Distance determination by GIY-YIG intron endonucleases: discrimination between repression and cleavage functions.
Distance determination by GIY-YIG intron endonucleases: discrimination between repression and cleavage functions.
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DOI：
10.1093/nar/gkl079
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发表时间：
2006
影响因子：
14.9
通讯作者：
Edgell DR
中科院分区：
文献类型：
--
作者：
Liu Q;Derbyshire V;Belfort M;Edgell DR
GIY-YIG归巢内切酶是一种模块化的蛋白质，其N端的保守催化结构域与C端的DNA结合区相连。I-TEVI是T4噬菌体GIY-YIG内含子内切酶，它既能促进TD内含子归位，又能作为转录自抑制因子发挥作用。抑制是通过与操纵子结合来实现的，操纵子被切割的效率比无内含子的定位位点降低了100倍。连接体包括锌指，它在距离确定中起作用，以约束催化结构域以在固定位置切割定位位点。在这里，我们发现I-BmoI，一个与GIY-YIG相关的缺乏锌指的内切酶，也具有一定的切割距离分辨能力。此外，通过交换I-BmoI和I-TEVI结构域构建的杂交内切酶活性高、精确度高，并表明锌指以外的特征有助于距离确定。最重要的是，I-Tevi锌指突变体比定位位点更有效地切割操纵子，而野生型蛋白质则相反。这些结果与锌指作为测量装置的作用是一致的，它指导定位位点的有效切割以促进内含子的移动，同时减少操纵子的切割以确保转录自抑制和噬菌体活性。
GIY-YIG homing endonucleases are modular proteins, with conserved N-terminal catalytic domains connected by linkers to C-terminal DNA-binding domains. I-TevI, the T4 phage GIY-YIG intron endonuclease, functions both in promoting td intron homing, and in acting as a transcriptional autorepressor. Repression is achieved by binding to an operator, which is cleaved at 100-fold reduced efficiency relative to the intronless homing site. The linker includes a zinc finger, which functions in distance determination, to constrain the catalytic domain to cleave the homing site at a fixed position. Here we show that I-BmoI, a related GIY-YIG endonuclease lacking a zinc finger, also possesses some cleavage distance discrimination. Furthermore, hybrid endonucleases constructed by swapping the domains of I-BmoI and I-TevI are active, precise and demonstrate that features other than the zinc finger facilitate distance determination. Most importantly, I-TevI zinc finger mutants cleave the operator more efficiently than the homing site, the converse of wild-type protein. These results are consistent with the zinc finger acting as a measuring device, directing efficient cleavage of the homing site to promote intron mobility, while reducing cleavage at the operator to ensure transcriptional autorepression and phage viability.
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影响因子：
10.5
作者：
Fricke, WM;Brill, SJ
通讯作者：
Brill, SJ
影响因子：
1.5
作者：
Patthy, L
通讯作者：
Patthy, L
影响因子：
11.4
作者：
BRYK, M;QUIRK, SM;BELFORT, M
通讯作者：
BELFORT, M
影响因子：
5.6
作者：
Edgell, DR;Stanger, MJ;Belfort, M
通讯作者：
Belfort, M
影响因子：
16.8
作者：
Edgell, DR;Derbyshire, V;Belfort, M
通讯作者：
Belfort, M