Development and Analysis of Autonomous Metabolite Valves

微生物将有机材料(原料)转化为大量化学产品的能力使它们非常适合作为化工厂。这项技术的进步需要将自然和工程的代谢途径(即细胞内的化学反应途径)重新定向到生产所需的化学物质。改变微生物代谢方向的一种方法是操纵途径，使所需数量的代谢物在适当的时间流向所需的途径。这类似于打开一个阀门，允许化学品在适当的时间和适当的数量流入工厂。在这项研究中，将开发一套生物工程工具，用于检测信号分子在微生物中的积累，并用于检测生长介质中营养物质的消耗。当达到信号分子或营养素的目标浓度时，特定代谢途径的表达将适当改变，以增加所需化学物质的产量。该工具包将广泛提供给合成生物学和代谢工程研究人员。将通过跨学科研究经验向年轻的职业科学家(本科生和研究生)提供劳动力培训和发展。人们非常有兴趣制定策略，在适当的时间和适当的数量控制代谢物的流量，以优化微生物合成的化学品的产量。要实现高产量，需要平衡增加细胞生物量的需要和将代谢物转向合成所需产品的需要。一个理想的系统将允许动态控制代谢产物流向产品合成，而不需要任何用户提供的输入。该项目旨在开发和分析自主代谢物阀门的使用，以在适当的时间通过异源途径控制适当的代谢物流量。与其他依赖外源改良剂控制代谢产物通量的系统不同，该系统将依赖于微生物合成信号在细胞内的积累或生长介质中特定营养物质的耗尽。在信号分子或营养素的目标浓度下，基因表达会发生变化，以增加代谢产物合成所需产品的流量。这项研究的目标是：1)开发和分析基于群体感应的代谢物阀；2)基于天然磷饥饿反应开发和分析代谢物阀；以及3)将群体感应和磷饥饿阀整合到单一菌株中，以评估系统的兼容性。虽然将研究特定代谢途径的瓣膜性能作为概念验证，但也可以预见这些设备将用于更一般的生理学研究，如代谢变化对细胞行为的影响。该奖项由分子和细胞生物科学部门的系统和合成生物学计划以及化学、生物工程、环境和运输系统部门的生物技术和生化工程计划联合资助。

The ability of microbes to convert organic materials (feedstock) into a vast array of chemical products makes them ideally suited as chemical factories. Advances in this technology require redirecting natural and engineered metabolic pathways (i.e. the pathways of chemical reactions in the cell) towards the production of the desired chemical. One means of redirecting microbial metabolism is to manipulate the pathways so that the desired amount of metabolites flows toward the desired pathway at the appropriate time. This is akin to opening a valve to permit the flow of chemicals into a factory at the appropriate time and in the appropriate amount. In this study a set of bioengineering tools will be developed for detecting accumulation in a microbe of a signalling molecule and for detecting the depletion of a nutrient in the growth medium. When the targeted concentrations of the signal molecule or nutrient are reached the expression of specific metabolic pathways will be altered appropriately to increase production of the desired chemical. This toolkit will be made broadly available to synthetic biology and metabolic engineering researchers. Workforce training and development will be provided to young career (undergraduate and graduate students) scientists through an interdisciplinary research experience. There has been great interest in developing strategies to control the flux of metabolites at the appropriate time and in the appropriate amounts to optimize yields of microbially synthesized chemicals. To achieve high product yields requires balancing the need for increasing cellular biomass with the diversion of metabolites towards the synthesis of the desired product. An ideal system would allow dynamic control of metabolite flow towards product synthesis without the need for any user-provided inputs. This project aims to develop and analyze the use of autonomous metabolite valves to control the appropriate flux of metabolites through a heterologous pathway at the appropriate time. In contrast to other systems that rely on exogenous amendments to control metabolite flux this system will rely on the intracellular accumulation of microbially synthesized signals or the depletion of a specific nutrient in the growth medium. At targeted concentrations of the signal molecule or nutrient, gene expression will altered to increase the flux of metabolites toward synthesis of the desired product. The objectives of this study will be to: 1) develop and analyze quorum sensing-based metabolite valves; 2) develop and analyze metabolite valves based on the natural phosphate starvation response; and 3) integrate the quorum sensing and phosphate starvation valves into a single strain to assess system compatibility. While a specific metabolic pathway will be investigated for valve performance as proof of concept, it is also envisioned that these devices will be useful for more general physiological studies such as the impact of metabolic shifts on cellular behavior.This award is funded jointly by the Systems and Synthetic Biology Program in the Division of Molecular and Cellular Biosciences and by the Biotechnology and Biochemical Engineering Program in the Division of Chemical, Bioengineering, Environmental and Transport Systems.