Structure and Function of S-adenosyl-L-methionine Riboswitches

描述（由申请人提供）：核糖开关是识别特定小分子（通常是关键代谢物）并在转录或翻译水平上“打开”或关闭下游基因表达的调节RNA。这些短顺式作用RNA元件的发现极大地改变了我们对遗传调控机制的理解。核糖开关在革兰氏阳性细菌中特别普遍，例如枯草芽孢杆菌作为模式生物，但也发现其控制重要病原体如炭疽芽孢杆菌、葡萄球菌、肠球菌、链球菌、李斯特菌、梭菌和分枝杆菌中的必需基因。这一特性和其他特性引起了人们对核糖开关介导调控的越来越多的关注。S-腺苷甲硫氨酸（SAM）核糖开关的三个不同的类别是最常见的核糖开关类的性质。这些RNA代表了实现特异性SAM识别的三种独立进化解决方案。我们最近确定了两类SAM核糖开关的晶体结构，E。faecalis SMK盒和B. subtilis S box.这些结构揭示了SAM是如何被特异性识别的，但没有提供足够的证据来支持在以前的遗传和生化研究中观察到的SAM依赖性构象变化。为了全面了解它们的结构-功能关系和构象动力学，我们提出：（1）了解SMK box核糖开关中的配体识别机制。(2)表征无配体的SMK构象并寻找真核核糖开关。(3)进行化学探测实验，以揭示SMK RNA中配体诱导的构象动力学 公共卫生相关性：所描述的SAM核糖开关的结构-功能研究包括合理设计可能含有抗生素活性的特定核糖开关抑制剂和开发荧光RNA标记系统以供研究界跟踪活细胞中RNA的位置、浓度和运动的实际应用。

DESCRIPTION (provided by applicant): Riboswitches are regulatory RNAs that recognize specific small molecules, usually key metabolites, and "switch" downstream gene expression on or off at either the transcriptional or translational level. The discovery of these short cis-acting RNA elements has drastically changed our understanding of genetic regulatory mechanisms. Riboswitches are especially prevalent in Gram-positive bacteria, exemplified by Bacillus subtilis as a model organism, but are also found to control essential genes in important pathogens such as Bacillus anthracis, Staphylococcus, Enterococcus, Streptococcus, Listeria, Clostridium, and Mycobacterium. This and other characteristics have attracted increasing attention to riboswitch-mediated regulation. The three distinct classes of S-adenosyl methionine (SAM) riboswitches are the most commonly found riboswitch classes in nature. These RNAs represent three independent evolution solutions to achieve specific SAM recognition. We recently determined the crystal structures of SAM riboswitches from two classes, the E. faecalis SMK box and the B. subtilis S box. These structures shed light into the how SAM is specifically recognized, but did not provide enough evidence to support the large SAM-dependent conformational changes observed in the previous genetic and biochemical studies. To fully understand their structure-functional relationship and conformational dynamics, we propose to: (1) Understand the ligand recognition mechanism in the SMK box riboswitch. (2) Characterize the ligand-free SMK conformation and search for eukaryotic riboswitches. (3) Carry out chemical probing experiments to reveal ligand-induced conformational dynamics in the SMK RNA PUBLIC HEALTH RELEVANCE: The described structure-function studies of SAM riboswitches include practical applications to rationally design specific riboswitch inhibitors that may contain antibiotic activities and to develop a fluorescent RNA tagging system for the research community to track the location, concentration, and movement of RNA in living cells.