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- EMDB-23543: cryoEM structure DrdV-DNA complex -

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Open data


ID or keywords:

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Basic information

Entry
Database: EMDB / ID: EMD-23543
TitlecryoEM structure DrdV-DNA complex
Map data
SampleDrdVDNA complex
  • Site-specific DNA-methyltransferase (adenine-specific)DNA methyltransferase
  • (nucleic-acidNucleic acid) x 2
  • (ligand) x 2
Function / homology
Function and homology information


site-specific DNA-methyltransferase (adenine-specific) / site-specific DNA-methyltransferase (adenine-specific) activity / N-methyltransferase activity / endonuclease activity / DNA binding
Similarity search - Function
Type ISP C-terminal specificity domain / Type ISP restriction-modification enzyme LLaBIII, C-terminal specificity domain / N-6 DNA Methylase / DNA methylase, adenine-specific / N-6 Adenine-specific DNA methylases signature. / DNA methylase, N-6 adenine-specific, conserved site / S-adenosyl-L-methionine-dependent methyltransferase
Similarity search - Domain/homology
Site-specific DNA-methyltransferase (adenine-specific)
Similarity search - Component
Biological speciesDeinococcus wulumuqiensis (bacteria) / synthetic construct (others)
Methodsingle particle reconstruction / cryo EM / Resolution: 3.25 Å
AuthorsShen BW / Stoddard BL
Funding support United States, 1 items
OrganizationGrant numberCountry
National Institutes of Health/National Institute of General Medical Sciences (NIH/NIGMS)GM105691 United States
CitationJournal: Structure / Year: 2021
Title: Coordination of phage genome degradation versus host genome protection by a bifunctional restriction-modification enzyme visualized by CryoEM.
Authors: Betty W Shen / Joel D Quispe / Yvette Luyten / Benjamin E McGough / Richard D Morgan / Barry L Stoddard /
Abstract: Restriction enzymes that combine methylation and cleavage into a single assemblage and modify one DNA strand are capable of efficient adaptation toward novel targets. However, they must reliably ...Restriction enzymes that combine methylation and cleavage into a single assemblage and modify one DNA strand are capable of efficient adaptation toward novel targets. However, they must reliably cleave invasive DNA and methylate newly replicated unmodified host sites. One possible solution is to enforce a competition between slow methylation at a single unmodified host target, versus faster cleavage that requires multiple unmodified target sites in foreign DNA to be brought together in a reaction synapse. To examine this model, we have determined the catalytic behavior of a bifunctional type IIL restriction-modification enzyme and determined its structure, via cryoelectron microscopy, at several different stages of assembly and coordination with bound DNA targets. The structures demonstrate a mechanism in which an initial dimer is formed between two DNA-bound enzyme molecules, positioning the endonuclease domain from each enzyme against the other's DNA and requiring further additional DNA-bound enzyme molecules to enable cleavage.
History
DepositionFeb 26, 2021-
Header (metadata) releaseMar 17, 2021-
Map releaseMar 17, 2021-
UpdateJun 16, 2021-
Current statusJun 16, 2021Processing site: RCSB / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.375
  • Imaged by UCSF Chimera
  • Download
  • Surface view colored by cylindrical radius
  • Surface level: 0.375
  • Imaged by UCSF Chimera
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  • Surface view with fitted model
  • Atomic models: PDB-7lvv
  • Surface level: 0.375
  • Imaged by UCSF Chimera
  • Download
Movie viewer
Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_23543.map.gz / Format: CCP4 / Size: 244.1 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.16 Å/pix.
x 400 pix.
= 464. Å
1.16 Å/pix.
x 400 pix.
= 464. Å
1.16 Å/pix.
x 400 pix.
= 464. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.16 Å
Density
Contour LevelBy AUTHOR: 0.375 / Movie #1: 0.375
Minimum - Maximum-0.7445625 - 2.2025354
Average (Standard dev.)0.00035449766 (±0.05004532)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin000
Dimensions400400400
Spacing400400400
CellA=B=C: 464.0 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.161.161.16
M x/y/z400400400
origin x/y/z0.0000.0000.000
length x/y/z464.000464.000464.000
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS000
NC/NR/NS400400400
D min/max/mean-0.7452.2030.000

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Supplemental data

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Sample components

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Entire DrdVDNA complex

EntireName: DrdVDNA complex / Details: dimer of DrdVDNA complex / Number of Components: 6

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Component #1: protein, DrdVDNA complex

ProteinName: DrdVDNA complex / Details: dimer of DrdVDNA complex / Recombinant expression: No
MassTheoretical: 240 kDa
SourceSpecies: Deinococcus wulumuqiensis (bacteria)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #2: protein, Site-specific DNA-methyltransferase (adenine-specific)

ProteinName: Site-specific DNA-methyltransferase (adenine-specific)DNA methyltransferase
Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 118.256859 kDa
SourceSpecies: Deinococcus wulumuqiensis (bacteria)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Component #3: nucleic-acid, DNA (28-MER)

nucleic acidName: DNA (28-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DC)(DA)(DG)(DC)(DC)(DC)(DA)(DT)(DG)(DG) (DA)(DC)(DC)(DC)(DA)(DG)(DA)(DA)(DC)(DC) (DA)(DC)(DC)(DC)(DA)(DC)(DC)(DC)(DG)
MassTheoretical: 8.748646 kDa
SourceSpecies: synthetic construct (others)

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Component #4: nucleic-acid, DNA (27-MER)

nucleic acidName: DNA (27-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DG)(DG)(DG)(DT)(DG)(DG)(DG)(DT)(DG)(DG) (DT)(DT)(DC)(DT)(DG)(DG)(DG)(DT)(DC)(DC) (DA)(DT)(DG)(DG)(DG)(DC)(DT)(DG)(DC)
MassTheoretical: 9.085788 kDa
SourceSpecies: synthetic construct (others)

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Component #5: ligand, S-ADENOSYLMETHIONINE

LigandName: S-ADENOSYLMETHIONINES-Adenosyl methionine / Number of Copies: 2 / Recombinant expression: No
MassTheoretical: 0.398437 kDa

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Component #6: ligand, CALCIUM ION

LigandName: CALCIUM IONCalcium / Number of Copies: 4 / Recombinant expression: No
MassTheoretical: 4.007805 MDa

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Experimental details

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Sample preparation

SpecimenSpecimen State: Particle / Method: cryo EM
Sample solutionSpecimen conc.: 0.25 mg/mL / Buffer solution: 20 nM HEPES pH7.5 150 NaCl, 2 mM CaCl2 / pH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen Name: ETHANE / Temperature: 298 K / Humidity: 95 %

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Electron microscopy imaging

ImagingMicroscope: TFS GLACIOS
Electron gunElectron Source: FIELD EMISSION GUN / Accelerating Voltage: 200 kV / Electron Dose: 40 e/Å2 / Illumination Mode: SPOT SCAN
LensMagnification: 37000.0 X (nominal) / Cs: 2.7 mm / Imaging Mode: BRIGHT FIELD / Defocus: 800.0 - 2300.0 nm
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER / Temperature: (70.0 - 70.0 K)
CameraDetector: FEI CETA (4k x 4k)

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Image acquisition

Image acquisitionNumber of Digital Images: 1200
Details: images were collected in movie-mode at 5 frames per second

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Image processing

ProcessingMethod: single particle reconstruction / Applied Symmetry: C1 (asymmetric) / Number of Projections: 34554
3D reconstructionSoftware: cryoSPARC / Resolution: 3.25 Å / Resolution Method: FSC 0.143 CUT-OFF

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Atomic model buiding

Output model

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