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- EMDB-30186: EcoR124I-DNA in the Translocation State -

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Basic information

Entry
Database: EMDB / ID: EMD-30186
TitleEcoR124I-DNA in the Translocation State
Map data
SampleEcoR124I-DNA
Function / homology
Function and homology information


type I site-specific deoxyribonuclease / type I site-specific deoxyribonuclease activity / site-specific DNA-methyltransferase (adenine-specific) / site-specific DNA-methyltransferase (adenine-specific) activity / N-methyltransferase activity / DNA restriction-modification system / DNA binding / ATP binding
Similarity search - Function
Antirestriction protein ArdA, domain 2 / Antirestriction / Antirestriction protein (ArdA) / Antirestriction protein ArdA, domain 3 / Antirestriction protein ArdA, domain 1 / Restriction endonuclease, type I, HsdM / N6 adenine-specific DNA methyltransferase, N-terminal domain / Type I restriction modification DNA specificity domain / SWI2/SNF2 ATPase / Restriction endonuclease, type I, HsdR, N-terminal ...Antirestriction protein ArdA, domain 2 / Antirestriction / Antirestriction protein (ArdA) / Antirestriction protein ArdA, domain 3 / Antirestriction protein ArdA, domain 1 / Restriction endonuclease, type I, HsdM / N6 adenine-specific DNA methyltransferase, N-terminal domain / Type I restriction modification DNA specificity domain / SWI2/SNF2 ATPase / Restriction endonuclease, type I, HsdR, N-terminal / Type I restriction enzyme R protein N terminus (HSDR_N) / Type I restriction and modification enzyme - subunit R C terminal / HsdM N-terminal domain / SWI2/SNF2 ATPase / Type-1 restriction enzyme R protein, C-terminal / N6 adenine-specific DNA methyltransferase, N-terminal domain superfamily / Type I restriction modification DNA specificity domain / Restriction endonuclease, type I, HsdR / N-6 DNA Methylase / DNA methylase, adenine-specific / N-6 Adenine-specific DNA methylases signature. / DNA methylase, N-6 adenine-specific, conserved site / Superfamilies 1 and 2 helicase ATP-binding type-1 domain profile. / DEAD-like helicases superfamily / Helicase superfamily 1/2, ATP-binding domain / S-adenosyl-L-methionine-dependent methyltransferase / P-loop containing nucleoside triphosphate hydrolase
Similarity search - Domain/homology
Antirestriction protein ArdA / Type I restriction enzyme EcoR124II M protein / Type-1 restriction enzyme EcoR124II specificity protein / Type I restriction enzyme EcoR124II R protein / Type I restriction enzyme R Protein
Similarity search - Component
Biological speciesEscherichia coli (E. coli)
Methodsingle particle reconstruction / cryo EM / Resolution: 6.72 Å
AuthorsGao P / Gao Y
CitationJournal: Nat Microbiol / Year: 2020
Title: Structural insights into assembly, operation and inhibition of a type I restriction-modification system.
Authors: Yina Gao / Duanfang Cao / Jingpeng Zhu / Han Feng / Xiu Luo / Songqing Liu / Xiao-Xue Yan / Xinzheng Zhang / Pu Gao /
Abstract: Type I restriction-modification (R-M) systems are widespread in prokaryotic genomes and provide robust protection against foreign DNA. They are multisubunit enzymes with methyltransferase, ...Type I restriction-modification (R-M) systems are widespread in prokaryotic genomes and provide robust protection against foreign DNA. They are multisubunit enzymes with methyltransferase, endonuclease and translocase activities. Despite extensive studies over the past five decades, little is known about the molecular mechanisms of these sophisticated machines. Here, we report the cryo-electron microscopy structures of the representative EcoR124I R-M system in different assemblies (RMS, RMS and MS) bound to target DNA and the phage and mobile genetic element-encoded anti-restriction proteins Ocr and ArdA. EcoR124I can precisely regulate different enzymatic activities by adopting distinct conformations. The marked conformational transitions of EcoR124I are dependent on the intrinsic flexibility at both the individual-subunit and assembled-complex levels. Moreover, Ocr and ArdA use a DNA-mimicry strategy to inhibit multiple activities, but do not block the conformational transitions of the complexes. These structural findings, complemented by mutational studies of key intermolecular contacts, provide insights into assembly, operation and inhibition mechanisms of type I R-M systems.
History
DepositionApr 2, 2020-
Header (metadata) releaseMay 27, 2020-
Map releaseMay 27, 2020-
UpdateSep 16, 2020-
Current statusSep 16, 2020Processing site: PDBj / Status: Released

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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  • Surface view colored by cylindrical radius
  • Surface level: 0.04
  • Imaged by UCSF Chimera
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Structure viewerEM map:
SurfViewMolmilJmol/JSmol
Supplemental images

Downloads & links

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Map

FileDownload / File: emd_30186.map.gz / Format: CCP4 / Size: 42.9 MB / Type: IMAGE STORED AS FLOATING POINT NUMBER (4 BYTES)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
1.35 Å/pix.
x 224 pix.
= 302.4 Å
1.35 Å/pix.
x 224 pix.
= 302.4 Å
1.35 Å/pix.
x 224 pix.
= 302.4 Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider.

Voxel sizeX=Y=Z: 1.35 Å
Density
Contour LevelBy AUTHOR: 0.04 / Movie #1: 0.04
Minimum - Maximum-0.020598693 - 0.12077804
Average (Standard dev.)0.0011962651 (±0.008593206)
SymmetrySpace group: 1
Details

EMDB XML:

Map geometry
Axis orderXYZ
Origin-112-112-112
Dimensions224224224
Spacing224224224
CellA=B=C: 302.4 Å
α=β=γ: 90.0 °

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.351.351.35
M x/y/z224224224
origin x/y/z0.0000.0000.000
length x/y/z302.400302.400302.400
α/β/γ90.00090.00090.000
MAP C/R/S123
start NC/NR/NS-112-112-112
NC/NR/NS224224224
D min/max/mean-0.0210.1210.001

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Supplemental data

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Sample components

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Entire EcoR124I-DNA

EntireName: EcoR124I-DNA / Number of Components: 1

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Component #1: protein, EcoR124I-DNA

ProteinName: EcoR124I-DNA / Recombinant expression: No
SourceSpecies: Escherichia coli (E. coli)
Source (engineered)Expression System: Escherichia coli (E. coli)

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Experimental details

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Sample preparation

SpecimenSpecimen State: Particle / Method: cryo EM
Sample solutionpH: 7.5
VitrificationCryogen Name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
ImagingMicroscope: FEI TITAN KRIOS
Electron gunElectron Source: FIELD EMISSION GUN / Accelerating Voltage: 300 kV / Electron Dose: 60 e/Å2 / Illumination Mode: FLOOD BEAM
LensImaging Mode: BRIGHT FIELD
Specimen HolderModel: OTHER
CameraDetector: GATAN K2 SUMMIT (4k x 4k)

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Image processing

ProcessingMethod: single particle reconstruction / Number of Projections: 15236
3D reconstructionResolution: 6.72 Å / Resolution Method: FSC 0.143 CUT-OFF

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