Genomic approaches of discovery broad-spectrum antimicrobial agents

发现广谱抗菌药物的基因组方法

• 批准号: 7669772
• 负责人: Jon Clardy
• 金额: $ 34.21万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-03-25 至 2014-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7669772
• 关键词: Actinobacteria class; Actinomyces; Actinomycetales; Antibiotics; Bacteria; Bioinformatics; Biological Factors; Chemistry; Culture Media; Ecology; Elements; Environment; Funding; Gene Cluster; Genetic; Genome; Genomics; Goals; Grant; Institutes; Methodology; Microbe; National Institute of Allergy and Infectious Disease; New England; Nutrient; Pilot Projects; Production; Resistance; Screening procedure; Simulate; antimicrobial; antimicrobial drug; base; biodefense; combat; microbial; novel; pathogen; programs; promoter; scaffold; small molecule; tool

Most clinically-used antibiotics are bacterially-produced small molecules known as natural products, or their derivatives. However, natural products are not being featured in antibiotic discovery programs due to high rates of rediscovery and low rates of new molecule discovery. This proposal describes a new genomicsbased approach to natural product discovery that employs a combination of bioinformatics, microbial ecology, genetics, and chemistry to identify cryptic biosynthetic loci and stimulate them to produce their encoded natural products. All molecules identified will be characterized for antimicrobial activity. Specific Aim 1. Use bioinformatics to identify biosynthetic gene clusters and predict their products. New bioinformatic tools will be developed to identify biosynthetic gene clusters in the genomes of actinomycetes and other microbes, and to predict structural elements of their small molecule products. These predictions will be leveraged toward antibiotic discovery by using them as the basis for efforts to stimulate the production of cryptic natural products, as described in Specific Aims 2 and 3: Specific Aim 2. Stimulate the production of cryptic metabolites by simulating a multispecies environment. Most screens for new natural products have been performed with strains grown as pure cultures in nutrientrich growth media. A new screening format has recently been developed in which strains are grown as microcolonies on nutrient-poor growth media. This microcolony screening methodology will be used to identify cryptic natural products with antimicrobial activity. Specific Aim 3. Stimulate the production of cryptic metabolites by genetically manipulating producers. Most natural product-encoding gene clusters are thought to be repressed under standard culture conditions. Using actinomycete strains whose biosynthetic gene clusters have been identified by the efforts described in Specific Aim 1, endogenous gene cluster promoters will be systematically replaced with a strong, inducible promoter, enabling the controlled synthesis and isolation of their small molecule products and the subsequent characterization of their antimicrobial activity. .

大多数临床使用的抗生素是细菌产生的小分子，称为天然产物，或其衍生物。 衍生物.然而，天然产品并没有在抗生素发现计划中得到重视， 重新发现的比率和新分子发现的低比率。该提案描述了一种新的基于基因组学的 结合生物信息学、微生物技术的天然产物发现方法 生态学、遗传学和化学，以确定隐藏的生物合成基因座，并刺激它们产生它们的 编码的天然产物。将对所有鉴定的分子进行抗微生物活性表征。 具体目标1.使用生物信息学来识别生物合成基因簇并预测其产物。新 将开发生物信息学工具来识别放线菌基因组中的生物合成基因簇 和其他微生物，并预测其小分子产物的结构元素。这些预测 将利用它们作为刺激生产的基础， 如具体目标2和3所述， 具体目标2。通过模拟多物种环境刺激隐蔽代谢物的产生。 大多数新天然产物的筛选都是用在nutrientrich中作为纯培养物生长的菌株进行的 生长介质。最近开发了一种新的筛选形式，其中菌株生长为 营养不良的生长培养基上的小菌落。这种小菌落筛选方法将用于 鉴定具有抗微生物活性的神秘天然产物。 具体目标3。通过基因操纵生产者刺激神秘代谢物的生产。最 天然产物编码基因簇被认为在标准培养条件下受到抑制。 使用放线菌菌株，其生物合成基因簇已经通过在 具体目标1，内源基因簇启动子将被系统地替换为强的、可诱导的 启动子，使得能够控制其小分子产物的合成和分离， 随后表征其抗微生物活性。.