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重新引导自然和工程代谢途径（即细胞中化学反应的途径），以生产所需的化学物质。重定向微生物代谢的一种方法是操纵代谢途径，使所需的代谢物量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以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，用于检测微生物中信号分子的积累，以及检测生长培养基中营养物质的消耗。当信号分子或营养物质达到目标浓度时，特定代谢途径的表达将被适当改变，以增加所需化学物质的产生。该工具包将广泛提供给合成生物学和代谢工程研究人员。通过跨学科的研究经验，为年轻的职业科学家（本科生和研究生）提供劳动力培训和发展。在适当的时间和适当的量来控制代谢物的通量以优化微生物合成化学品的产量的策略一直是人们非常感兴趣的。为了实现高产品产量，需要平衡增加细胞生物量的需要与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物流向产品合成，而不需要任何用户提供的输入。本项目旨在开发和分析利用自主代谢物阀，在适当的时间通过异源途径控制代谢物的适当通量。与其他依赖外源性修正来控制代谢物通量的系统相反，该系统将依赖于微生物合成信号的细胞内积累或生长培养基中特定营养物质的消耗。当信号分子或营养物质达到目标浓度时，基因表达将发生改变，从而增加代谢产物合成所需产物的通量。本研究的目标是：1)开发和分析基于群体感应的代谢物阀；2)基于天然磷酸盐饥饿反应开发和分析代谢物阀；3)将群体感应和磷酸盐饥饿阀集成到一个菌株中，以评估系统兼容性。虽然将研究一种特定的代谢途径来研究阀门性能，作为概念的证明，但也设想这些设备将用于更一般的生理研究，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的系统和合成生物学项目以及化学、生物工程、环境和运输系统部的生物技术和生化工程项目共同资助。

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.