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

Ian M. Slaymaker | Zendy; Pablo Mesa | Zendy; Max J. Kellner | Zendy; Soumya Kannan | Zendy; Edward J. Brignole | Zendy; Jeremy Koob | Zendy; Patrícia R. Feliciano | Zendy; Stefano Stella | Zendy; Omar O. Abudayyeh | Zendy; Jonathan S. Gootenberg | Zendy; Jonathan Strecker | Zendy; Guillermo Montoya | Zendy; Feng Zhang | Zendy

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

Author(s) -

Ian M. Slaymaker,

Pablo Mesa,

Max J. Kellner,

Soumya Kannan,

Edward J. Brignole,

Jeremy Koob,

Patrícia R. Feliciano,

Stefano Stella,

Omar O. Abudayyeh,

Jonathan S. Gootenberg,

Jonathan Strecker,

Guillermo Montoya,

Feng Zhang

Publication year - 2019

Publication title -

cell reports

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 6.264

H-Index - 154

eISSN - 2639-1856

pISSN - 2211-1247

DOI - 10.1016/j.celrep.2019.02.094

Subject(s) - trans activating crrna , crispr , rna , nuclease , effector , nucleic acid structure , computational biology , biology , nucleic acid , guide rna , rna editing , cleavage (geology) , rna silencing , microbiology and biotechnology , dna , cas9 , genetics , rna interference , gene , paleontology , fracture (geology)

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.

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