Structure of the mini-RNA-guided endonuclease CRISPR-Cas12j3

Research output: Contribution to journal › Journal article › Research › peer-review

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Structure of the mini-RNA-guided endonuclease CRISPR-Cas12j3. / Carabias, Arturo; Fuglsang, Anders; Temperini, Piero; Pape, Tillmann; Sofos, Nicholas; Stella, Stefano; Erlendsson, Simon; Montoya, Guillermo.

In: Nature Communications, Vol. 12, 4476, 2021.

Research output: Contribution to journal › Journal article › Research › peer-review

Harvard

Carabias, A, Fuglsang, A, Temperini, P, Pape, T, Sofos, N, Stella, S, Erlendsson, S & Montoya, G 2021, 'Structure of the mini-RNA-guided endonuclease CRISPR-Cas12j3', Nature Communications, vol. 12, 4476. https://doi.org/10.1038/s41467-021-24707-3

APA

Carabias, A., Fuglsang, A., Temperini, P., Pape, T., Sofos, N., Stella, S., Erlendsson, S., & Montoya, G. (2021). Structure of the mini-RNA-guided endonuclease CRISPR-Cas12j3. Nature Communications, 12, [4476]. https://doi.org/10.1038/s41467-021-24707-3

Vancouver

Carabias A, Fuglsang A, Temperini P, Pape T, Sofos N, Stella S et al. Structure of the mini-RNA-guided endonuclease CRISPR-Cas12j3. Nature Communications. 2021;12. 4476. https://doi.org/10.1038/s41467-021-24707-3

Author

Carabias, Arturo ; Fuglsang, Anders ; Temperini, Piero ; Pape, Tillmann ; Sofos, Nicholas ; Stella, Stefano ; Erlendsson, Simon ; Montoya, Guillermo. / Structure of the mini-RNA-guided endonuclease CRISPR-Cas12j3. In: Nature Communications. 2021 ; Vol. 12.

Bibtex

@article{dd6a5b10b2cd4ac59ecea5ba9dfc0b17,
title = "Structure of the mini-RNA-guided endonuclease CRISPR-Cas12j3",
abstract = "CRISPR-Cas12j is a recently identified family of miniaturized RNA-guided endonucleases from phages. These ribonucleoproteins provide a compact scaffold gathering all key activities of a genome editing tool. We provide the first structural insight into the Cas12j family by determining the cryoEM structure of Cas12j3/R-loop complex after DNA cleavage. The structure reveals the machinery for PAM recognition, hybrid assembly and DNA cleavage. The crRNA-DNA hybrid is directed to the stop domain that splits the hybrid, guiding the T-strand towards the catalytic site. The conserved RuvC insertion is anchored in the stop domain and interacts along the phosphate backbone of the crRNA in the hybrid. The assembly of a hybrid longer than 12-nt activates catalysis through key functional residues in the RuvC insertion. Our findings suggest why Cas12j unleashes unspecific ssDNA degradation after activation. A site-directed mutagenesis analysis supports the DNA cutting mechanism, providing new avenues to redesign CRISPR-Cas12j nucleases for genome editing.",
keywords = "Bacteriophages/enzymology, CRISPR-Associated Proteins/chemistry, CRISPR-Cas Systems, Catalytic Domain, Cryoelectron Microscopy, DNA Cleavage, Endodeoxyribonucleases/chemistry, Escherichia coli Proteins/chemistry, Gene Editing, Models, Molecular, Mutagenesis, Site-Directed, Protein Conformation, RNA, Guide/genetics, RNA, Viral/genetics",
author = "Arturo Carabias and Anders Fuglsang and Piero Temperini and Tillmann Pape and Nicholas Sofos and Stefano Stella and Simon Erlendsson and Guillermo Montoya",
note = "{\textcopyright} 2021. The Author(s).",
year = "2021",
doi = "10.1038/s41467-021-24707-3",
language = "English",
volume = "12",
journal = "Nature Communications",
issn = "2041-1723",
publisher = "nature publishing group",

}

RIS

TY - JOUR

T1 - Structure of the mini-RNA-guided endonuclease CRISPR-Cas12j3

AU - Carabias, Arturo

AU - Fuglsang, Anders

AU - Temperini, Piero

AU - Pape, Tillmann

AU - Sofos, Nicholas

AU - Stella, Stefano

AU - Erlendsson, Simon

AU - Montoya, Guillermo

N1 - © 2021. The Author(s).

PY - 2021

Y1 - 2021

N2 - CRISPR-Cas12j is a recently identified family of miniaturized RNA-guided endonucleases from phages. These ribonucleoproteins provide a compact scaffold gathering all key activities of a genome editing tool. We provide the first structural insight into the Cas12j family by determining the cryoEM structure of Cas12j3/R-loop complex after DNA cleavage. The structure reveals the machinery for PAM recognition, hybrid assembly and DNA cleavage. The crRNA-DNA hybrid is directed to the stop domain that splits the hybrid, guiding the T-strand towards the catalytic site. The conserved RuvC insertion is anchored in the stop domain and interacts along the phosphate backbone of the crRNA in the hybrid. The assembly of a hybrid longer than 12-nt activates catalysis through key functional residues in the RuvC insertion. Our findings suggest why Cas12j unleashes unspecific ssDNA degradation after activation. A site-directed mutagenesis analysis supports the DNA cutting mechanism, providing new avenues to redesign CRISPR-Cas12j nucleases for genome editing.

AB - CRISPR-Cas12j is a recently identified family of miniaturized RNA-guided endonucleases from phages. These ribonucleoproteins provide a compact scaffold gathering all key activities of a genome editing tool. We provide the first structural insight into the Cas12j family by determining the cryoEM structure of Cas12j3/R-loop complex after DNA cleavage. The structure reveals the machinery for PAM recognition, hybrid assembly and DNA cleavage. The crRNA-DNA hybrid is directed to the stop domain that splits the hybrid, guiding the T-strand towards the catalytic site. The conserved RuvC insertion is anchored in the stop domain and interacts along the phosphate backbone of the crRNA in the hybrid. The assembly of a hybrid longer than 12-nt activates catalysis through key functional residues in the RuvC insertion. Our findings suggest why Cas12j unleashes unspecific ssDNA degradation after activation. A site-directed mutagenesis analysis supports the DNA cutting mechanism, providing new avenues to redesign CRISPR-Cas12j nucleases for genome editing.

KW - Bacteriophages/enzymology

KW - CRISPR-Associated Proteins/chemistry

KW - CRISPR-Cas Systems

KW - Catalytic Domain

KW - Cryoelectron Microscopy

KW - DNA Cleavage

KW - Endodeoxyribonucleases/chemistry

KW - Escherichia coli Proteins/chemistry

KW - Gene Editing

KW - Models, Molecular

KW - Mutagenesis, Site-Directed

KW - Protein Conformation

KW - RNA, Guide/genetics

KW - RNA, Viral/genetics

U2 - 10.1038/s41467-021-24707-3

DO - 10.1038/s41467-021-24707-3

M3 - Journal article

C2 - 34294706

VL - 12

JO - Nature Communications

JF - Nature Communications

SN - 2041-1723

M1 - 4476

ER -

ID: 275949393