Crystal Structure of Cpf1 in Complex with Guide RNA and Target DNA.
Crystal Structure of Cpf1 in Complex with Guide RNA and Target DNA.
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DOI：
10.1016/j.cell.2016.04.003
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发表时间：
2016-05-05
期刊：
影响因子：
64.5
通讯作者：
Nureki O
中科院分区：
文献类型：
--
作者：
Yamano T;Nishimasu H;Zetsche B;Hirano H;Slaymaker IM;Li Y;Fedorova I;Nakane T;Makarova KS;Koonin EV;Ishitani R;Zhang F;Nureki O
Cpf1是一种rna引导的V型CRISPR-Cas系统内切酶，最近已被用于基因组编辑。在这里，我们报道了酸胺球菌sp. Cpf1 （AsCpf1）与引导RNA及其靶DNA复合物的晶体结构，分辨率为2.8 Å。AsCpf1采用双叶结构，RNA-DNA异双工结合在中心通道内。AsCpf1与Cas9（一种II型CRISPR-Cas核酸酶）的结构比较揭示了它们惊人的相似性和主要的差异，从而解释了它们不同的功能。AsCpf1含有RuvC结构域和一个假定的新型核酸酶结构域，它们分别负责非靶链和靶链的切割，并共同产生交错的DNA双链断裂。AsCpf1通过碱基和形状读出机制识别5 ‘ - ttn -3 ’原间隔邻近基序。我们的发现为Cpf1在rna引导下的DNA切割提供了机制见解，并为CRISPR-Cpf1工具箱的合理工程建立了框架。
Cpf1 is an RNA-guided endonuclease of a type V CRISPR-Cas system that has been recently harnessed for genome editing. Here, we report the crystal structure of Acidaminococcus sp. Cpf1 (AsCpf1) in complex with the guide RNA and its target DNA, at 2.8 Å resolution. AsCpf1 adopts a bilobed architecture, with the RNA–DNA heteroduplex bound inside the central channel. The structural comparison of AsCpf1 with Cas9, a type II CRISPR-Cas nuclease, reveals both striking similarity and major differences, thereby explaining their distinct functionalities. AsCpf1 contains the RuvC domain and a putative novel nuclease domain, which are responsible for the cleavage of the non-target and target strands, respectively, and jointly generate staggered DNA double-strand breaks. AsCpf1 recognizes the 5′-TTTN-3′ protospacer adjacent motif by base and shape readout mechanisms. Our findings provide mechanistic insights into RNA-guided DNA cleavage by Cpf1, and establish a framework for rational engineering of the CRISPR-Cpf1 toolbox.