Leipply Desirae, Lambert Dominic, Draper David E
Department of Biophysics, Johns Hopkins University, Baltimore, Maryland, USA.
Methods Enzymol. 2009;469:433-63. doi: 10.1016/S0076-6879(09)69021-2. Epub 2009 Nov 17.
RNA secondary and tertiary structures are strongly stabilized by added salts, and a quantitative thermodynamic analysis of the relevant ion-RNA interactions is an important aspect of the RNA folding problem. Because of long-range electrostatic forces, an RNA perturbs the distribution of both cations and anions throughout a large volume. Binding formalisms that require a distinction between "bound" and "free" ions become problematic in such situations. A more fundamental thermodynamic framework is developed here, based on preferential interaction coefficients; linkage equations derived from this framework provide a model-free description of the "uptake" or "release" of cations and anions that accompany an RNA conformational transition. Formulas appropriate for analyzing the dependence of RNA stability on either mono- or divalent salt concentration are presented and their application to experimental data is illustrated. Two example datasets are analyzed with respect to the monovalent salt dependence of tertiary structure formation in different RNAs, and three different experimental methods for quantitating the "uptake" of Mg(2+) ions are applied to the folding of a riboswitch RNA. Advantages and limitations of each method are discussed.
RNA的二级和三级结构会因添加的盐而得到强烈稳定，对相关离子与RNA相互作用进行定量热力学分析是RNA折叠问题的一个重要方面。由于长程静电力的存在，RNA会扰乱整个大体积区域内阳离子和阴离子的分布。在这种情况下，需要区分“结合”离子和“游离”离子的结合形式会出现问题。本文基于优先相互作用系数建立了一个更基本的热力学框架；从该框架导出的连锁方程提供了一个无模型的描述，用于描述伴随RNA构象转变的阳离子和阴离子的“摄取”或“释放”。给出了适用于分析RNA稳定性对单价或二价盐浓度依赖性的公式，并说明了它们在实验数据中的应用。针对不同RNA中三级结构形成对单价盐的依赖性，分析了两个示例数据集，并将三种不同的定量Mg(2+)离子“摄取”的实验方法应用于核糖开关RNA的折叠。讨论了每种方法的优缺点。
