Collaborative Research: RUI: Elucidating Design Rules for non-NRPS Incorporation of Amino Acids on Polyketide Scaffolds

合作研究：RUI：阐明聚酮化合物支架上非 NRPS 氨基酸掺入的设计规则

• 批准号: 2300890
• 负责人: Katharine Watts
• 金额: $ 60.82万
• 依托单位: California Polytechnic State University Foundation
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-04-15 至 2027-03-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2300890&HistoricalAwards=false
• 关键词: Collaborative; Research; RUI; Elucidating; Design

More bacteria have become resistant to current antibiotics. Natural compounds with antibiotic properties are assembled in a modular fashion inside cells. The modular nature allows for the design of hybrid pathways to produce non-natural molecules. The mechanisms of individual bond-forming reactions in an antibiotic molecule will be investigated. The intention is to develop a degree of insight that would enable the shuffling of building blocks to produce novel antibiotics. The concept is akin to molecular building blocks. In addition, this work will provide a foundation for training undergraduate students in conducting biotechnological research and communicating science to their community. The project will attempt to uncover novel enzymatic mechanisms that facilitate production of unique natural products. Specifically, this work will elucidate mechanistic underpinnings of amino acid incorporation on polyketide scaffolds. The project focuses on the model molecule TLN-05220, which is an Echinosporamicin-type antibiotic with antibacterial activity against several strains of methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococci (VRE), and cytotoxic activities against several human tumor cell lines. Individual gene functions in TLN-05220 biosynthesis will be identified using gene knockouts in the wild-type producing organism. Potential substrate binding sites and catalytic residues of amino acid-incorporating enzymes will be identified using homology modeling. Expression and characterization of individual enzymes will elucidate mechanism(s) of amino acid incorporation into natural product structures. The minimally necessary genes to produce TLN-05220 will be cloned and expressed in a heterologous host.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

越来越多的细菌对目前的抗生素产生了抗药性。具有抗生素特性的天然化合物在细胞内以模块化的方式组装。模块化性质允许设计混合途径以产生非天然分子。将研究抗生素分子中单个键形成反应的机制。其目的是发展一定程度的洞察力，使改组的积木，以产生新的抗生素。 这个概念类似于分子构建块。 此外，这项工作将为培养本科生进行生物技术研究和科学传播到他们的社区奠定基础。该项目将试图揭示新的酶促机制，促进独特的天然产物的生产。具体而言，这项工作将阐明聚酮化合物支架上的氨基酸掺入的机制基础。该项目的重点是模型分子TLN-05220，这是一种棘孢霉素型抗生素，对几种耐甲氧西林金黄色葡萄球菌和耐万古霉素肠球菌（VRE）菌株具有抗菌活性，对几种人类肿瘤细胞系具有细胞毒性活性。将使用野生型生产生物体中的基因敲除来鉴定TLN-05220生物合成中的单个基因功能。 潜在的底物结合位点和催化残基的氨基酸结合酶将被确定使用同源建模。 单个酶的表达和表征将阐明氨基酸掺入天然产物结构的机制。生产TLN-05220所需的最低限度基因将被克隆并在异源宿主中表达。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。