Abstract：
:High-resolution Cas9 structures have yet to reveal catalytic conformations due to HNH nuclease domain positioning away from the cleavage site. Nme1Cas9 and Nme2Cas9 are compact nucleases for in vivo genome editing. Here, we report structures of meningococcal Cas9 homologs in complex with sgRNA, dsDNA, or the AcrIIC3 anti-CRISPR protein. DNA-bound structures represent an early step of target recognition, a later HNH pre-catalytic state, the HNH catalytic state, and a cleaved-target-DNA-bound state. In the HNH catalytic state of Nme1Cas9, the active site is seen poised at the scissile phosphodiester linkage of the target strand, providing a high-resolution view of the active conformation. The HNH active conformation activates the RuvC domain. Our structures explain how Nme1Cas9 and Nme2Cas9 read distinct PAM sequences and how AcrIIC3 inhibits Nme1Cas9 activity. These structures provide insights into Cas9 domain rearrangements, guide-target engagement, cleavage mechanism, and anti-CRISPR inhibition, facilitating the optimization of these genome-editing platforms.
journal_name
Mol Celljournal_title
Molecular cellauthors
Sun W,Yang J,Cheng Z,Amrani N,Liu C,Wang K,Ibraheim R,Edraki A,Huang X,Wang M,Wang J,Liu L,Sheng G,Yang Y,Lou J,Sontheimer EJ,Wang Ydoi
10.1016/j.molcel.2019.09.025subject
Has Abstractpub_date
2019-12-19 00:00:00pages
938-952.e5issue
6eissn
1097-2765issn
1097-4164pii
S1097-2765(19)30730-0journal_volume
76pub_type
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