Local kinetic measures of macromolecular structure reveal partitioning among multiple parallel pathways from the earliest steps in the folding of a large RNA molecule
Local kinetic measures of macromolecular structure reveal partitioning among multiple parallel pathways from the earliest steps in the folding of a large RNA molecule
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DOI：
10.1016/j.jmb.2006.02.075
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发表时间：
2006-05-12
影响因子：
5.6
通讯作者：
Altman, Russ B.
中科院分区：
文献类型：
--
作者：
Laederach, Alain;Shcherbakova, Inna;Altman, Russ B.
RNA 折叠问题的核心是大多数大型 RNA 分子折叠途径中中间体的数量、结构和关系。对这些中间体结构动力学的独特见解可以从大分子构象局部探针随时间变化的变化中获得（例如，通过羟基自由基（(OH)-O-中心点）足迹提供的关于单个核苷酸溶剂可及性的报告）。分布在大分子周围的局部测量单独阐明了构成折叠反应的单独变化的整体。由于随着独立局部测量数量的增加，可能的动力学模型的组合爆炸，从大量这些单独测量中重建折叠通路是令人畏惧的。幸运的是，时间进度曲线的聚类充分降低了数据的维数，从而使重建在计算上易于处理。然后可以通过在超级计算机网格上详尽地列举所有可能的动力学模型来识别最可能的折叠拓扑和中间体。使用这种实验和计算相结合的方法，确定了嗜热四膜虫 I 组内含子的 Mg2+ 和 Na+ 介导的折叠途径和通过它们的相对通量的测量。 Mg2+介导的折叠过程中的通量被分配到许多平行路径中。相比之下，Na + 介导的反应期间的通量主要通过三种途径受到限制，其中之一是没有可检测到的通过中间体的通道。在这两种条件下，折叠反应都是高度平行的，没有单一途径占分子通量的 50% 以上。这表明，即使在折叠的最初阶段，RNA 折叠在各种不同的实验条件下也是非顺序的。这项研究提供了一个模板，用于系统分析来自局部测量集合的 RNA 结构的时间演化，这将阐明该过程中每个步骤的化学和物理特征。讨论了这种分析方法对其他大分子的适用性。 (c) 2006 Elsevier Ltd. 保留所有权利。
At the heart of the RNA folding problem is the number, structures, and relationships among the intermediates that populate the folding pathways of most large RNA molecules. Unique insight into the structural dynamics of these intermediates can be gleaned from the time-dependent changes in local probes of macromolecular conformation (e.g. reports on individual nucleotide solvent accessibility offered by hydroxyl radical ((OH)-O-center dot) footprinting). Local measures distributed around a macromolecule individually illuminate the ensemble of separate changes that constitute a folding reaction. Folding pathway reconstruction from a multitude of these individual measures is daunting due to the combinatorial explosion of possible kinetic models as the number of independent local measures increases. Fortunately, clustering of time progress curves sufficiently reduces the dimensionality of the data so as to make reconstruction computationally tractable. The most likely folding topology and intermediates can then be identified by exhaustively enumerating all possible kinetic models on a super-computer grid. The folding pathways and measures of the relative flux through them were determined for Mg2+ and Na+-mediated folding of the Tetrahymena thermophila group I intron using this combined experimental and computational approach. The flux during Mg2+-mediated folding is divided among numerous parallel pathways. In contrast, the flux during the Na+-mediated reaction is predominantly restricted through three pathways, one of which is without detectable passage through intermediates. Under both conditions, the folding reaction is highly parallel with no single pathway accounting for more than 50% of the molecular flux. This suggests that RNA folding is non-sequential under a variety of different experimental conditions even at the earliest stages of folding. This study provides a template for the systematic analysis of the time-evolution of RNA structure from ensembles of local measures that will illuminate the chemical and physical characteristics of each step in the process. The applicability of this analysis approach to other macromolecules is discussed. (c) 2006 Elsevier Ltd. All rights reserved.