VISUALIZING ALLOSTERY IN THE GENE-REGULATORY LYSINE RIBOSWITCH

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目及 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 核糖开关是近年来发现的一类RNA结构域，在细菌和真核生物中直接与细胞代谢产物结合，控制基因顺式表达。由于它们在细胞内稳态中的作用以及它们作为抗菌药物靶点的潜力，核糖开关的结构研究具有直接的意义。已经报道了与其同源代谢物结合的几种类型的核糖开关的晶体结构。然而，由于缺乏关于无代谢物构象的结构信息，核糖开关功能的机制尚未得到很好的理解。 本研究的目的是通过小角X射线散射（SAXS）技术来探测四种核糖开关的无代谢物构象和代谢物结合构象的溶液结构：焦磷酸硫胺素（TPP）、黄素单核苷酸（FMN）、甘氨酸（Gly）和赖氨酸（Lys）核糖开关。甘氨酸核糖开关的先前研究已经证明了SAXS研究（Lipfert等人，JMB，2007）在研究这些RNA适体的整体大小和形状方面的效用。结合代谢物后诱导的构象变化预计约为5 - 20埃，非常适合在APS进行实验。对代谢物自由构象和代谢物结合构象进行SAXS研究将提供对核糖开关基因调控反应所涉及的全局结构变化的深入了解。获得这些构象的Rg和P（r）图将使我们能够在基因调控转换事件之前和之后对RNA进行建模。这些研究将补充并行结构测定的代谢产物结合构象的X-射线晶体学，核糖开关功能提供了详细的结构解释。

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. Riboswitches are recently described RNA domains that directly bind to cellular metabolites and control gene expression in cis in bacteria and eukaryotes. Due to their role in cellular homeostasis and their potential as antibacterial drug targets, structural studies of riboswitches are of immediate interest. Crystal structures of several types of riboswitches bound to their cognate metabolites have been reported. However, the mechanism of riboswitch function is not well understood due to the lack of structural information regarding the metabolite-free conformation. The goal of this proposal is to probe the solution structure of the metabolite-free conformation and the metabolite-bound conformation of four riboswitches by SAXS: the thiamine pyrophosphate (TPP), flavin mononucleotide (FMN), glycine (Gly), and lysine (Lys) riboswitches. Previous studies of the glycine riboswitch have demonstrated the utility of SAXS studies (Lipfert et al, JMB, 2007) to investigate the global size and shape of these RNA aptamers. The conformational change induced upon binding metabolite is expected to be on the order of 5 - 20 Angstroms and is well suited for experimentation at the APS. Performing SAXS studies on both the metabolite-free and metabolite-bound conformations will provide insight into the global structural changes involved in the gene-regulatory response of riboswitches. Obtaining Rg and P(r) plots for these conformations will allow us to model the RNA before and after the gene-regulatory switching event. These studies will complement concurrent structure determination of the metabolite bound conformation by X-ray crystallography, providing a detailed structural explanation of riboswitch function.