CHARACTERIZATION OF LIGAND RECOGNITION OF A METABOLITE-RESPONSIVE RIBOZYME

代谢物响应核酶的配体识别特征

• 批准号: 7725203
• 负责人: JEFF SOUKUP
• 金额: $ 3.94万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7725203
• 关键词: 5' Untranslated Regions; Address; Affect; Amines; Catalysis; Catalytic RNA; Cell Wall; Cells; Computer Retrieval of Information on Scientific Projects Database; Enzymes; Funding; Gene Expression; Genetic; Glucosamine; Glucose; Gram-Positive Bacteria; Grant; Institution; Length; Ligands; Measures; Mediating; Messenger RNA; Metabolic; Molecular; Names; Proteins; RNA; Rate; Reaction; Research; Research Personnel; Resources; Role; Source; Translational Repression; United States National Institutes of Health; base; functional group; glucosamine 6-phosphate; inorganic phosphate; interest; novel; response; serinol

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. The metabolic state of a cell typically affects gene expression through protein-mediated mechanisms of transcriptional or translational control. Yet recent studies have shown that messenger RNAs (mRNAs) can directly sense and respond to specific metabolites through intrinsic domains termed riboswitches. In the presence of metabolites, structural changes in riboswitch domains can result in either transcriptional termination or translational repression. Recently a novel catalytic riboswitch has been discovered that exerts genetic control through self-cleavage of the nascent RNA in response to cellular metabolite concentration. The metabolite-dependent ribozyme resides in the 5'-untranslated region of the glmS mRNA of numerous Gram-positive bacteria and it catalyzes an internal phosphoester transfer reaction that results in cleavage and inactivation of the mRNA. The ribozyme selectively recognizes and is 1000-fold activated by glucosamine-6-phosphate, the metabolic product of the GlmS enzyme, and an important component of bacterial cell walls. We are interested in understanding the molecular basis of ligand recognition and catalysis by the glmS riboswitch. To address this we measured the rate of self-cleavage of the ribozyme/riboswitch in response to a panel of related but distinct ligands, serinol, glucose and glucosamine, to name a few. We have determined that the ribozyme makes important contacts to the amine and phosphate in the natural ligand. We are interested in determining how these two functional groups are interacting with the full-length glmS ribozyme in order to fully understand its role in catalysis.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 细胞的代谢状态通常通过蛋白质介导的转录或翻译控制机制影响基因表达。 然而最近的研究表明，信使 RNA (mRNA) 可以通过称为核糖开关的内在结构域直接感知特定代谢物并对其做出反应。 在代谢物存在的情况下，核糖开关结构域的结构变化可能导致转录终止或翻译抑制。 最近发现了一种新型催化核糖开关，它通过响应细胞代谢物浓度而自我切割新生 RNA 来发挥遗传控制。 代谢物依赖性核酶位于许多革兰氏阳性细菌的 g​​lmS mRNA 的 5'-非翻译区，它催化内部磷酸酯转移反应，导致 mRNA 的裂解和失活。 该核酶选择性识别 6-磷酸葡萄糖胺并被其激活 1000 倍，6-磷酸葡萄糖胺是 GlmS 酶的代谢产物，也是细菌细胞壁的重要组成部分。 我们有兴趣了解 glmS 核糖开关配体识别和催化的分子基础。 为了解决这个问题，我们测量了核酶/核糖开关响应一组相关但不同的配体（丝氨醇、葡萄糖和葡萄糖胺）的自裂解速率。 我们已经确定核酶与天然配体中的胺和磷酸盐具有重要的接触。 我们有兴趣确定这两个官能团如何与全长 glmS 核酶相互作用，以便充分了解其在催化中的作用。