A Cell-Free Tool for Rapid Screening of Antifungal Peptides

快速筛选抗真菌肽的无细胞工具

• 批准号: 2879917
• 金额: --
• 依托单位: Queen Mary University of London
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2023
• 资助国家: 英国
• 起止时间: 2023 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2879917
• 关键词: Cell; Free; Tool; Rapid; Screening

To date, only one antifungal RiPP has been identified (e.g., pinensin), which is a significant area of opportunity for the discovery of novel RiPPs. As an enabling technology, we will use cell-free transcription-translation (TX-TL) systems, developed by the Moore group3,4. TX-TL systems require DNA, a cell extract, and an energy solution to make recombinant peptides/proteins within minutes to hours, and up to a milligram scale. This method of production also overcomes key challenges and time scale problems with engineering RiPPs in cell-based systems, including known peptide solubility issues. Additionally, TX-TL reactions are microscale and compatible with automation5. TX-TL has been used to create new RiPPs with improved biological activity2,6.Overall, our project will address the following question: can we use TX-TL to discover novel antifungal RiPPs that inhibit plant pathogens? The specific aims and objectives (1-4) of this proposal are:Objective 1 - Produce a model antibacterial RiPP using TX-TL Objective 2 - Develop an automated RiPP screening platform at Syngenta Objective 3 - Characterise new antifungal RiPPs Objective 4 - Automate screening of antifungal RiPP Biosynthetic Gene Clusters at SyngentaProject impact: This project will (i.) provide a tool to make different RiPP variants for bioactivity screening; (ii.) characterise novel enzymes and secondary metabolites from RiPP biosynthesis; (iii.) develop an automated platform for discovery of antifungal RiPP natural products as a biological control measure for emerging crop pathogens.

迄今为止，仅鉴定出一种抗真菌RiPP（例如，pinensin），这是发现新RIPP的重要机会领域。作为一项使能技术，我们将使用由摩尔小组开发的无细胞转录-翻译（TX-TL）系统3，4。TX-TL系统需要DNA、细胞提取物和能量解决方案，以在数分钟至数小时内制造重组肽/蛋白质，并达到毫克规模。这种生产方法还克服了在基于细胞的系统中工程化RIPP的关键挑战和时间尺度问题，包括已知的肽溶解度问题。此外，TX-TL反应是微型的，与自动化兼容。TX-TL已被用于创建新的RIPPs与改善的生物活性2，6.总体而言，我们的项目将解决以下问题：我们可以使用TX-TL发现新的抗真菌RIPPs，抑制植物病原体？本提案的具体目的和目标（1-4）是：目标1 -使用TX-TL生产模型抗菌RiPP目标2 -在先正达开发自动化RiPP筛选平台目标3 -表征新的抗真菌RiPP目标4 -在先正达自动化筛选抗真菌RiPP生物合成基因簇项目影响：本项目将（i.）提供制备用于生物活性筛选的不同RiPP变体的工具;（ii.）从RiPP生物合成中获得新的酶和次级代谢物;（iii.）开发一个自动化平台，用于发现抗真菌RiPP天然产物，作为新出现的作物病原体的生物控制措施。