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- PDB-6v9q: Cryo-EM structure of Cascade-TniQ binary complex -

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Basic information

Entry
Database: PDB / ID: 6v9q
TitleCryo-EM structure of Cascade-TniQ binary complex
Components
  • (Type I-F CRISPR-associated ...) x 2
  • Cas8
  • RNA (61-MER)
  • TniQ family protein
KeywordsIMMUNE SYSTEM / CRISPR-Cas system / Cascade- TniQ
Function / homologyCRISPR-associated endoribonuclease Cas6/Csy4 / Alpha-Beta Plaits / 2-Layer Sandwich / Alpha Beta / RNA / RNA (> 10)
Function and homology information
Biological speciesVibrio cholerae (bacteria)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.9 Å
AuthorsJia, N. / Patel, D.J.
CitationJournal: Cell Res / Year: 2020
Title: Structure-function insights into the initial step of DNA integration by a CRISPR-Cas-Transposon complex.
Authors: Ning Jia / Wei Xie / M Jason de la Cruz / Edward T Eng / Dinshaw J Patel /
History
DepositionDec 15, 2019Deposition site: RCSB / Processing site: RCSB
Revision 1.0Jan 29, 2020Provider: repository / Type: Initial release
Revision 1.1Feb 19, 2020Group: Database references / Category: citation
Item: _citation.journal_volume / _citation.page_first / _citation.page_last
Revision 1.2Mar 6, 2024Group: Data collection / Database references / Category: chem_comp_atom / chem_comp_bond / database_2
Item: _database_2.pdbx_DOI / _database_2.pdbx_database_accession

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Structure visualization

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Assembly

Deposited unit
K: RNA (61-MER)
A: Cas8
J: TniQ family protein
I: TniQ family protein
G: Type I-F CRISPR-associated protein Csy3
B: Type I-F CRISPR-associated protein Csy3
C: Type I-F CRISPR-associated protein Csy3
D: Type I-F CRISPR-associated protein Csy3
E: Type I-F CRISPR-associated protein Csy3
F: Type I-F CRISPR-associated protein Csy3
H: Type I-F CRISPR-associated endoribonuclease Cas6/Csy4
hetero molecules


Theoretical massNumber of molelcules
Total (without water)445,76515
Polymers445,50411
Non-polymers2624
Water00
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551
Buried area51290 Å2
ΔGint-342 kcal/mol
Surface area142890 Å2

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Components

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Protein , 2 types, 3 molecules AJI

#2: Protein Cas8


Mass: 72294.930 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vibrio cholerae (bacteria) / Gene: FXE93_08750 / Production host: Escherichia coli (E. coli)
#3: Protein TniQ family protein


Mass: 45597.867 Da / Num. of mol.: 2
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vibrio cholerae (bacteria) / Gene: FXE93_08745 / Production host: Escherichia coli (E. coli)

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Type I-F CRISPR-associated ... , 2 types, 7 molecules GBCDEFH

#4: Protein
Type I-F CRISPR-associated protein Csy3


Mass: 39886.031 Da / Num. of mol.: 6
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vibrio cholerae (bacteria) / Gene: csy3, FXE93_08755 / Production host: Escherichia coli (E. coli)
#5: Protein Type I-F CRISPR-associated endoribonuclease Cas6/Csy4


Mass: 23130.430 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vibrio cholerae (bacteria) / Gene: cas6f, FXE93_08760 / Production host: Escherichia coli (E. coli)

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RNA chain / Non-polymers , 2 types, 5 molecules K

#1: RNA chain RNA (61-MER)


Mass: 19566.543 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Vibrio cholerae (bacteria) / Production host: Escherichia coli (E. coli)
#6: Chemical
ChemComp-ZN / ZINC ION


Mass: 65.409 Da / Num. of mol.: 4 / Source method: obtained synthetically / Formula: Zn

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Details

Has ligand of interestN

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Experimental details

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Experiment

ExperimentMethod: ELECTRON MICROSCOPY
EM experimentAggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction

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Sample preparation

ComponentName: Cascade-TniQ binary complex / Type: COMPLEX / Entity ID: #1-#5 / Source: RECOMBINANT
Molecular weightValue: 0.4 MDa / Experimental value: YES
Source (natural)Organism: Vibrio cholerae (bacteria)
Source (recombinant)Organism: Escherichia coli (E. coli)
Buffer solutionpH: 7.5 / Details: 20 mM HEPES, pH 7.5, 150 mM NaCl, 2 mM DTT
Buffer componentFormula: Tris
SpecimenConc.: 0.5 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationInstrument: FEI VITROBOT MARK IV / Cryogen name: ETHANE / Humidity: 100 %

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: FEI TITAN KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD
Image recordingElectron dose: 2.16 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM softwareName: RELION / Version: 2.1 / Category: 3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 134856 / Symmetry type: POINT

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