TY - JOUR
T1 - High-Resolution Structure of Cas13b and Biochemical Characterization of RNA Targeting and Cleavage
AU - Slaymaker, Ian M
AU - Mesa, Pablo
AU - Kellner, Max J
AU - Kannan, Soumya
AU - Brignole, Edward
AU - Koob, Jeremy
AU - Feliciano, Patricia R
AU - Stella, Stefano
AU - Abudayyeh, Omar O
AU - Gootenberg, Jonathan S
AU - Strecker, Jonathan
AU - Montoya, Guillermo
AU - Zhang, Feng
PY - 2019
Y1 - 2019
N2 - 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.
AB - 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.
U2 - 10.1016/j.celrep.2019.02.094
DO - 10.1016/j.celrep.2019.02.094
M3 - Journal article
C2 - 30917325
VL - 26
SP - 3741-3751.e5
JO - Cell Reports
JF - Cell Reports
SN - 2211-1247
IS - 13
ER -