Type V CRISPR-Cas Cpf1 endonuclease employs a unique mechanism for crRNA-mediated target DNA recognition
Type V CRISPR-Cas Cpf1 endonuclease employs a unique mechanism for crRNA-mediated target DNA recognition
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DOI：
10.1038/cr.2016.88
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发表时间：
2016-08-01
期刊：
影响因子：
44.1
通讯作者：
Patel, Dinshaw J.
中科院分区：
文献类型：
--
作者：
Gao, Pu;Yang, Hui;Patel, Dinshaw J.
CRISPR-Cas9和CRISPR-Cpf 1系统已成功用于基因组编辑。在CRIS-PR-Cas9系统中，预先排序的A型RNA种子序列和预先形成的蛋白质PAM相互作用裂缝对于Cas9形成DNA转录活性结构是必需的。CRISPR-Cpf 1系统是否采用类似的靶DNA识别机制尚不清楚。在这里，我们已经确定了与crRNA和靶DNA复合的Acidaminococcus sp. Cpf 1（AsCpf 1）的晶体结构。AsCpf 1-crRNA-DNA三元复合物与最近报道的毛螺菌科细菌Cpf 1（LbCpf 1）-crRNA二元复合物之间的结构比较鉴定了Cpf 1用于靶识别的独特机制。初始DNA询问所需的种子序列在Cpf 1-cRNA二元复合物中是无序的，但在三元复合物形成后变得有序。此外，Cpf 1的PAM相互作用裂缝在靶DNA结合后经历“开放到闭合”的构象变化，这反过来诱导Cpf 1内的结构变化以适应有序的A型种子RNA片段。Cpf 1的这种独特的靶向识别机制与先前报道的Cas9不同。
CRISPR-Cas9 and CRISPR-Cpf1 systems have been successfully harnessed for genome editing. In the CRIS-PR-Cas9 system, the preordered A-form RNA seed sequence and preformed protein PAM-interacting cleft are essential for Cas9 to form a DNA recognition-competent structure. Whether the CRISPR-Cpf1 system employs a similar mechanism for target DNA recognition remains unclear. Here, we have determined the crystal structure of Acidaminococcus sp. Cpf1 (AsCpf1) in complex with crRNA and target DNA. Structural comparison between the AsCpf1-crRNA-DNA ternary complex and the recently reported Lachnospiraceae bacterium Cpf1 (LbCpf1)-crRNA binary complex identifies a unique mechanism employed by Cpf1 for target recognition. The seed sequence required for initial DNA interrogation is disordered in the Cpf1-cRNA binary complex, but becomes ordered upon ternary complex formation. Further, the PAM interacting cleft of Cpf1 undergoes an "open-to-closed" conformational change upon target DNA binding, which in turn induces structural changes within Cpf1 to accommodate the ordered A-form seed RNA segment. This unique mechanism of target recognition by Cpf1 is distinct from that reported previously for Cas9.