A Two-State Model for the Dynamics of the Pyrophosphate Ion Release in Bacterial RNA Polymerase

The dynamics of the PPi release during the transcription elongation of bacterial RNA polymerase and its effects on the Trigger Loop (TL) opening motion are still elusive. Here, we built a Markov State Model (MSM) from extensive all-atom molecular dynamics (MD) simulations to investigate the mechanism of the PPi release. Our MSM has identified a simple two-state mechanism for the PPi release instead of a more complex four-state mechanism observed in RNA polymerase II (Pol II). We observed that the PPi release in bacterial RNA polymerase occurs at sub-microsecond timescale, which is ∼3-fold faster than that in Pol II. After escaping from the active site, the (Mg-PPi)2− group passes through a single elongated metastable region where several positively charged residues on the secondary channel provide favorable interactions. Surprisingly, we found that the PPi release is not coupled with the TL unfolding but correlates tightly with the side-chain rotation of the TL residue R1239. Our work sheds light on the dynamics underlying the transcription elongation of the bacterial RNA polymerase.

细菌RNA聚合酶转录延伸过程中焦磷酸(PPi)的释放动力学及其对触发环(Trigger Loop，TL)开放运动的影响仍不明晰。本研究基于大规模全原子分子动力学(molecular dynamics，MD)模拟构建了马尔可夫状态模型(Markov State Model，MSM)，以探究焦磷酸释放的分子机制。我们所构建的MSM揭示了焦磷酸释放的简单双态机制，而非此前在RNA聚合酶II(RNA polymerase II，Pol II)中观测到的更为复杂的四态机制。研究发现，细菌RNA聚合酶中的焦磷酸释放发生在亚微秒时间尺度，其速率约为RNA聚合酶II的3倍。当焦磷酸从活性位点逃逸后，(Mg-PPi)²⁻基团会经历一段单一的伸长型亚稳态区域，该区域内二级通道上的多个带正电残基可提供有利的相互作用。令人意外的是，我们发现焦磷酸的释放并不与触发环的解折叠过程偶联，而是与TL残基R1239的侧链旋转紧密相关。本研究阐明了细菌RNA聚合酶转录延伸背后的内在动力学机制。