Molecular Analysis of Corrinoid Specificity

类咕啉特异性的分子分析

• 批准号: 9311076
• 负责人: Michiko E. Taga
• 金额: $ 37.08万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9311076
• 关键词: Affinity; Anabolism; Bacteria; Bacterial Genome; Bacterial Physiology; Binding; Biochemical Genetics; Bioinformatics; Biological Assay; Biosensor; Cobalt; Communities; Corrinoids; Data; Development; Disease; Elements; Enzymes; Family; Fluorescence; Genetic; Genome; Goals; Growth; Health; Human; Human Microbiome; In Vitro; Intestines; Knowledge; Ligands; Link; Measurement; Measures; Metabolic Pathway; Metabolism; Methods; Microbe; Modeling; Molecular; Molecular Analysis; Mutagenesis; Mutase; Organism; Pathway interactions; Population; Prokaryotic Cells; RNA; RNA Sequences; Regulatory Element; Reporter; Research; Robotics; Role; Sequence Analysis; Specificity; Testing; Tetrapyrroles; Variant; Vitamin B 12; Work; analog; base; cofactor; design; enzyme model; gain of function; gut microbiome; gut microbiota; improved; in vivo; interest; member; methylmalonyl-coenzyme A; microbial community; microbiota; microorganism; mutant; novel strategies; preference; prevent; tool

Project Summary The composition of microorganisms that reside in the human intestine (gut microbiota) has a significant impact on human health. Despite recent advances in understanding the role of the gut microbiota in health, there remains a need for effective mechanisms of manipulating the gut microbiota to treat and prevent disease. One potential target for manipulation of the gut community is the corrinoid family of cofactors (vitamin B12 and related cobalt-containing modified tetrapyrroles), which are required by the majority of bacteria in the gut, though they are produced only by a fraction of the population. A variety of structural forms of corrinoids exist that are not functionally equivalent, yet little is known about the link between bacterial genome sequences and adaptations for the use of particular corrinoids. The goal of this proposal is to determine the relationship between genome sequence and corrinoid preferences in gut microbes. This goal will be accomplished by examining bacterial corrinoid preferences at the molecular, organismal, and community levels. Corrinoid specificity of homologs of a model corrinoid-dependent enzyme will be dissected by a combination of biochemical, genetic, and bioinformatic approaches. In parallel, corrinoid specificity of riboswitch RNAs, which are present in the majority of genomes in the human microbiome, will be examined by using an in vitro fluorescence-based binding assay and in vivo reporter assays. These data will be used to predict sequence signatures of corrinoid specificity and test these predictions by targeted mutagenesis. Additionally, corrinoid preferences will be examined in cultured human gut bacteria by measuring growth under corrinoid-requiring conditions using a high-throughput robotics platform. This work will lead to an improved ability to interpret genome sequence information and provide a detailed characterization of metabolic processes critical to the majority of human gut microbes. The results will promote the development of novel strategies to prevent and treat diseases associated with the gut microbiota.

项目摘要 存在于人肠（肠道菌群）的微生物的组成具有重大影响 关于人类健康。尽管最近在了解肠道微生物群在健康中的作用方面取得了进步，但 仍然需要有效操纵肠道菌群治疗和预防疾病的机制。一 操纵肠道群落的潜在目标是辅因子的类型（维生素B12及相关的） 含钴的修饰四吡咯），这是肠道中大多数细菌所要求的 仅由人口的一小部分产生。存在多种不存在的结构形式 在功能上等效，但对细菌基因组序列与适应之间的联系知之甚少 用于使用特定的类似石。该提议的目的是确定基因组之间的关系 肠道微生物中的序列和类cor骨偏好。该目标将通过检查细菌来实现 分子，生物和社区层面的类corrinoid偏好。同源物的类contrinofics 模型依赖性酶将通过生化，遗传和生物学的组合解剖 方法。同时，核糖开关RNA的类crorinoid特异性，这些特异性存在于大多数基因组中 人类微生物组将通过使用基于体外荧光的结合测定和体内检查 记者测定法。这些数据将用于预测类corrinoid特异性的序列特异性和测试 通过靶向诱变进行预测。此外，将在培养的人类肠道中检查类corinoid偏好 通过使用高通量机器人平台测量菌皮征收条件下的生长，细菌。 这项工作将提高解释基因组序列信息并提供详细的能力 代谢过程的表征对大多数人类肠道微生物至关重要。结果将促进 预防和治疗与肠道菌群相关的疾病的新型策略的发展。