RNA Cold Denaturation

描述(由申请人提供)：RNA在生物化学的许多方面发挥着核心作用。为了执行这些细胞功能，RNA保持其复杂的二级、三级和四级结构至关重要。此外，越来越明显的是，作为正常生物功能的一部分，RNA很容易在两个或更多稳定(或伪稳定)结构之间进行相互转换。这些重排可能参与核酶活性的控制，如对接的情况，或通过在5‘-或3’-UTRs中形成替代结构而对mRNA的翻译控制。这个项目围绕着理解在某些RNA中观察到的结构变化的热力学基础。我们关注的一种结构变化是冷变性过程，在这个过程中，RNA在低温下展开。对这些过程特别重要的是折叠时热容的变化(增量C-p)，这一特性以前被认为是微不足道的，但现在在某些情况下被证明是显著的。锤头状核酶的冷变性是迄今为止研究得最好的，也是第一个特定目标的焦点。然后，将冷变性的热力学与高温和热变剂引起的变性进行比较。作为特定目标2的一部分，将探讨冷变性的一般性，以及冷变性与冷休克反应中涉及的核转录调节过程之间的关系。来自大肠杆菌的一种名为DSRA的非编码小RNA参与了这一过程的调节。我们正在探索这种RNA的结构变化，以及这些结构变化如何影响它与蛋白质(Hfq)和其他RNA形成的复合体。这一建议的第三个主要目标是研究核酸折叠中DeltaH和DeltaC-p的潜在性质，并了解金属水化/脱水反应对RNA三级结构形成的整体能量学的热力学贡献。

DESCRIPTION (provided by applicant): RNA plays a central role in many aspects of biochemistry. To carry out these cellular functions, it is critical that RNAs maintain their complex secondary, tertiary and quaternary structures. Furthermore, it is becoming evident that RNAs readily interconvert between two or more stable (or pseudo-stable) structures as part of their normal biological function. These rearrangements can be involved in control of ribozyme activity, as in the case of docking, or translational control of an mRNA through formation of alternative structures in the 5'- or 3'-UTRs. This project revolves around understanding the thermodynamic basis for the structural changes observed in certain RNAs. One type of structural change we focus on is the process of cold denaturation, where RNAs unfold at low temperature. Of particular importance to these processes, is the change in heat capacity (deltaC-p) upon folding, a property previously believed to be negligible, but now shown to be significant in certain cases. The cold denaturation of the hammerhead ribozyme is the best studied to date and the focus of the first specific aim. The thermodynamics to cold denaturation will then be compared to denaturation induced by high temperature and chaotropic agents. The generality of cold denaturation will be probed as part of specific aim 2, as well as the relationship between cold denaturation and riboregulation processes involved in the cold shock response. A small non-coding RNA from E. coil called DsrA is involved in regulating this process. We are probing the structural changes of this RNA and how these structural changes affect the complexes it makes with proteins (Hfq) and other RNAs. The 3rd major goal of this proposal is to look at the underlying nature of deltaH and deltaC-p in nucleic acid folding and understand the thermodynamic contribution of metal hydration/dehydration reactions on the overall energetics of tertiary structure formation of RNAs.