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De Dong¹, Kuan Ren¹, Xiaolin Qiu¹

¹School of Life Science and Technology, Harbin Institute of Technology, Harbin 150080, China.

|April 21, 2016

概括

CRISPR-Cpf1系统是一个新的基因组编辑工具, 了解它的RNA识别机制有助于设计更精确的基因编辑技术.

科学领域:

分子生物学
遗传学
生物化学

背景情况:

克里斯普尔-卡斯系统是 prokaryotes 的适应性免疫机制.
在人体细胞中,CRISPR-Cpf1 (第二类) 提供了与CRISPR-Cas9不同的DNA干扰.
在指导RNA和基质特异性方面,Cpf1和Cas9有所不同.

研究的目的:

确定与CRISPRRNA (crRNA) 结合的Lachnospiraceae细菌ND2006 Cpf1 (LbCpf1) 的晶体结构.
阐明通过LbCpf1识别crRNA的机制.
为基因组编辑应用提供crRNA引导的基底结合的见解.

主要方法:

结合crRNA的LbCpf1的X射线晶体 (2.38 Å分辨率)
对LbCpf1架构和crRNA结合的结构分析.
研究crRNA结合时的形状变化.

主要成果:

LbCpf1呈现出一个三角形结构,其中有一个正电道.
通过相互作用和离子稳定,crRNA在寡核结域内采用扭曲的构造.
在没有寡合化的情况下,crRNA结合会诱导LbCpf1的结构变化.

结论:

这项研究揭示了由LbCpf1识别crRNA的结构基础.
对crRNA引导的基质结合的洞察为LbCpf1工程提供了基础.
这项工作促进了基于LbCpf1的改进基因组编辑工具的开发,提高了效率和特异性.

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