Structural Biology of RNA

描述（由申请人提供）：我们希望了解生物RNA的三维（3D）结构与其作用机制之间的关系。我们使用X射线晶体学和生物化学的组合来研究这两种RNA，折叠和自主功能，以及RNA和蛋白质的细胞复合物。我们的模型系统是核糖开关和假尿苷合成。核糖开关是控制古细菌、细菌和真核生物中基因表达的RNA。核糖开关以高亲和力和特异性结合小分子代谢物，并在结合后转换构象。尽管是“裸”RNA，核糖开关显示复杂的生化行为。它们中的一些通过表现出协同性的多个位点结合其效应器。另一些是催化RNA，它们被配体变构激活。假尿苷激酶是负责细胞RNA的最丰富类型的转录后修饰的蛋白酶。我们特别感兴趣的是由假尿苷合酶Dyskerin，3个辅助蛋白和盒H/ACA RNA形成的RNA-蛋白质复合物。这种核糖核蛋白（RNP）复合物在古细菌和人类之间是保守的，并且对于核糖体RNA的加工和成熟是必需的。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组装和稳定性都是必不可少的。我们将使用X射线晶体学可视化核糖开关和框H/ACA RNP在多个功能状态。我们将结合联合收割机的结构信息与生化探测和溶液酶学来推断这些分子机器是如何工作的。蛋白质和RNA，使生命成为可能的分子机器，具有复杂的结构。为了了解细胞RNA机器是如何工作的，我们将在它们执行功能的不同时间捕获它们，并使用一种称为X射线晶体学的技术来找出它们的组成原子在不同时间点的位置。这些“分子电影”将提供基本的理解，这将有助于操纵RNA在人类健康和疾病以及病原体生命周期中的功能。

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.