College of Chemistry and Chemical Engineering, Shanghai University of Engineering Science, Shanghai 201620, China.
Department of Pharmaceutical Sciences, University of North Texas System College of Pharmacy, University of North Texas Health Science Center, Fort Worth, TX 76107, USA.
Curr Opin Struct Biol. 2020 Jun;62:166-174. doi: 10.1016/j.sbi.2020.01.013. Epub 2020 Feb 18.
The CRISPR-Cas9 system from Streptococcus pyogenes has been exploited as a programmable RNA-guided DNA-targeting and DNA-editing platform. This evolutionary tool enables diverse genetic manipulations with unprecedented precision and ease. Cas9 is an allosteric enzyme, which is allosterically regulated in conformational activation, target recognition, and DNA cleavage. Here, we outline the underlying allosteric control over the Cas9 complex assembly and targeting specificity. We further review the strategies for mitigating intrinsic Cas9 off-target effects through allosteric modulations and the advances in engineering controllable Cas9 systems that are responsive to external allosteric signals. Future development of highly specific, tunable CRISPR-Cas9 systems through allosteric modulations would greatly benefit applications that require both conditional control and high precision.
CRISPR-Cas9 系统源自酿脓链球菌，现已被开发为一种可编程的 RNA 引导的 DNA 靶向和 DNA 编辑平台。这一进化工具使各种基因操作具有前所未有的精确性和简易性。Cas9 是一种变构酶，在构象激活、靶标识别和 DNA 切割过程中受到变构调节。在这里，我们概述了 Cas9 复合物组装和靶向特异性的变构控制。我们进一步综述了通过变构调节减轻 Cas9 固有脱靶效应的策略，以及构建对外源变构信号有响应的可控 Cas9 系统的进展。通过变构调节开发高度特异性、可调谐的 CRISPR-Cas9 系统将极大地有益于需要条件控制和高精度的应用。
