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RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2

Author

Listed:
  • Jack P. K. Bravo

    (University of Texas at Austin)

  • Thomson Hallmark

    (Utah State University)

  • Bronson Naegle

    (Utah State University)

  • Chase L. Beisel

    (Helmholtz Institute for RNA-based Infection Research (HIRI), Helmholtz-Centre for Infection Research (HZI)
    University of Würzburg)

  • Ryan N. Jackson

    (Utah State University)

  • David W. Taylor

    (University of Texas at Austin
    University of Texas at Austin
    University of Texas at Austin
    LIVESTRONG Cancer Institutes, Dell Medical School)

Abstract

Cas12a2 is a CRISPR-associated nuclease that performs RNA-guided, sequence-nonspecific degradation of single-stranded RNA, single-stranded DNA and double-stranded DNA following recognition of a complementary RNA target, culminating in abortive infection1. Here we report structures of Cas12a2 in binary, ternary and quaternary complexes to reveal a complete activation pathway. Our structures reveal that Cas12a2 is autoinhibited until binding a cognate RNA target, which exposes the RuvC active site within a large, positively charged cleft. Double-stranded DNA substrates are captured through duplex distortion and local melting, stabilized by pairs of ‘aromatic clamp’ residues that are crucial for double-stranded DNA degradation and in vivo immune system function. Our work provides a structural basis for this mechanism of abortive infection to achieve population-level immunity, which can be leveraged to create rational mutants that degrade a spectrum of collateral substrates.

Suggested Citation

  • Jack P. K. Bravo & Thomson Hallmark & Bronson Naegle & Chase L. Beisel & Ryan N. Jackson & David W. Taylor, 2023. "RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2," Nature, Nature, vol. 613(7944), pages 582-587, January.
    • Handle: RePEc:nat:nature:v:613:y:2023:i:7944:d:10.1038_s41586-022-05560-w
    DOI: 10.1038/s41586-022-05560-w
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