Title
RNA targeting unleashes indiscriminate nuclease activity of CRISPR–Cas12a2
Author
Jack P. K. Bravo, Thomson Hallmark, Bronson Naegle, Chase L. Beisel, Ryan N. Jackson & David W. Taylor
Year
2023
Journal
Nature
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.
Full Article
Instrument
J-1500
Keywords
RNA, nuclease activity, CRISPR–Cas12a2