High-Resolution Structure of Cas13b and Biochemical Characterization of RNA Targeting and Cleavage

Research output: Contribution to journalJournal articleResearchpeer-review

  • Ian M Slaymaker
  • Mesa, Pablo
  • Max J Kellner
  • Soumya Kannan
  • Edward Brignole
  • Jeremy Koob
  • Patricia R Feliciano
  • Stefano Stella
  • Omar O Abudayyeh
  • Jonathan S Gootenberg
  • Jonathan Strecker
  • Montoya, Guillermo
  • Feng Zhang

Type VI CRISPR-Cas systems contain programmable single-effector RNA-guided RNases, including Cas13b, one of the four known family members. Cas13b, which has been used for both RNA editing and nucleic acid detection, is unique among type VI CRISPR effectors in its linear domain architecture and CRISPR RNA (crRNA) structure. Here, we report the crystal structure of Prevotella buccae Cas13b (PbuCas13b) bound to crRNA at 1.65 Å resolution. This structure, combined with biochemical experiments assaying the stability, kinetics, and function of Cas13b, provides a mechanistic model for Cas13b target RNA recognition and identifies features responsible for target and cleavage specificity. Based on these observations, we generated Cas13b variants with altered cleavage preferences, which may expand the utility of nuclease-based RNA detection assays and other applications of Cas13b in mammalian cells.

Original languageEnglish
JournalCell Reports
Issue number13
Pages (from-to)3741-3751.e5
Number of pages17
Publication statusPublished - 2019

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