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- PDB-8inb: Cryo-EM structure of Cas12j-SF05-crRNA-dsDNA complex -

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Basic information

Entry
Database: PDB / ID: 8inb
TitleCryo-EM structure of Cas12j-SF05-crRNA-dsDNA complex
Components
  • Cas12j-SF05
  • NTS-DNA
  • TS-DNA
  • crRNA
KeywordsRNA BINDING PROTEIN/RNA/DNA / CRISPR / RNA BINDING PROTEIN-RNA complex / RNA BINDING PROTEIN / RNA BINDING PROTEIN-RNA-DNA complex
Function / homologyDNA / DNA (> 10) / RNA / RNA (> 10)
Function and homology information
Biological speciesBiggievirus Mos11
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 3.1 Å
AuthorsZhang, X. / Duan, Z.Q. / Zhu, J.K.
Funding support China, 1items
OrganizationGrant numberCountry
National Science Foundation (NSF, China) China
CitationJournal: Cell Discov / Year: 2023
Title: Molecular basis for DNA cleavage by the hypercompact Cas12j-SF05.
Authors: Zhiqiang Duan / Xi Zhang / Jun-Tao Zhang / Shanshan Li / Ruiheng Liu / Jie Sun / Qingzhi Zhao / Nannan Jia / Ning Jia / Jian-Kang Zhu /
History
DepositionMar 9, 2023Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Mar 13, 2024Provider: repository / Type: Initial release
Revision 1.1Mar 20, 2024Group: Database references / Category: citation / citation_author
Item: _citation.country / _citation.journal_abbrev ..._citation.country / _citation.journal_abbrev / _citation.journal_id_CSD / _citation.journal_id_ISSN / _citation.journal_volume / _citation.page_first / _citation.page_last / _citation.pdbx_database_id_DOI / _citation.pdbx_database_id_PubMed / _citation.title / _citation.year

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

Downloads & links

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Assembly

Deposited unit
A: Cas12j-SF05
B: crRNA
C: TS-DNA
D: NTS-DNA


Theoretical massNumber of molelcules
Total (without water)135,5254
Polymers135,5254
Non-polymers00
Water0
1


  • Idetical with deposited unit
  • defined by author
  • Evidence: gel filtration
TypeNameSymmetry operationNumber
identity operation1_5551

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Components

#1: Protein Cas12j-SF05


Mass: 83449.094 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) Biggievirus Mos11 / Production host: Escherichia coli BL21(DE3) (bacteria)
#2: RNA chain crRNA


Mass: 18179.771 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Biggievirus Mos11
#3: DNA chain TS-DNA


Mass: 16994.910 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Biggievirus Mos11
#4: DNA chain NTS-DNA


Mass: 16900.799 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) Biggievirus Mos11

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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: Ternary complex of Cas12j-SF05 with crRNA and target dsDNA
Type: COMPLEX / Entity ID: all / Source: RECOMBINANT
Source (natural)Organism: Biggievirus Mos11
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.4
SpecimenEmbedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
VitrificationCryogen name: ETHANE

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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 FIELDBright-field microscopy / Nominal defocus max: 2500 nm / Nominal defocus min: 1000 nm
Image recordingElectron dose: 50 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 3.1 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 59719 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0035569
ELECTRON MICROSCOPYf_angle_d0.5227833
ELECTRON MICROSCOPYf_dihedral_angle_d19.1951265
ELECTRON MICROSCOPYf_chiral_restr0.037898
ELECTRON MICROSCOPYf_plane_restr0.006775

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