COUNTERION DEPENDENT FOLDING OF RNA

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。对RNA组装成三维结构的基本原理的理解仍然缺乏。解决这一问题将提高人们对聚电解质的认识，也是设计和操作RNA的必要条件，而RNA是一种很有可能应用于制药工业和生物技术的物质。我们用小角X射线散射(SAXS)技术观察了固氮弧菌核酶的大小与镁离子和温度的关系。我们的分析表明，双态跃迁可以用来模拟折叠过程。这种转变强烈依赖于温度，在12-38℃范围内，随着温度的降低，这种转变变得不那么合作。我们通过突变破坏了固氮藻中的一个三级相互作用，研究了三级相互作用对崩塌转变的影响。我们的测量表明，三级相互作用在塌陷态的稳定中起着重要的作用。在突变样本中，崩塌转变的中点显著增加，转变变得不那么合作。塌陷状态不如野生型核酶致密。结果表明，在塌陷态中确实形成了三级相互作用，它们对稳定塌陷态起到了积极作用。在我们未来的实验中，我们将观察额外突变的RNA分子的折叠行为，以衡量不同结构域之间在崩溃转变中相互作用的相关性。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. An understanding of basic principles governing the RNA assembly in to three dimensional structures is still missing. Solving this problem will improve the knowledge on polyelectrolytes, and is a requisite for the design and manipulation of RNA, which is a strong candidate to be used in pharmaceutical industry and in biotechnology. We used the Small Angle X-Ray Scattering (SAXS) technique to observe the size of the Azoarcus Ribozyme as a function of Mg2+ and temperature. Our analysis has indicated that a two-state transition can be used to model the folding process. This transition was strongly temperature dependent and became less cooperative with decreasing temperature over the range of 12-38C. We studied the effect of tertiary interactions on the collapse transition by disrupting one of the tertiary interactions in Azoarcus through a mutation. Our measurements pointed that the tertiary interactions play an important role in the stabilization of the collapsed state. In the mutated sample, the midpoint of the collapse transition increased significantly and the transition became less cooperative. The collapsed state was not as compact as the wild type ribozyme. The results show that tertiary interactions do form in the collapsed state, and they play an active role in stabilizing the collapsed state. In our future experiments, we will observe the folding behavior of additional mutated RNA molecules, in order to measure the relevance of interactions between different domains in the collapse transition.