Structures of CRISPR Cas3 offer mechanistic insights into Cascade-activated DNA unwinding and degradation.
Structures of CRISPR Cas3 offer mechanistic insights into Cascade-activated DNA unwinding and degradation.
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DOI：
10.1038/nsmb.2875
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发表时间：
2014-09
影响因子：
16.8
通讯作者：
Ke A
中科院分区：
文献类型：
--
作者：
Huo Y;Nam KH;Ding F;Lee H;Wu L;Xiao Y;Farchione MD Jr;Zhou S;Rajashankar K;Kurinov I;Zhang R;Ke A
CRISPR利用rna介导的干扰机制驱动原核生物适应侵入性核酸，如噬菌体和质粒。对I型CRISPR-Cas系统的干扰需要靶向级联复合体和降解机器Cas3， Cas3同时含有核酸酶和解旋酶活性。在这里，我们报道了结合到ss-DNA底物的Cas3的晶体结构，并表明它是一个义务的3 ‘到5 ’ ss-DNA酶，优先从解旋酶部分直接接受底物。hd型核酸酶中的保守残基协调两个铁进行ss-DNA切割。sf2型解旋酶片段显示ATP配位和构象柔韧性。Cas3通过与非靶底物链和CasA蛋白的物理相互作用，被特异性地引导到具有正确PAM序列的级联结合靶DNA。识别事件的级联确保了通过cascade和Cas3对外来DNA的靶向和降解进行良好控制。
CRISPR drives prokaryotic adaptation to invasive nucleic acids such as phages and plasmids using an RNA-mediated interference mechanism. Interference in Type I CRISPR-Cas systems requires a targeting Cascade complex and a degradation machine Cas3, which contains both nuclease and helicase activities. Here we report the crystal structures of Cas3 bound to ss-DNA substrate and show that it is an obligated 3′-to-5′ ss-DNase preferentially accepting substrate directly from the helicase moiety. Conserved residues in the HD-type nuclease coordinate two irons for ss-DNA cleavage. ATP coordination and conformational flexibility are revealed for the SF2-type helicase moiety. Cas3 is specifically guided towards Cascade-bound target DNA with a correct PAM sequence, through physical interactions to both the non-target substrate strand and the CasA protein. The cascade of recognition events ensures a well-controlled DNA targeting and degradation of alien DNA by Cascade and Cas3.
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DOI：
10.1126/science.1165771
发表时间：
2008-12-19
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Marraffini LA;Sontheimer EJ
通讯作者：
Sontheimer EJ
影响因子：
11.4
作者：
Beloglazova, Natalia;Petit, Pierre;Yakunin, Alexander F.
通讯作者：
Yakunin, Alexander F.
DOI：
10.1126/science.1159689
发表时间：
2008-08-15
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Brouns SJ;Jore MM;Lundgren M;Westra ER;Slijkhuis RJ;Snijders AP;Dickman MJ;Makarova KS;Koonin EV;van der Oost J
通讯作者：
van der Oost J
影响因子：
64.5
作者：
Lee, Jae Young;Yang, Wei
通讯作者：
Yang, Wei
影响因子：
5.5
作者：
Makarova, Kira S.;Grishin, Nick V.;Koonin, Eugene V.
通讯作者：
Koonin, Eugene V.