EAGER: SusChEM: Developing Temporal Compartmentalization Tools in Yeast to Enhance the Bioproduction of Phytosterols

植物类固醇是一种复杂的天然分子，具有广泛的制药和农业应用；然而，它们通常在植物中含量很少，而且很难用化学方法合成。这个研究项目研究了改造酵母产生这些分子的方法。为了实现这一目标，有必要引入酵母对特定蛋白质进行靶向降解的能力。该项目的研究人员正在采用先进的生化方法来实现酵母的这种变化。这个研究项目包括为高中学生和当地社区大学的本科生举办的具体活动，这将有助于为生物技术产业培养训练有素的技术劳动力。本研究为酵母中复杂的异源植物通路的可编程时间调控机制的开发提供了基础。这种机制采用酵母内源性时间区隔化策略和与内质网（ER）相关的诱导蛋白降解系统。将复杂的植物途径引入包括酵母在内的微生物宿主是一项极具挑战性的工作，主要原因是：1)大量植物酶的表达带来的高负担；2)许多异源酶步骤之间缺乏有效的协调。需要一个植物生物合成平台来整合和验证所提出的时间调节策略，以及靶向蛋白质降解（TPD）方法。植物类固醇激素brassinosteroids （BRs）的生物合成途径正在酵母中得到验证和建立，这将使下游建立所提出的时间调控策略成为可能。此外，正在评估在拟议的时间调节策略中引入TPD的可行性。

Plant steroids are complex, naturally occurring molecules that have a wide variety of pharmaceutical and agricultural applications; however, they are usually found in quite small amounts in plants, and they are difficult to synthesize chemically. This research project investigates ways to engineer yeasts to produce these molecules. To accomplish this goal, it is necessary to introduce into the yeast the ability to perform the targeted degradation of specific proteins. The researchers on this project are employing advanced biochemical methods in order to realize this alteration in the yeast. This research project include specific activities for high school students and for undergraduate students from a local community college, which will help in developing a highly trained technical workforce for the biotechnology industry. This research project provides a foundation for the development of programmable temporal regulation machinery for complex heterologous plant pathways in yeast. Such machinery employs yeast endogenous temporal compartmentalization strategies and an inducible protein degradation system associated with the endoplasmic reticulum (ER). Introducing complex plant pathways into microbial hosts including yeast has been highly challenging, mainly due to 1) the high burden from the expression of large numbers of plant enzymes; and 2) the lack of efficient coordination among the many heterologous enzymatic steps. A plant biosynthetic platform is required to incorporate and validate the proposed temporal regulation strategy, as is a targeted protein degradation (TPD) approach. The biosynthetic pathway of the plant steroid hormone brassinosteroids (BRs) is being verified and established in yeast, which will enable the downstream establishment of the proposed temporal regulation strategies. In addition, the feasibility of introducing TPD in the proposed temporal regulation strategy is being evaluated.