Structure-based cleavage mechanism of Thermus thermophilus Argonaute DNA guide strand-mediated DNA target cleavage
Structure-based cleavage mechanism of Thermus thermophilus Argonaute DNA guide strand-mediated DNA target cleavage
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嗜热栖热菌 Argonaute DNA 引导链介导的 DNA 靶标切割的基于结构的切割机制
DOI：
10.1073/pnas.1321032111
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发表时间：
2014-01-14
影响因子：
11.1
通讯作者：
Wang, Yanli
中科院分区：
文献类型：
--
作者：
Sheng, Gang;Zhao, Hongtu;Wang, Yanli
我们报道了三元Thermus thermophilus Argonaute（TtAgo）复合物与5 '-磷酸化的指导DNA和一系列DNA靶标的晶体结构。这些三元复合物结构的裂解不兼容，裂解兼容，和postcleavage状态解决在提高分辨率高达2.2埃提供了分子的见解，协调定位的催化残基，一对Mg 2+阳离子，和假定的水亲核试剂定位为在线攻击的可裂解磷酸TtAgo介导的目标裂解的RNA酶H型机制。此外，这些三元复合物结构提供了对蛋白质和DNA构象变化的深入了解，这些构象变化促进了切割不相容状态和切割相容状态之间的转变，包括Glu指在产生具有切割能力的催化Asp-Glu-Asp-Asp四分体中的作用。切割后，种子片段与靶链的互补片段形成稳定的双链体。
We report on crystal structures of ternary Thermus thermophilus Argonaute (TtAgo) complexes with 5'-phosphorylated guide DNA and a series of DNA targets. These ternary complex structures of cleavage-incompatible, cleavage-compatible, and postcleavage states solved at improved resolution up to 2.2 angstrom have provided molecular insights into the orchestrated positioning of catalytic residues, a pair of Mg2+ cations, and the putative water nucleophile positioned for in-line attack on the cleavable phosphate for TtAgo-mediated target cleavage by a RNase H-type mechanism. In addition, these ternary complex structures have provided insights into protein and DNA conformational changes that facilitate transition between cleavage-incompatible and cleavage-compatible states, including the role of a Glu finger in generating a cleavage-competent catalytic Asp-Glu-Asp-Asp tetrad. Following cleavage, the seed segment forms a stable duplex with the complementary segment of the target strand.