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- EMDB-8827: Cas1-Cas2-IHF-DNA holo-complex -

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Basic information

Entry
Database: EMDB / ID: 8827
TitleCas1-Cas2-IHF-DNA holo-complex
SampleCas1-Cas2-IHF-DNA holo complex
SourceEscherichia coli / bacteria / エシェリキア・コリ, 大腸菌 /
Map dataCas1-Cas2-IHF-DNA holo-complex
Methodsingle particle reconstruction, at 3.64 Å resolution
AuthorsWright AV / Liu JJ
CitationScience, 2017, 357, 1113-1118

Science, 2017, 357, 1113-1118 Yorodumi Papers
Structures of the CRISPR genome integration complex.
Addison V Wright / Jun-Jie Liu / Gavin J Knott / Kevin W Doxzen / Eva Nogales / Jennifer A Doudna

Validation ReportPDB-ID: 5wfe

SummaryFull reportAbout validation report
DateDeposition: Jul 11, 2017 / Header (metadata) release: Aug 2, 2017 / Map release: Aug 2, 2017 / Last update: Sep 27, 2017

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Structure visualization

Movie
  • Surface view with section colored by density value
  • Surface level: 0.8
  • Imaged by UCSF CHIMERA
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  • Surface view colored by radius
  • Surface level: 0.8
  • Imaged by UCSF CHIMERA
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  • Surface view with fitted model
  • Atomic models: : PDB-5wfe
  • Surface level: 0.8
  • Imaged by UCSF CHIMERA
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3D viewer


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Supplemental images

Downloads & links

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Map

Fileemd_8827.map.gz (map file in CCP4 format, 108001 KB)
Projections & slices

Image control

Size
Brightness
Contrast
Others
AxesZ (Sec.)Y (Row.)X (Col.)
300 pix
1.07 Å/pix.
= 321. Å
300 pix
1.07 Å/pix.
= 321. Å
300 pix
1.07 Å/pix.
= 321. Å

Surface

Projections

Slices (1/3)

Slices (1/2)

Slices (2/3)

Images are generated by Spider package.

Voxel sizeX=Y=Z: 1.07 Å
Density
Contour Level:0.8 (by author), 0.8 (movie #1):
Minimum - Maximum-2.0546746 - 4.251078
Average (Standard dev.)0.00078146334 (0.08141421)
Details

EMDB XML:

Space Group Number1
Map Geometry
Axis orderXYZ
Dimensions300300300
Origin-150-150-150
Limit149149149
Spacing300300300
CellA=B=C: 321.00003 Å
α=β=γ: 90 deg.

CCP4 map header:

modeImage stored as Reals
Å/pix. X/Y/Z1.071.071.07
M x/y/z300300300
origin x/y/z0.0000.0000.000
length x/y/z321.000321.000321.000
α/β/γ90.00090.00090.000
start NX/NY/NZ
NX/NY/NZ
MAP C/R/S123
start NC/NR/NS-150-150-150
NC/NR/NS300300300
D min/max/mean-2.0554.2510.001

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Supplemental data

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Sample components

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Entire Cas1-Cas2-IHF-DNA holo complex

EntireName: Cas1-Cas2-IHF-DNA holo complex / Number of components: 9

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Component #1: protein, Cas1-Cas2-IHF-DNA holo complex

ProteinName: Cas1-Cas2-IHF-DNA holo complex / Recombinant expression: No
SourceSpecies: Escherichia coli / bacteria / エシェリキア・コリ, 大腸菌 /
Source (engineered)Expression System: Escherichia coli / bacteria / エシェリキア・コリ, 大腸菌 /

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Component #2: protein, CRISPR-associated endonuclease Cas1

ProteinName: CRISPR-associated endonuclease Cas1 / Details: Cas1 / Recombinant expression: No
MassTheoretical: 33.235418 kDa
Source (engineered)Expression System: Escherichia coli k-12 / bacteria / image: Escherichia coli
Strain: K12

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Component #3: protein, CRISPR-associated endoribonuclease Cas2

ProteinName: CRISPR-associated endoribonuclease Cas2 / Recombinant expression: No
MassTheoretical: 11.55327 kDa
Source (engineered)Expression System: Escherichia coli k-12 / bacteria / image: Escherichia coli
Strain: K12

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Component #4: nucleic-acid, DNA (28-MER)

Nucleic-acidName: DNA (28-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DA)(DA)(DA)(DC)(DA)(DC)(DC)(DA)(DG)(DA) (DA)(DC)(DG)(DA)(DG)(DT)(DA)(DG)(DT)(DA) (DA)(DA)(DT)(DT)(DG)(DG)(DG)(DC)
MassTheoretical: 8.680646 kDa

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Component #5: nucleic-acid, DNA (45-MER)

Nucleic-acidName: DNA (45-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DA)(DT)(DT)(DT)(DA)(DC)(DT)(DA)(DC)(DT) (DC)(DG)(DT)(DT)(DC)(DT)(DG)(DG)(DT)(DG) (DT)(DT)(DT)(DC)(DT)(DC)(DG)(DT)(DG)(DT) (DG)(DT)(DT)(DC)(DC)(DC)(DC)(DG)(DC)(DG) (DC)(DC)(DA)(DG)(DC)(DG)(DG)(DG)(DG)(DA) (DT)(DA)(DA)(DA)(DC)(DC)(DG)(DA)(DG)(DC) (DA)
MassTheoretical: 18.745965 kDa

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Component #6: nucleic-acid, DNA (76-MER)

Nucleic-acidName: DNA (76-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DT)(DG)(DC)(DT)(DC)(DG)(DG)(DT)(DT)(DT) (DA)(DT)(DC)(DC)(DC)(DC)(DG)(DC)(DT)(DG) (DG)(DC)(DG)(DC)(DG)(DG)(DG)(DG)(DA)(DA) (DC)(DA)(DC)(DT)(DC)(DT)(DA)(DA)(DA)(DC) (DA)(DT)(DA)(DA)(DC)(DC)(DT)(DA)(DT)(DT) (DA)(DT)(DT)(DA)(DA)(DT)(DT)(DA)(DA)(DT) (DG)(DA)(DT)(DT)(DT)(DT)(DT)(DT)(DA)(DA) (DG)(DC)(DC)(DA)(DG)(DT)(DC)(DA)(DC)(DA) (DA)(DT)(DC)(DT)(DA)(DC)(DC)(DA)(DA)(DC) (DT)(DT)(DT)(DA)(DT)
MassTheoretical: 29.101656 kDa

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Component #7: nucleic-acid, DNA (61-MER)

Nucleic-acidName: DNA (61-MER) / Class: DNA / Structure: OTHER / Synthetic: No
Sequence:
(DA)(DT)(DA)(DA)(DA)(DG)(DT)(DT)(DG)(DG) (DT)(DA)(DG)(DA)(DT)(DT)(DG)(DT)(DG)(DA) (DC)(DT)(DG)(DG)(DC)(DT)(DT)(DA)(DA)(DA) (DA)(DA)(DA)(DT)(DC)(DA)(DT)(DT)(DA)(DA) (DT)(DT)(DA)(DA)(DT)(DA)(DA)(DT)(DA)(DG) (DG)(DT)(DT)(DA)(DT)(DG)(DT)(DT)(DT)(DA) (DG)(DA)
MassTheoretical: 19.286447 kDa

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Component #8: protein, Integration host factor subunit alpha

ProteinName: Integration host factor subunit alpha / Recombinant expression: No
MassTheoretical: 11.373952 kDa
Source (engineered)Expression System: Escherichia coli s88 / bacteria / image: Escherichia coli
Strain: S88 / ExPEC

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Component #9: protein, Integration host factor subunit beta

ProteinName: Integration host factor subunit beta / Recombinant expression: No
MassTheoretical: 10.671178 kDa
Source (engineered)Expression System: Escherichia coli s88 / bacteria / image: Escherichia coli
Strain: S88 / ExPEC

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Experimental details

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Sample preparation

Specimen stateparticle
Sample solutionSpecimen conc.: 0.3 mg/ml
Buffer solution: 20 mM HEPES, pH 7.5, 150 mM KCl, 5 mM EDTA, 1 mM DTT, and 0.1% glycerol
pH: 7.5
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Temperature: 281.15 K / Humidity: 100 %

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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: 1.5 e/Å2 / Illumination mode: FLOOD BEAM
LensCs: 2.6 mm / Imaging mode: BRIGHT FIELD
Specimen HolderModel: FEI TITAN KRIOS AUTOGRID HOLDER
CameraDetector: GATAN K2 (4k x 4k)

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Image acquisition

Image acquisitionNumber of digital images: 3000

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Image processing

ProcessingMethod: single particle reconstruction / Applied symmetry: C1 (asymmetric) / Number of projections: 86000
3D reconstructionSoftware: cryoSPARC / Resolution: 3.64 Å / Resolution method: FSC 0.143 CUT-OFF

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Atomic model buiding

Output model

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