Structural Biology of RNA

描述(申请人提供)：我们希望了解生物RNA的三维(3D)结构和它们的作用机制之间的关系。我们使用X射线结晶学和生物化学相结合的方法来研究这两者，即自主折叠和发挥功能的RNA，以及RNA和蛋白质的细胞复合体。我们的模型系统是核糖开关和假尿苷合成。核糖开关是控制古生菌、细菌和真核生物中基因表达的RNA。核糖开关以高亲和力和高特异性结合小分子代谢物，并在结合时改变构象。尽管核糖开关是“裸露的”RNA，但它们表现出复杂的生化行为。它们中的一些通过表现出协作性的多个位置绑定它们的效应器。其他的是催化的RNA，由它们的配体变构激活。假尿苷合成酶是一种蛋白质酶，负责细胞RNA转录后修饰的最丰富类型。我们特别感兴趣的是由伪尿苷合成酶Dyskerin、3个辅助蛋白和Box H/ACA RNAs形成的RNA-蛋白质复合体。这种核糖核蛋白(RNP)复合体在古生物和人类之间保守，对于核糖体RNA的加工和成熟是必不可少的。在脊椎动物中，盒H/ACA RNP也是端粒酶RNP的一部分。端粒酶RNA的H/ACA结构域与Dyskerin的正确结合对于端粒酶的组装和稳定性都是必不可少的。我们将使用X射线结晶学来可视化处于多种功能状态的核糖开关和盒H/ACA RNPs。我们将结合结构信息、生化探测和溶液酶学来推断这些分子机器是如何工作的。简单描述：蛋白质和RNA，这些使生命成为可能的分子机器，有着复杂的结构。为了了解细胞RNA机器的工作原理，我们将在它们执行功能的不同时间捕获它们，并使用一种名为X射线结晶学的技术来找出它们的组成原子在不同的时间点上的位置。这些“分子电影”将提供基本的理解，将有助于操纵RNA在人类健康和疾病中的功能，以及在病原体的生命周期中的功能。

DESCRIPTION (provided by applicant): We wish to understand the relationships between the three-dimensional (3D)architectures of biological RNAs and their mechanisms of action. We use a combination of X-ray crystallography and biochemistry to study both, RNAs that fold and function autonomously, and cellular complexes of RNAs and proteins. Our model systems are riboswitches and pseudouridine syntheses. Riboswitches are RNAs that control gene expression in archaea, bacteria and eukaryotes. Riboswitches bind small-molecule metabolites with high affinity and specificity and switch conformation upon binding. Despite being 'naked' RNAs, riboswitches display sophisticated biochemical behaviors. Some of them bind their effectors through multiple sites that exhibit cooperativity. Others are catalytic RNAs that are allosterically activated by their ligands. Pseudouridine synthases are protein enzymes responsible for the most abundant type of post-transcriptional modification of cellular RNAs. We are particularly interested in an RNA-protein complex formed by the pseudouridine synthase Dyskerin, 3 accessory proteins, and the box H/ACA RNAs. This ribonucleoprotein (RNP) complex is conserved between archaea and human, and is essential for processing and maturation of ribosomal RNA. In vertebrates, a box H/ACA RNP is also part of the telomerase RNP. Correct association of the H/ACA domain of telomerase RNA with Dyskerin is essential both for telomerase assembly and stability. We will use X-ray crystallography to visualize riboswitches and box H/ACA RNPs in multiple functional states. We will combine structural information with biochemical probing and solution enzymology to deduce how these molecular machines work. Lay Description: Proteins and RNAs, the molecular machines that make life possible, have complicated architectures. In order to understand how cellular RNA machines work, we will trap them at different times while they carry out their function, and use a technique called X-ray crystallography to find out where their constituent atoms are at different points in time. These 'molecular movies' will provide the basic understanding that will help manipulate the functions of RNAs in human health and disease, and in the life cycle of pathogenic organisms.