Mining Actinomycetal Genomes for Natural Product Discovery and Biosynthesis

挖掘放线菌基因组用于天然产物发现和生物合成

• 批准号: 10064144
• 负责人: Ben Shen
• 金额: $ 79.55万
• 依托单位: SCRIPPS FLORIDA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-01-01 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10064144
• 关键词: Acids; Actinomycetales; Anabolism; Biological; Biological Models; Biology; Chemical Structure; Chemistry; Collection; Complex; Crude Extracts; Diterpenes; Engineering; Enzymatic Biochemistry; Evaluation; Evaluation Studies; Family; Gene Cluster; Genome; Goals; Grant; Hybrids; Knowledge; Laboratories; Libraries; Medicine; Metabolic Pathway; Mining; Modeling; Molecular; National Institute of General Medical Sciences; Natural Products; Outcome; Peptides; Pharmaceutical Preparations; Production; Research; Research Institute; Source; Structure; Sulfur; Surveys; Technology; analog; drug discovery; genetic manipulation; genome sequencing; insight; leinamycin; member; microbial; microorganism; novel; programs; scaffold; small molecule

PROJECT SUMMARY Natural products (NPs) continue to inspire novel chemistry, biology, and medicine, but the rate of discovery of novel NPs has slowed dramatically as most gene clusters encoding their biosynthesis are silent when the microorganisms are cultured under standard laboratory conditions. In this MIRA application, we propose to discover new NPs by mining actinomycetal genomes, develop enabling technologies to produce the target NPs in enough quantities for structural elucidation and biological evaluation, and characterize their biosynthetic machineries to discover new chemistry and enzymology. Our hypotheses have been: (i) genome survey of the actinomycetale strain collection at The Scripps Research Institute (TSRI) will allow us to identify potential producers of the targeted NP scaffolds, (ii) genome sequencing of the potential producers for the targeted biosynthetic gene clusters (BGCs) will allow us to predict the structural novelty of the new NPs, (iii) genetic manipulation of the most promising BGCs in their native producers or expression of them in heterologous model hosts will allow us to produce and isolate the new NPs, and (iv) characterization of the biosynthetic machinery of these new NPs will allow us to discover new chemistry and enzymology. Studies from two past and two current NIGMS grants have cumulated into (i) the construction of the Natural Products Library at TSRI that currently consists of ~211,000 microbial strains, ~461,000 crude extracts and partially purified fractions prepared from selected strains, and ~600 pure NPs, (ii) the establishment of C-1027 and tiancimycin as model systems for enediyne NP biosynthesis, leinamycin and guangnanmycin as model systems for biosynthesis of the leinamycin family of NPs (i.e., hybrid peptide-polyketide macrolactams featuring unprecedented sulfur functionalities), and platensimycin and platencin as model systems for bacterial diterpenoid and thiocarboxylic acid-containing/derived NP biosynthesis, respectively, and (iii) the identification of 190, 43, 66, and 264 distinct BGCs, encoding new enediynes, new members of the leinamycin family, new bacterial diterpenoids, and new thiocarboxylic acid-containing/derived NPs, respectively. These findings set the stage in the current application to mine the actinomycetal genomes for NP discovery and biosynthesis. The outcomes of this application include (i) fundamental contributions to genome mining and activation of large BGCs, in native producers and heterologous model hosts, for NP production and structural diversity by metabolic pathway engineering, (ii) discovery of new NPs, with privileged scaffolds, to inspire new chemistry, biology, and medicine, and (iii) new insights into biosynthetic machineries and novel chemistry and enzymology for the biosynthesis of enediynes, the leinamycin family of NPs, bacterial diterpenoids, and thiocarboxylic acid- containing/derived NPs. The long-term goal of our research is to understand at a molecular level how microorganisms synthesize complex NPs and to exploit this knowledge to discover novel NPs and engineer their analogues for drug discovery.

项目概要 天然产物 (NP) 继续激发新的化学、生物学和医学的灵感，但其发现速度 新型纳米粒子的速度已显着减慢，因为大多数编码其生物合成的基因簇在 微生物在标准实验室条件下培养。在此 MIRA 应用程序中，我们建议 通过挖掘放线菌基因组发现新的纳米颗粒，开发生产目标纳米颗粒的技术 足够数量用于结构阐明和生物学评价，并表征其生物合成 发现新化学和酶学的机器。我们的假设是：（i）基因组调查 斯克里普斯研究所 (TSRI) 的放线菌菌株收藏将使我们能够识别潜在的 目标 NP 支架的生产者，(ii) 目标 NP 潜在生产者的基因组测序 生物合成基因簇 (BGC) 将使我们能够预测新 NP 的结构新颖性，(iii) 遗传 在其本地生产者中操纵最有前途的 BGC 或在异源中表达它们 模型宿主将使我们能够生产和分离新的纳米粒子，以及（iv）生物合成的表征 这些新纳米粒子的机制将使我们能够发现新的化学和酶学。过去两届的研究 目前 NIGMS 的两项赠款已累计用于 (i) TSRI 天然产物图书馆的建设 目前由约 211,000 种微生物菌株、约 461,000 种粗提物和部分纯化的馏分组成 由选定的菌株和约 600 个纯 NP 制备，(ii) 建立 C-1027 和天赐霉素作为模型 烯二炔 NP 生物合成系统，来那霉素和广南霉素作为生物合成的模型系统 NP 的莱纳霉素家族（即杂合肽-聚酮大环内酰胺，具有前所未有的硫 功能），以及platensimycin和platencin作为细菌二萜和硫代羧酸的模型系统 分别鉴定含酸/衍生的 NP 生物合成，以及 (iii) 鉴定 190、43、66 和 264 个不同的 BGC，编码新的烯二炔、莱那霉素家族的新成员、新的细菌二萜类化合物和新的 分别是含硫代羧酸/衍生的纳米颗粒。这些发现为当前的研究奠定了基础 应用挖掘放线菌基因组以进行 NP 发现和生物合成。这样做的结果 应用包括（i）对基因组挖掘和大型 BGC 激活的基本贡献，在天然 生产者和异源模型宿主，通过代谢途径实现 NP 生产和结构多样性 工程，(ii) 发现新的纳米粒子，具有特殊的支架，以激发新的化学、生物学和 医学，以及（iii）对生物合成机械和新型化学和酶学的新见解 烯二炔、莱纳霉素纳米颗粒家族、细菌二萜类化合物和硫代羧酸的生物合成 含有/衍生的纳米颗粒。我们研究的长期目标是在分子水平上了解如何 微生物合成复杂的纳米颗粒，并利用这些知识来发现新的纳米颗粒并设计 它们用于药物发现的类似物。