STTR Phase I: De Novo Production of Aroma Compounds by Nonconventional Recombinant Yeast

这个小企业技术转让（STTR）项目的更广泛的影响/商业潜力是开发一个工程化的非模型酵母平台，用于经济和可持续地生产天然芳香化合物。高品质的芳香化合物具有多种应用，从香料和芳香剂到化妆品和药物。随着消费者对食品和个人护理产品中化学合成成分的天然替代品的强烈需求，通过发酵生产这些成分将为供应链增加可再生资源，并将减少对植物来源的需求。这项工作将推进通过13 C-代谢物追踪，代谢通量分析，发酵优化和动力学建模创建新型微生物细胞工厂的方法和知识。这项研究的经验教训将为克服生物合成障碍和优化生物过程操作提供一般指导方针，并将促进可转让技术的发展，以廉价生产广泛用于人类健康和保健的各种天然产品。 该STTR第一阶段项目建议整合合成生物学和生物工艺工程，以优化微生物菌株，用于从头生产天然芳香化合物。由于植物携带极少量的芳香化合物，因此来自植物的商业提取物在经济上不可行。为了克服这些挑战，该项目将开发一种新的发酵工艺，用于生产具有高产品特异性和产率的这些化合物。具体而言，非模型酵母菌株将通过PUSH（增加前体乙酰辅酶A的供应）、PALG（过表达关键生物合成途径基因）、POWER（增强ATP和NADPH生成）策略进行工程化。此外，13 C-代谢通量分析将用于严格测量体内酶功能，描绘乙酰辅酶A供应/消耗途径，并确定代谢负担或瓶颈因素。同时，将建立新的同时发酵和产品回收，以提高生产率。通量体信息和发酵动力学将为合理的遗传修饰和生物过程优化提供指导。最终，这种环保工艺将提供一种新的、可靠的和自然的制造方案，以满足消费者对更健康产品日益增长的需求。

The broader impact/commercial potential of this Small Business Technology Transfer (STTR) project is to develop an engineered nonmodel yeast platform for the economical and sustainable production of natural aroma compounds. High quality aroma compounds have a variety of applications, ranging from flavor and fragrance to cosmetics and pharmaceuticals. With the strong consumer demand for more natural alternatives to chemically synthesized ingredients in both food and personal care items, the production of these ingredients by fermentation will add a renewable source to the supply chain and will reduce demand on botanical sources. This work will advance methodologies and knowledge for creating novel microbial cell factories via 13C-metabolite tracing, metabolic flux analysis, fermentation optimization, and kinetic modeling. The lessons from this research will offer general guidelines for overcoming biosynthesis hurdles and optimizing bioprocess operations, and will facilitate the development of transferable technologies for inexpensive production of diverse natural products that are broadly used for human health and wellness. This STTR Phase I project proposes to integrate synthetic biology and bioprocess engineering to optimize the microbial strain for the de novo production of natural aroma compounds. Because plants carry aroma compounds in extremely low amounts, commercial extracts from plants are not economically viable. To overcome these challenges, this project will develop a novel fermentation process for production of these compounds with high product specificity and yields. Specifically, a nonmodel yeast strain will be engineered via PUSH (increase the supply of the precursor acetyl-CoA), PULL (overexpress key biosynthetic pathway genes), POWER (enhance ATP and NADPH generation) strategies. Further, 13C-metabolic flux analysis will be used to rigorously measure in vivo enzyme functions, to delineate acetyl-CoA supply/consumption pathways, and to identify metabolic burdens or bottleneck factors. Meanwhile, novel simultaneous fermentation and product recovery will be built to improve productivity. The fluxome information and fermentation kinetics will offer guidelines for rational genetic modifications and bioprocess optimizations. Ultimately, this environmentally friendly process would provide a new, reliable and natural manufacture scheme to meet the growing demand of consumers for healthier products.