.02 e/Å2
Electron beamAcceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron opticsC2 aperture diameter: 100.0 µm / Calibrated magnification: 37837 / Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 0.01 mm / Nominal defocus max: 4.5 µm / Nominal defocus min: 3.0 µm / Nominal magnification: 18000
Sample stageSpecimen holder model: FEI TITAN KRIOS AUTOGRID HOLDER / Cooling holder cryogen: NITROGEN
Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company

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Image processing

Final reconstructionNumber classes used: 1 / Applied symmetry - Point group: C1 (asymmetric) / Algorithm: BACK PROJECTION / Resolution.type: BY AUTHOR / Resolution: 13.7 Å / Resolution method: FSC 0.143 CUT-OFF / Software - Name: RELION (ver. 2.1) / Number subtomograms used: 16372
ExtractionNumber tomograms: 15 / Number images used: 42548 / Reference model: 1DPS / Method: template matching
Software:
Namedetails
EMAN2 (ver. 2.22)selection
RELION (ver. 2.1)extraction
CTF correctionSoftware - Name: CTFFIND (ver. 4)
Details: 3D models for the contrast transfer function (CTF) in RELION2
Final 3D classificationNumber classes: 2 / Avg.num./class: 21274 / Software - Name: RELION (ver. 2.1)
Final angle assignmentType: MAXIMUM LIKELIHOOD / Software - Name: RELION (ver. 2.1)
FSC plot (resolution estimation)

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Atomic model buiding 1

Initial modelPDB ID:

1dps

RefinementSpace: REAL / Protocol: RIGID BODY FIT / Target criteria: Cross-correlation coefficient

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