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- PDB-9jfs: Structure of Cas12p-TrxA-guide RNA-target DNA complex(29nt TS and... -

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Basic information

Entry
Database: PDB / ID: 9jfs
TitleStructure of Cas12p-TrxA-guide RNA-target DNA complex(29nt TS and 11nt NTS)
Components
  • Cas12p
  • DNA (29-MER)
  • DNA (5'-D(P*TP*CP*AP*AP*CP*TP*GP*TP*AP*TP*G)-3')
  • RNA (246-MER)
  • Thioredoxin
KeywordsRNA BINDING PROTEIN/RNA/DNA / CRISPR-Cas / nuclease / Heterodimer / RNA BINDING PROTEIN-RNA-DNA complex
Function / homologyDNA / DNA (> 10) / RNA / RNA (> 10) / RNA (> 100) / :
Function and homology information
Biological speciesunidentified (others)
Escherichia coli BL21 (bacteria)
synthetic construct (others)
MethodELECTRON MICROSCOPY / single particle reconstruction / cryo EM / Resolution: 2.67 Å
AuthorsWang, Z.P. / Wang, Y.J. / Ji, Q.J.
Funding support China, 1items
OrganizationGrant numberCountry
National Natural Science Foundation of China (NSFC)22277078 China
CitationJournal: To Be Published
Title: Host thioredoxin is a CRISPR enhancer of phage-encoded miniature Cas12p nucleases
Authors: Wang, Z.P. / Wang, Y.J. / Ji, Q.J.
History
DepositionSep 5, 2024Deposition site: PDBJ / Processing site: PDBC
Revision 1.0Sep 10, 2025Provider: repository / Type: Initial release
Revision 1.0Sep 10, 2025Data content type: EM metadata / Data content type: EM metadata / Provider: repository / Type: Initial release
Revision 1.0Sep 10, 2025Data content type: Additional map / Part number: 1 / Data content type: Additional map / Provider: repository / Type: Initial release
Revision 1.0Sep 10, 2025Data content type: Half map / Part number: 1 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Sep 10, 2025Data content type: Half map / Part number: 2 / Data content type: Half map / Provider: repository / Type: Initial release
Revision 1.0Sep 10, 2025Data content type: Image / Data content type: Image / Provider: repository / Type: Initial release
Revision 1.0Sep 10, 2025Data content type: Primary map / Data content type: Primary map / Provider: repository / Type: Initial release

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Structure visualization

Structure viewerMolecule:
MolmilJmol/JSmol

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Assembly

Deposited unit
A: Cas12p
B: RNA (246-MER)
C: DNA (29-MER)
D: DNA (5'-D(P*TP*CP*AP*AP*CP*TP*GP*TP*AP*TP*G)-3')
E: Thioredoxin


Theoretical massNumber of molelcules
Total (without water)188,3735
Polymers188,3735
Non-polymers00
Water00
1


  • Idetical with deposited unit
  • defined by author&software
  • Evidence: electron microscopy, not applicable
TypeNameSymmetry operationNumber
identity operation1_555x,y,z1

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Components

#1: Protein Cas12p


Mass: 84964.945 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) unidentified (others) / Production host: Escherichia coli BL21(DE3) (bacteria)
#2: RNA chain RNA (246-MER)


Mass: 79190.891 Da / Num. of mol.: 1
Source method: isolated from a genetically manipulated source
Source: (gene. exp.) unidentified (others) / Production host: Escherichia coli BL21(DE3) (bacteria)
#3: DNA chain DNA (29-MER)


Mass: 9049.875 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#4: DNA chain DNA (5'-D(P*TP*CP*AP*AP*CP*TP*GP*TP*AP*TP*G)-3')


Mass: 3348.209 Da / Num. of mol.: 1 / Source method: obtained synthetically / Source: (synth.) synthetic construct (others)
#5: Protein Thioredoxin


Mass: 11818.582 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Escherichia coli BL21(DE3) (bacteria) / References: UniProt: A0A140SSC0
Has protein modificationY

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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: complex of Cas12p-TrxA-gRNA-target DNA(29-nt TS and 11-nt NTS)
Type: COMPLEX / Entity ID: all / Source: MULTIPLE SOURCES
Source (natural)Organism: unidentified (others)
Source (recombinant)Organism: Escherichia coli BL21(DE3) (bacteria)
Buffer solutionpH: 7.5
SpecimenConc.: 3 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen supportGrid material: GOLD / Grid mesh size: 300 divisions/in. / Grid type: Homemade
VitrificationCryogen name: ETHANE

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Electron microscopy imaging

Experimental equipment
Model: Titan Krios / Image courtesy: FEI Company
MicroscopyModel: TFS KRIOS
Electron gunElectron source: FIELD EMISSION GUN / Accelerating voltage: 300 kV / Illumination mode: FLOOD BEAM
Electron lensMode: BRIGHT FIELD / Nominal defocus max: 1800 nm / Nominal defocus min: 1200 nm
Image recordingElectron dose: 20 e/Å2 / Film or detector model: GATAN K3 (6k x 4k)

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Processing

EM software
IDNameVersionCategory
1cryoSPARCparticle selection
9PHENIX1.21.1_5286:model refinement
13cryoSPARC3D reconstruction
CTF correctionType: PHASE FLIPPING AND AMPLITUDE CORRECTION
3D reconstructionResolution: 2.67 Å / Resolution method: FSC 0.143 CUT-OFF / Num. of particles: 508876 / Symmetry type: POINT
Refine LS restraints
Refine-IDTypeDev idealNumber
ELECTRON MICROSCOPYf_bond_d0.0059337
ELECTRON MICROSCOPYf_angle_d0.51313438
ELECTRON MICROSCOPYf_dihedral_angle_d19.0014382
ELECTRON MICROSCOPYf_chiral_restr0.0391613
ELECTRON MICROSCOPYf_plane_restr0.0041071

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