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

这个小型企业技术转移（STTR）项目的更广泛的影响/商业潜力是开发一个工程的非模型酵母平台，用于经济和可持续地生产天然香气化合物。高质量的芳香化合物具有多种用途，从香精香料到化妆品和药品。随着消费者对食品和个人护理用品中化学合成成分的更多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来源，并将减少对植物来源的需求。这项工作将通过13c代谢物示踪、代谢通量分析、发酵优化和动力学建模来推进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，并将促进开发可转让技术，以廉价生产广泛用于人类健康和保健的各种天然产品。本STTR一期项目拟将合成生物学和生物工艺工程相结合，优化天然芳香化合物从头生产的微生物菌株。由于植物携带的芳香化合物含量极低，从植物中提取的商业提取物在经济上是不可行的。为了克服这些挑战，该项目将开发一种新的发酵工艺来生产这些具有高产品特异性和产量的化合物。具体来说，将通过PUSH（增加前体乙酰辅酶a的供应），PULL（过度表达关键生物合成途径基因），POWER（增强ATP和NADPH的产生）策略来设计非模型酵母菌株。此外，13c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的供应/消耗途径，并确定代谢负担或瓶颈因素。同时，将建立新的同步发酵和产品回收，以提高生产效率。fluxome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、自然的生产方案，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.