Cytochrome P450 Enzymes from Streptomyces: Diverse Biocatalysts in Natural Products Biosynthesis

来自链霉菌的细胞色素 P450 酶：天然产物生物合成中的多种生物催化剂

• 批准号: 9371636
• 负责人: Jeffrey Daniel Rudolf
• 金额: $ 9万
• 依托单位: SCRIPPS FLORIDA
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-09-18 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9371636
• 关键词: Actinomyces Infections; Actinomycetales; Advisory Committees; Anabolism; Biochemical; Bioinformatics; Biological; Biological Process; Biology; Biomedical Engineering; Biotechnology; Breathing; Catalysis; Chemicals; Chemistry; Collection; Cytochrome P450; Data; Databases; Development; Development Plans; Diterpenes; Drug Metabolic Detoxication; Engineering; Ensure; Environment; Enzymatic Biochemistry; Enzymes; Foundations; Future; Generations; Genetic; Genome; Genomics; Goals; Gram-Positive Bacteria; Hemeproteins; Homologous Gene; Hormones; Human; Hydroxylation; In Vitro; Life; Mammals; Mentors; Metabolism; Mining; Modeling; Molecular; Natural Product Drug; Natural Products; Nature; Organic Synthesis; Outcome; Pathway Analysis; Pathway interactions; Phase; Physiological; Positioning Attribute; Proteins; Research; Research Institute; Research Project Grants; Research Training; Roentgen Rays; Role; Science; Site-Directed Mutagenesis; Skeleton; Streptomyces; Structure; Structure-Activity Relationship; Terpenes; Testing; Time; Training; Universities; Utah; Variant; X-Ray Crystallography; Xenobiotics; analog; biosynthetic product; career; career development; chemical reaction; design; drug development; drug metabolism; experience; genetic manipulation; hormone biosynthesis; human disease; in vivo; innovation; interdisciplinary approach; microbial; molecular array; novel; post-doctoral training; preference; programs; scaffold; small molecule; structural biology; tenure track

Project Summary/Abstract Research. The goal of this K99/R00 project is to test the hypothesis that cytochromes P450 from Actinomycetales, a diverse subfamily of remarkable biocatalysts, may be used as a means to discover natural products with novel functionalities. We propose to (Aim 1) investigate the sequence–structure–function relationships of novel P450s from selected natural product biosynthetic pathways, (Aim 2) use a P450-focused genome mining approach to discover natural products that have undergone unique P450-catalyzed transformations, and (Aim 3) employ the newly identified P450s, and their homologues, as models for P450 enzymology and bioengineering. The multidisciplinary approach involves bioinformatics analysis, in vitro enzymology, protein X-ray crystallography, in vivo pathway engineering, (un)natural product isolation and structural determination. Superb preliminary data supports both the feasibility of the proposed mentored phase in a two-year time span, and a successful transition to an independent research program. The innovation of this application lies in leveraging the diversity of natural P450 variants from a proprietary Actinomycetales strain collection for discovering unprecedented chemistries that imbue natural products with new functionalities. The significance of this project is the advancement of P450 catalysis and enzymology and the discovery of novel natural products, laying the foundation for future treatments of human diseases. Candidate and Environment. Dr. Rudolf obtained graduate training in mechanistic enzymology and organic synthesis from Dr. C. Dale Poulter, a preeminent terpenoid biochemist, at the University of Utah. He then joined Dr. Ben Shen, a recognized leader in natural products discovery and biosynthesis, at The Scripps Research Institute (TSRI), and received postdoctoral training in actinomycetes genetics and natural products biosynthesis. His long-term career goals include establishing an independent, competitive research program that emphasizes using natural products as inspirations for the discovery and study of novel chemistries, enzymologies, and biologies. His immediate career goals include obtaining training in P450 enzymology and structural biology, and in utilizing genomics for the discovery of novel enzymes and natural products. Career development plan. This proposal was designed to ensure the realization of his career goals by providing intellectual, technical, and professional training during the mentored phase that will lead to an effective transition to an independent, tenure-track position. The primary mentor, Dr. Shen, and co-mentor, Dr. George Phillips, an expert in X-ray crystallography and heme proteins, were chosen for their scientific expertise and mentoring experience. An advisory committee, including Drs. Bradley Moore, Wilfred van der Donk, Sean Brady, and Michael Smanski, all experts in natural products, was explicitly assembled to oversee both the science and career development aspects of this proposal. The research training and excellent mentoring will be supplemented with numerous opportunities for scientific and professional development available at TSRI.

项目总结/摘要 Research.这个K99/R 00项目的目标是检验细胞色素P450从 放线菌目（Actinomycetales）是一种多样的生物催化剂亚科，可用于发现天然的生物催化剂。 具有新功能的产品。我们提出（目的1）研究序列结构功能 从选定的天然产物生物合成途径的新型P450的关系，（目的2）使用P450为重点， 基因组挖掘方法，以发现经过独特的P450催化的天然产物 转化，（目的3）采用新鉴定的P450及其同系物作为P450的模型 酶学和生物工程。多学科方法包括生物信息学分析，体外 酶学，蛋白质X射线晶体学，体内途径工程，（非）天然产物分离和 结构决定一流的初步数据支持的可行性，建议辅导 在为期两年的时间跨度阶段，并成功过渡到一个独立的研究计划。的 这种应用的创新在于利用天然P450变体的多样性， 放线菌目菌株收集，用于发现前所未有的化学物质， 新功能。该项目的意义在于P450催化和酶学的进步， 发现新的天然产物，为未来治疗人类疾病奠定基础。 候选人与环境Rudolf博士获得了机械酶学和有机酶学的研究生培训。 由C博士合成。犹他州大学杰出的萜类生物化学家戴尔·保尔特说。然后他 加入了Ben Shen博士，他是天然产物发现和生物合成领域公认的领导者， 研究所（TSRI），并接受放线菌遗传学和天然产物博士后培训 生物合成他的长期职业目标包括建立一个独立的、有竞争力的研究项目 它强调使用天然产物作为发现和研究新化学的灵感， 酶学和生物学。他的近期职业目标包括获得P450酶学方面的培训， 结构生物学，以及利用基因组学发现新的酶和天然产物。职业生涯 发展规划这一建议旨在确保实现他的职业目标， 知识，技术和专业培训在指导阶段，这将导致有效的 过渡到一个独立的终身职位主要导师沈博士和共同导师乔治博士 菲利普斯是X射线晶体学和血红素蛋白的专家，他的科学专长被选中， 指导经验。一个咨询委员会，包括布拉德利摩尔博士，威尔弗雷德货车德唐克，肖恩 布雷迪和迈克尔·斯曼斯基都是天然产品方面的专家，他们被明确地召集起来， 科学和职业发展方面的建议。研究培训和优秀的指导将 在TSRI提供的科学和专业发展的众多机会的补充。

项目成果

The Natural Products Atlas 2.0: a database of microbially-derived natural products.

• DOI: 10.1093/nar/gkab941
• 发表时间: 2022-01-07
• 期刊: Nucleic acids research
• 影响因子: 14.9
• 作者: van Santen JA;Poynton EF;Iskakova D;McMann E;Alsup TA;Clark TN;Fergusson CH;Fewer DP;Hughes AH;McCadden CA;Parra J;Soldatou S;Rudolf JD;Janssen EM;Duncan KR;Linington RG
• 通讯作者: Linington RG