Defining the ligandability of bacterial RNAs

定义细菌 RNA 的配体能力

• 批准号: 10315245
• 负责人: Jordan T Koehn
• 金额: $ 6.64万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10315245
• 关键词: Address; Adopted; Affinity; Antibiotics; Bacillus subtilis; Bacteria; Bacterial RNA; Binding; Binding Sites; Bioinformatics; Biological; Biological Assay; Biology; Cell Culture Techniques; Cell physiology; Cells; Chemicals; Chemistry; Complementary DNA; Complex; Coupling; Data; Data Set; Development; Drug Design; Drug resistance; Escherichia coli; Eukaryotic Cell; Generations; Genes; Goals; Growth; High-Throughput Nucleotide Sequencing; Higher Order Chromatin Structure; Human; Hybrids; Individual; Knowledge; Leadership; Learning; Libraries; Ligand Binding; Ligands; Link; Maps; Measures; Messenger RNA; Methods; Molecular Biology; Natural Products; Nucleotides; Pharmaceutical Preparations; Positioning Attribute; Proteins; RNA; RNA Binding; RNA Sequences; Research Personnel; Research Project Grants; Research Training; Resolution; Reverse Transcription; Ribosomal RNA; Ribosomes; Scientist; Site; Structure; Technology; Thiamine; Thiamine Pyrophosphate; Training; Training Programs; Transcript; Validation; Vision; base; career; cell growth; chemical synthesis; complex data; design; experimental study; high throughput technology; infancy; interest; member; new technology; novel; post-doctoral training; skills; small molecule; structural genomics; targeted treatment; therapeutic target; training project; transcriptome

PROJECT SUMMARY Most bioactive small molecules function by targeting proteins. Complex RNA structures are, in principle, also targetable with small molecules. However, apart from bacterial riboswitches, natural products that bind a small number of sites in the ribosome, and a handful of human-designed ligands, we know very little of the capacity of small molecules to bind RNA or to modulate RNA function. During the first year of my postdoctoral training, I demonstrated the feasibility of a new technology, called Frag-JuMP, that simultaneously identifies RNA-binding ligands and maps RNA-ligand binding sites at nucleotide resolution. The current version of the technology requires gene-specific primers. Therefore, one goal of this project is to develop a generic approach for examining the ligand-binding ability of large RNAs. A second goal is to use Frag-JuMP to interrogate ligand-binding by the bacterial ribosomal RNA in live cells to understand fundamental principles that allow ligands to bind selectively to complex RNAs and to identify RNA-ligand binding sites in functional regions of the ribosome. These studies are expected to characterize RNA structures capable of binding small molecules and to identify the subset of these that constitute ligand- binding sites likely to modulate ribosome function. The long-term, sustained impacts of this research and training project include the creation of a comprehensive RNA-ligand discovery strategy that can immediately be used to discover RNA-binding molecules and to provide an unprecedented picture of ligand-binding sites in any cellular RNA. Molecules identified by Frag-JuMP are also readily linkable, facilitating generation of potent bivalent ligands by linking co-binding fragments. The field of small- molecule, RNA-targeted therapeutics is in its infancy and represents an enormous opportunity to manipulate the functions of diverse cellular processes in bacterial and eukaryotic cells. Training will emphasize development of new expertise in RNA chemical probing methods, technologies based on high- throughput sequencing, bioinformatics analyses, cell culture methods, and small-molecule fragment chemistry. This training, combined with my previously developed expertise in chemical synthesis, will equip me with unique, career transforming, skills in chemical biology and structural genomics. Upon completion of this cross-disciplinary training program, I will be equipped for a leadership position at the interface of chemistry and biology, focused on understanding how RNA structure governs biological mechanism and on developing RNA-targeted therapeutics.

项目摘要 大多数生物活性小分子通过靶向蛋白质发挥作用。复杂的RNA结构，原则上， 也可被小分子靶向。然而，除了细菌核糖开关， 结合核糖体中的少量位点，以及少数人类设计的配体，我们非常清楚， 小分子结合RNA或调节RNA功能的能力很小。第一年 在我的博士后培训中，我展示了一种名为Frag-JuMP的新技术的可行性， 同时识别RNA结合配体并以核苷酸分辨率绘制RNA配体结合位点。 目前的技术版本需要基因特异性引物。因此，本项目的一个目标是 开发一种通用的方法来检查大RNA的配体结合能力。第二个目标是 使用Frag-JuMP询问活细胞中细菌核糖体RNA的配体结合，以了解 允许配体选择性结合复杂RNA并识别RNA配体的基本原理 核糖体功能区的结合位点。这些研究有望表征RNA 能够结合小分子的结构，并确定构成配体的这些结构的子集， 可能调节核糖体功能的结合位点。这项研究的长期、持续影响， 培训项目包括创建一个全面的RNA配体发现策略， 立即被用来发现RNA结合分子，并提供一个前所未有的图片， 任何细胞RNA中的配体结合位点。通过Frag-JuMP鉴定的分子也容易被识别， 通过连接共结合片段促进有效的二价配体的产生。小的领域- 分子中，RNA靶向疗法正处于起步阶段，这代表了一个巨大的机会， 操纵细菌和真核细胞中各种细胞过程的功能。培训将 强调RNA化学探测方法的新专业知识的发展，基于高， 通量测序、生物信息学分析、细胞培养方法和小分子片段 化学.这次培训，结合我以前在化学合成方面的专业知识， 装备我在化学生物学和结构基因组学的独特的，职业转型，技能。后 完成这个跨学科的培训计划，我将配备一个领导职位， 化学和生物学的界面，专注于了解RNA结构如何控制生物学 机制和开发RNA靶向治疗。