研究人员重组了大肠杆菌的耦合系统，并证明核糖体可以防止RNA聚合酶（RNAP）的暂停和终止，并在牺牲保真度的情况下使总转录率增加一倍。此外，研究人员监测了与核糖体耦合的单个RNAP，显示耦合增加了聚合酶的无暂停速度，并且机械辅助力足以解释耦合的大部分效果。另外，通过冷冻电镜，研究人员观察到具有末端错配的RNAP采用了回溯的构象，而耦合的核糖体则异构诱导这些聚合酶走向催化活性的反旋转状态。最后，研究人员证明，长时间的RNAP停顿对细胞的生存能力是有害的，这可以通过耦合的核糖体重新激活聚合酶来防止。
据了解，在原核生物中，翻译可以发生在正在转录的mRNA上，这个过程称为耦合。在耦合过程中，核糖体如何影响RNAP还不是很清楚。
附：英文原文
Title: A trailing ribosome speeds up RNA polymerase at the expense of transcript fidelity via force and allostery
Author: Liang Meng Wee, Alexander B. Tong, Alfredo Jose Florez Ariza, Cristhian Caari-Chumpitaz, Patricia Grob, Eva Nogales, Carlos J. Bustamante
Issue&Volume: 2023/03/16
Abstract: 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.
DOI: 10.1016/j.cell.2023.02.008
Source: https://www.cell.com/cell/fulltext/S0092-8674(23)00112-5