maintenance of CRISPR repeat elements / endonuclease activity / defense response to virus / 加水分解酵素; エステル加水分解酵素 / DNA binding / RNA binding / metal ion binding 類似検索 - 分子機能
National Natural Science Foundation of China (NSFC)
21922705
中国
National Natural Science Foundation of China (NSFC)
22277078
中国
引用
ジャーナル: ACS Synth Biol / 年: 2024 タイトル: Molecular Basis and Genome Editing Applications of a Compact CRISPR-Cas9 System. 著者: Na Tang / Zhaowei Wu / Yan Gao / Weizhong Chen / Zixiao Wang / Mengjiao Su / Wenxin Ji / Quanjiang Ji / 要旨: CRISPR-Cas9 systems have been widely harnessed for diverse genome editing applications because of their ease of use and high efficiency. However, the large molecular sizes and strict PAM requirements ...CRISPR-Cas9 systems have been widely harnessed for diverse genome editing applications because of their ease of use and high efficiency. However, the large molecular sizes and strict PAM requirements of commonly used CRISPR-Cas9 systems restrict their broad applications in therapeutics. Here, we report the molecular basis and genome editing applications of a novel compact type II-A CRISPR-Cas9 system (EvCas9) with 1107 residues and distinct 5'-NNGDGN-3' (where D represents A, T, or G) PAM specificity. We determine the cryo-EM structure of EvCas9 in a complex with an sgRNA and a target DNA, revealing the detailed PAM recognition and sgRNA and target DNA association mechanisms. Additionally, we demonstrate the robust genome editing capacity of EvCas9 in bacteria and human cells with superior fidelity compared to SaCas9 and SpCas9, and we engineer it to be efficient base editors by fusing a cytidine or adenosine deaminase. Collectively, our results facilitate further understanding of CRISPR-Cas9 working mechanisms and expand the compact CRISPR-Cas9 toolbox.