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A trailing ribosome speeds up RNA polymerase at the expense of transcript fidelity via force and allostery
Cell ( IF 64.5 ) Pub Date : 2023-03-16 , DOI: 10.1016/j.cell.2023.02.008
Liang Meng Wee 1 , Alexander B Tong 2 , Alfredo Jose Florez Ariza 3 , Cristhian Cañari-Chumpitaz 4 , Patricia Grob 5 , Eva Nogales 6 , Carlos J Bustamante 7
Affiliation  

In prokaryotes, translation can occur on mRNA that is being transcribed in a process called coupling. How the ribosome affects the RNA polymerase (RNAP) during coupling is not well understood. Here, we reconstituted the E. coli coupling system and demonstrated that the ribosome can prevent pausing and termination of RNAP and double the overall transcription rate at the expense of fidelity. Moreover, we monitored single RNAPs coupled to ribosomes and show that coupling increases the pause-free velocity of the polymerase and that a mechanical assisting force is sufficient to explain the majority of the effects of coupling. Also, by cryo-EM, we observed that RNAPs with a terminal mismatch adopt a backtracked conformation, while a coupled ribosome allosterically induces these polymerases toward a catalytically active anti-swiveled state. Finally, we demonstrate that prolonged RNAP pausing is detrimental to cell viability, which could be prevented by polymerase reactivation through a coupled ribosome.



中文翻译:

尾部核糖体通过力和变构以牺牲转录保真度为代价加速 RNA 聚合酶

在原核生物中,翻译可以发生在正在转录的 mRNA 上,这个过程称为偶联。核糖体在偶联过程中如何影响 RNA 聚合酶 (RNAP) 尚不清楚。在这里,我们重组了大肠杆菌偶联系统并证明核糖体可以防止 RNAP 的暂停和终止,并以牺牲保真度为代价使总转录率加倍。此外,我们监测了与核糖体偶联的单个 RNAP,并表明偶联增加了聚合酶的无暂停速度,并且机械辅助力足以解释偶联的大部分影响。此外,通过冷冻电镜,我们观察到具有末端错配的 RNAP 采用回溯构象,而偶联的核糖体变构诱导这些聚合酶进入具有催化活性的反旋转状态。最后,我们证明延长的 RNAP 暂停不利于细胞活力,这可以通过偶联核糖体聚合酶再激活来防止。

更新日期:2023-03-16
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