Structural basis of transcription arrest by coliphage HK022 Nun in an Escherichia coli RNA polymerase elongation complex
Structural basis of transcription arrest by coliphage HK022 Nun in an Escherichia coli RNA polymerase elongation complex
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DOI：
10.7554/elife.25478
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发表时间：
2017-03-20
期刊：
影响因子：
7.7
通讯作者：
Darst, Seth A.
中科院分区：
文献类型：
--
作者：
Kang, Jin Young;Olinares, Paul Dominic B.;Darst, Seth A.
大肠杆菌噬菌体HK 022 Nun通过阻止RNA聚合酶（RNAP）特异性地在λ DNA上易位来阻断大肠杆菌噬菌体λ的双重感染。为了提供一个结构框架来理解Nun如何阻止RNAP易位，我们通过单粒子冷冻电子显微镜确定了有和没有Nun的大肠杆菌RNAP三元延伸复合物（TEC）的结构。Nun通过利用RNAP和核酸之间的间隙而紧密地融入TEC。Nun的C-末端区段与RNAP活性位点裂缝内的RNAP β和β '亚基以及与核酸支架的几乎每个元件相互作用，基本上交联RNAP和核酸以防止易位，这是由Nun氨基酸取代的作用支持的机制。RNAP活性位点裂缝内Nun相互作用的性质表明RNAP钳夹打开是Nun建立其相互作用所必需的，这解释了为什么Nun作用于暂停的TEC。
Coliphage HK022 Nun blocks superinfection by coliphage lambda, by stalling RNA polymerase (RNAP) translocation specifically on lambda, DNA. To provide a structural framework to understand how Nun blocks RNAP translocation, we determined structures of Escherichia coli RNAP ternary elongation complexes (TECs) with and without Nun by single-particle cryo-electron microscopy. Nun fits tightly into the TEC by taking advantage of gaps between the RNAP and the nucleic acids. The C-terminal segment of Nun interacts with the RNAP beta and beta' subunits inside the RNAP active site cleft as well as with nearly every element of the nucleic acid scaffold, essentially crosslinking the RNAP and the nucleic acids to prevent translocation, a mechanism supported by the effects of Nun amino acid substitutions. The nature of Nun interactions inside the RNAP active site cleft suggests that RNAP clamp opening is required for Nun to establish its interactions, explaining why Nun acts on paused TECs.