Journal
ELIFE
Volume 8, Issue -, Pages -Publisher
ELIFE SCIENCES PUBLICATIONS LTD
DOI: 10.7554/eLife.46500
Keywords
-
Categories
Funding
- Shanghai Municipal Education Commission
- National Science Foundation [MCB-1716423]
- National Institute of General Medical Sciences [P20GM103640]
- University of North Texas (UNT) Health Science Center
Ask authors/readers for more resources
The CRISPR-associated endonuclease Cas9 from Streptococcus pyogenes (SpyCas9), along with a programmable single-guide RNA (sgRNA), has been exploited as a significant genome-editing tool. Despite the recent advances in determining the SpyCas9 structures and DNA cleavage mechanism, the cleavage-competent conformation of the catalytic HNH nuclease domain of SpyCas9 remains largely elusive and debatable. By integrating computational and experimental approaches, we unveiled and validated the activated Cas9-sgRNA-DNA ternary complex in which the HNH domain is neatly poised for cleaving the target DNA strand. In this catalysis model, the HNH employs the catalytic triad of D839-H840-N863 for cleavage catalysis, rather than previously implicated D839-H840-D861, D837-D839-H840, or D839-H840-D861-N863. Our study contributes critical information to defining the catalytic conformation of the HNH domain and advances the knowledge about the conformational activation underlying Cas9-mediated DNA cleavage.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available