Discovery and Biosynthesis of Bacterial Terpenoids

细菌萜类化合物的发现和生物合成

• 批准号: 10667588
• 负责人: Jeffrey Daniel Rudolf
• 金额: $ 36.3万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10667588
• 关键词: Actinobacteria class; Address; Anabolism; Antibiotics; Antimalarials; Bacteria; Biochemistry; Bioinformatics; Biological; Biomedical Engineering; Biotechnology; Chemicals; Clinical; Collaborations; Complex; Cyclization; Cytochrome P450; Dedications; Development; Engineering; Enzymatic Biochemistry; Enzymes; Family; Future; Gene Cluster; Genome; Genomics; Immunosuppressive Agents; In Vitro; Libraries; Medicine; Methodology; Mining; Natural Product Drug; Natural Products; Nature; Outcomes Research; Pathway interactions; Pharmaceutical Preparations; Property; Proteins; Research; Source; Structure-Activity Relationship; Systems Development; Terpenes; Terpenoid Biosynthesis Pathway; Up-Regulation; X-Ray Crystallography; anti-cancer; combinatorial; drug development; drug discovery; experience; in vivo; innovation; insight; interdisciplinary approach; metabolomics; novel; novel therapeutics; programs; small molecule; success; synthetic biology; terpene synthase; tool

PROJECT SUMMARY/ABSTRACT The long-term objectives of our research program are to (i) discover novel bacterial natural products (NPs), (ii) elucidate the biosynthetic pathways and regulatory mechanisms of these NPs, and (iii) characterize and utilize the discovered NPs and their biosynthetic enzymes for biomedical and biotechnological applications. NPs are highly functionalized and evolutionarily optimized small molecules that possess unrivaled chemical and structural diversities, resulting in a wide range of biological activities. Terpenoids, the largest and most structurally diverse family of NPs, are considered rare in bacteria; only ~1.2% of known terpenoids are of bacterial origin. However, genomics studies revealed that the biosynthetic enzymes responsible for terpenoid biosynthesis are widely distributed in bacteria, particularly actinobacteria. We hypothesize that (i) bacterial terpenoids are considerably underestimated among current NP libraries and the discovery and characterization of novel terpenoids will lead to new drug leads and (ii) understanding the sequence-structure-function relationships of terpenoid biosynthetic enzymes will lead to new opportunities in genome mining, combinatorial biosynthesis, and oxidative biocatalysis. Our initial efforts follow two research directions that address immediate needs and will set the stage for continued success in the field of terpenoid discovery and biosynthesis. In the first direction, we will use an integrated genomics–metabolomics approach to discovery novel bacterial terpenoids from bacteria. This will include the development of new and innovative methodologies for targeted identification of complex bacterial terpenoids and the activation or upregulation of terpenoid biosynthetic gene clusters. In the second direction, we will elucidate the biosynthetic pathways of both new and known bacterial terpenoids and functionally, mechanistically, and structurally characterize terpene synthases and their associated oxidative enzymes, particularly cytochrome P450s. We will use a rigorous multidisciplinary approach involving genome mining, bioinformatics analysis, in vivo pathway engineering, (un)natural product isolation and structural determination, in vitro enzymology, and protein X-ray crystallography. Our experience in terpenoid biosynthesis and enzymology and our significant progress in both research directions supports the feasibility of the proposed research and that we are well-suited to establish and sustain a successful independent program in this field. In addition, we have established several key collaborations with leaders in the fields of synthetic biology, NP drug discovery, and X-ray crystallography that further strengthen this research program. Expected outcomes of this research program include the revelation of the bacterial terpenome, understanding the underlying principles of how terpene synthases dictate terpene cyclization, and the exploitation of naturally evolved oxidative enzymes to create a toolbox of biocatalysts.

项目摘要/摘要 我们的研究计划的长期目标是（i）发现新型细菌天然产品 （NP），（ii）阐明这些NP的生物合成途径和调节机制，（iii）表征 并利用发现的NP及其生物合成酶进行生物医学和生物技术应用。 NP具有高度功能化和进化优化的小分子，具有无与伦比的化学物质 和结构多样性，导致广泛的生物学活动。 Terpenoids，最大，最大的 结构上多样的NP家族在细菌中被认为很少见。只有约1.2％的已知萜类化合物是 细菌起源。然而，基因组学研究表明，负责萜类化合物的生物合成酶 生物合成广泛分布在细菌中，尤其是静脉细菌。我们假设（i）细菌 当前的NP库和发现和表征，萜类化合物被仔细地低估了 新型萜类化合物将导致新药铅，（ii）了解序列结构函数 萜类生物合成酶的关系将导致基因组开采的新机会 生物合成和氧化生物催化。我们最初的努力遵循两个研究方向，解决了 直接需求，并将为在Terpenoid Discovery和 生物合成。在第一个方向上，我们将使用集成的基因组学 - 实谢代谢方法进行发现 来自细菌的新型细菌萜类化合物。这将包括开发新的和创新的 靶向鉴定复杂细菌的方法以及激活或上调的方法 萜类生物合成基因簇。在第二个方向上，我们将阐明 新的和已知的细菌萜类化合物，以及在功能，机械上和结构上表征的 萜烯合酶及其相关的氧化酶，尤其是细胞色素P450。我们将使用一个 严格的多学科方法涉及基因组挖掘，生物信息学分析，体内途径 工程，（联合国）天然产品隔离和结构测定，体外酶学和蛋白质X射线 晶体学。我们在萜类生物合成和酶学方面的经验以及我们在 这两个研究方向都支持拟议研究的可行性，我们非常适合 在该领域建立并维持成功的独立计划。此外，我们已经建立了几个 与合成生物学，NP药物发现和X射线领域领导者的主要合作 晶体学进一步加强了该研究计划。该研究计划的预期结果 包括细菌萜烯组的启示，了解萜烯的基本原理 合成酶决定萜烯环化，并剥削自然进化的氧化酶以创建一种 生物催化剂的工具箱。