Dynamics changes of CRISPR-Cas9 system induced by high fidelity mutations

报告人：慕宇光 教授

南洋理工大学，生物科学学院

邀请人：李伟峰 教授

报告时间：12月20日 上午9：30  

报告地点: 离子束实验室二楼报告厅

报告人简介：

1997年在买球最好的网站取得博士学位。1999年获洪堡奖学金赴德国留学，2003年获新加坡李光耀基金赴南洋理工大学生物科学学院工作至今。慕教授的研究领域为生物物理，主要从事蛋白质、DNA/RNA折叠的分子动力学研究以及动力学模拟方法和数据分析方法的发展， 是这些领域的世界知名学者。其研究成果多次发表在Nature Materials、Nature Communications、JACS等国际著名的刊物上。

报告摘要：

CRISPR-Cas9 as a powerful genome editing tool has widely been applied in biological fields. Ever since the discovery of CRISPR-Cas9 as an adaptive immune system, it has been gradually modified to perform precise genome editing in eukaryotic cells by creating double-strand breaks. Though being robust and efficient, current CRISPR-Cas9 system faces a major flaw: the off-target effect, which has not been well understood. Several Cas9 mutants show significant improvement, with very low off-target effect, however relatively lower cleavage efficiency for on-target sequences as well. In this study, the dynamics of the wild-type Cas9 from Streptococcus pyogenes and the high fidelity Cas9 mutant has been explored using molecular dynamics simulations. It turns out that the mutations cause reduction of electrostatic interactions between Cas9 and R-loop. Consequently, the flexibility of the tDNA/sgRNA heteroduplex is reduced, which may explain the reason of less tolerance of mismatches in the heteroduplex region. The mutations also affect the protein dynamics and the correlation networks among Cas9 domains. In mutant Cas9, weakened communications between two catalytic domains, as well as the mutations induced slight opening of the conformation, account for the lower on-target cleavage efficiency, and probably lower off-target as well. These findings would facilitate more precise Cas9 engineering in future.