Model-based design of a sensor-circuit for autonomous control of biosynthesis pathways

Micro-organisms have been engineered to manufacture diverse chemical products through the heterologous expression of biosynthesis pathways. However, the productivities of most pathways are anti-coupled with the host’s growth rate, due to competition for metabolic precursors. Here, we  engineered a rationally designed sensor-circuit device that autonomously regulates the expression of a metabolic pathway in response to its load, greatly increasing the organism’s growth rate and the pathway’s productivity. Specifically, we utilized a newly discovered intracellular stress sensor that detects imbalances in glycolytic flux, and activates proteolytic degradation of a targeted regulator. We show that this sensor increased the degradation rate of a targeted protein by 5-fold whenever a heterologous pathway over-consumes glycolytic metabolites. We then combined this sensor with a 2-regulator genetic circuit to create a modular sensor-circuit controller. We applied the sensor-circuit device to regulate the expression of a 3-enzyme terpenoid biosynthesis pathway and characterized its dynamic response over a 100 hour time period. We found that the sensor-circuit device’s control dynamics exhibited classic overshoot behavior; pathway expression increased 14-fold before settling at an optimal set-point. By perturbing the strength of the negative feedback loop, we showed that the sensor-circuit device was responsible for increasing the host’s terpenoid productivity by over 75-fold. Our load-sensing sensor-circuit devices can be re-used with a variety of biosynthesis pathways, and can provide dynamic, autonomous control to further increase their productivities during long-time fermentations.