High-Resolution Structure of Cas13b and Biochemical Characterization of RNA Targeting and Cleavage
Name
1-s2.0-S2211124719302803-main.pdf
Description
Published version
Size
4.46 MB
Format
Adobe PDF
Checksum (MD5)
212da44c1ba9a42dc00a1ea0fdc6ac3a
Author(s)
Slaymaker, Ian
Mesa, Pablo
Kellner, Max J.
Kannan, Soumya
Brignole, Edward J
Koob, Jeremy
Feliciano, Patricia Rosa
Stella, Stefano
Abudayyeh, Omar O.
Gootenberg, Jonathan S
Date Issued
March 2019
Journal
Cell Reports
Publisher
Elsevier BV
Citation
Slaymaker, Ian M. et al. "High-Resolution Structure of Cas13b and Biochemical Characterization of RNA Targeting and Cleavage." Cell Reports 26, 16 (March 2019): P3741-3751.e5 © 2019 The Author(s)
Version
Final published version
Abstract
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. Slaymaker et al. present a high-resolution structure of Cas13b in combination with biochemical studies showing that Cas13b is a highly dynamic, multi-turnover enzyme distinct from other Cas13 family members. Additionally, the authors demonstrate Cas13b can be rationally engineered to change RNA cleavage specificity and create a minimal variant suitable for delivery with viral vectors.
MIT Department
McGovern Institute for Brain Research at MIT
Massachusetts Institute of Technology. Department of Brain and Cognitive Sciences
Massachusetts Institute of Technology. Department of Biological Engineering
Massachusetts Institute of Technology. Department of Biology
Harvard University--MIT Division of Health Sciences and Technology
Broad Institute of MIT and Harvard
Terms of Use
Creative Commons Attribution 4.0 International license
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DOI of Published Version
https://doi.org/10.1016/j.celrep.2019.02.094
