A Cas9-guide RNA complex preorganized for target DNA recognition
A Cas9-guide RNA complex preorganized for target DNA recognition
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DOI：
10.1126/science.aab1452
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发表时间：
2015-06-26
期刊：
影响因子：
56.9
通讯作者：
Doudna, Jennifer A.
中科院分区：
文献类型：
--
作者：
Jiang, Fuguo;Zhou, Kaihong;Doudna, Jennifer A.
细菌适应性免疫使用CRISPR（簇簇插入了散布的短篇小语重复序列）与蛋白质相关（CAS）蛋白以及CRISPR转录本以用于外国DNA降解。在II型CRISPR-CAS系统中，通过未知机制发生CAS9核酸内切酶以引导RNA结合的DNA识别。 Cas9的晶体结构与单个指南RNA结合的晶体结构揭示了与APO和DNA结合状态不同的构象，其中初始DNA询问所需的10-核苷酸RNA“种子”序列以A形式构象预订。该导向RNA的这一段对于CAS9形成DNA识别能力的结构至关重要，该结构有望参与双链DNA靶序列。我们将其解释为“种子”机制的收敛演化，让人联想到真核生物RNA干扰期间Argonaute蛋白所使用的机制。
Bacterial adaptive immunity uses CRISPR (clustered regularly interspaced short palindromic repeats)-associated (Cas) proteins together with CRISPR transcripts for foreign DNA degradation. In type II CRISPR-Cas systems, activation of Cas9 endonuclease for DNA recognition upon guide RNA binding occurs by an unknown mechanism. Crystal structures of Cas9 bound to single-guide RNA reveal a conformation distinct from both the apo and DNA-bound states, in which the 10-nucleotide RNA "seed" sequence required for initial DNA interrogation is preordered in an A-form conformation. This segment of the guide RNA is essential for Cas9 to form a DNA recognition-competent structure that is poised to engage double-stranded DNA target sequences. We construe this as convergent evolution of a "seed" mechanism reminiscent of that used by Argonaute proteins during RNA interference in eukaryotes.