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Characterization of the mechanical unfolding of RNA pseudoknots

机译：RNA假结的机械展开的表征

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The pseudoknot is an important RNA structural element that provides an excellent model system for studying the contributions of tertiary interactions to RNA stability and to folding kinetics. RNA pseudoknots are also of interest because of their key role in the control of ribosomal frameshifting by viral RNAs. Their mechanical properties are directly relevant to their unfolding by ribosomes during translation. We have used optical tweezers to study the kinetics and thermodynamics of mechanical unfolding and refolding of single RNA molecules. Here we describe the unfolding of the frameshifting pseudoknot from infectious bronchitis virus (IBV), three constituent hairpins, and three mutants of the IBV pseudoknot. All four pseudoknots cause -1 programmed ribosomal frameshifting. We have measured the free energies and rates of mechanical unfolding and refolding of the four frameshifting pseudoknots. Our results show that the IBV pseudoknot requires a higher force than its corresponding hairpins to unfold. Furthermore, its rate of unfolding changes little with increasing force, in contrast with the rate of hairpin unfolding. The presence of Mg2+ significantly increases the kinetic barriers to unfolding the IBV pseudoknot, but has only a minor effect on the hairpin unfolding. The greater mechanical stability of pseudoknots compared to hairpins, and their kinetic insensitivity to force supports the hypothesis that -1 frameshifting depends on the difficulty of unfolding the mRNA. (C) 2007 Elsevier Ltd. All rights reserved.
机译：假结是重要的RNA结构元件，为研究三次相互作用对RNA稳定性和折叠动力学的贡献提供了一个出色的模型系统。 RNA假结也很受关注，因为它们在通过病毒RNA控制核糖体移码中起关键作用。它们的机械性质与核糖体在翻译过程中的展开直接相关。我们已经使用光镊来研究单个RNA分子的机械展开和重折叠的动力学和热力学。在这里，我们描述了传染性支气管炎病毒（IBV），三个组成发夹和IBV假结的三个突变体的移码假结的展开。所有四个伪结均导致-1编程的核糖体移码。我们已经测量了四个移码假结的自由能和机械展开和重新折叠的速率。我们的结果表明，IBV假结展开所需的力要大于其相应的发夹。此外，与发夹的展开速度相反，其展开的速度随力的增加变化很小。 Mg2 +的存在显着增加了解开IBV假结的动力学障碍，但对发夹的解开作用很小。与发夹相比，假结的机械稳定性更高，并且它们对力的动力学不敏感，支持-1移码取决于展开mRNA的难度的假说。 （C）2007 Elsevier Ltd.保留所有权利。

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