Department of Chemistry, University of Texas, Austin, TX 78712, USA.
School of Pharmacy, University of Nottingham, Nottingham, UK.
Nucleic Acids Res. 2023 Oct 27;51(19):10737-10751. doi: 10.1093/nar/gkad755.
Folding of ribozymes into well-defined tertiary structures usually requires divalent cations. How Mg2+ ions direct the folding kinetics has been a long-standing unsolved problem because experiments cannot detect the positions and dynamics of ions. To address this problem, we used molecular simulations to dissect the folding kinetics of the Azoarcus ribozyme by monitoring the path each molecule takes to reach the folded state. We quantitatively establish that Mg2+ binding to specific sites, coupled with counter-ion release of monovalent cations, stimulate the formation of secondary and tertiary structures, leading to diverse pathways that include direct rapid folding and trapping in misfolded structures. In some molecules, key tertiary structural elements form when Mg2+ ions bind to specific RNA sites at the earliest stages of the folding, leading to specific collapse and rapid folding. In others, the formation of non-native base pairs, whose rearrangement is needed to reach the folded state, is the rate-limiting step. Escape from energetic traps, driven by thermal fluctuations, occurs readily. In contrast, the transition to the native state from long-lived topologically trapped native-like metastable states is extremely slow. Specific collapse and formation of energetically or topologically frustrated states occur early in the assembly process.
核酶折叠成明确的三级结构通常需要二价阳离子。Mg2+ 离子如何指导折叠动力学一直是一个长期未解决的问题，因为实验无法检测离子的位置和动态。为了解决这个问题，我们使用分子模拟来通过监测每个分子到达折叠状态所采取的路径来剖析 Azoarcus 核酶的折叠动力学。我们定量地确定了 Mg2+ 与特定位点的结合，加上单价阳离子的抗衡离子释放，刺激了二级和三级结构的形成，导致了不同的途径，包括直接快速折叠和在错误折叠结构中捕获。在一些分子中，当 Mg2+ 离子在折叠的最早阶段结合到特定的 RNA 位点时，形成关键的三级结构元素，导致特定的折叠和快速折叠。在其他分子中，需要重新排列非天然碱基对才能达到折叠状态，这是限速步骤。由热波动驱动的从能量陷阱中逃逸很容易发生。相比之下，从拓扑上困住的类似天然的亚稳态向天然状态的转变非常缓慢。特定的折叠和形成能量上或拓扑上受阻的状态在组装过程的早期发生。
