An Optimized Bistable Metabolic Switch To Decouple Phenotypic States during Anaerobic Fermentation

Metabolic engineers aim to genetically modify microorganisms to improve their ability to produce valuable compounds. Despite the prevalence of growth-coupled production processes, these strategies can significantly limit production rates. Instead, rates can be improved by decoupling and optimizing growth and production independently, and operating with a growth stage followed by a production stage. Here, we implement a bistable transcriptional controller to decouple and switch between these two states. We optimize the controller in anaerobic conditions, typical of industrial fermentations, to ensure stability and tight expression control, while improving switching dynamics. The stability of this controller can be maintained through a simulated seed train scale-up from 5 mL to 500 000 L, indicating industrial feasibility. Finally, we demonstrate a two-stage production process using our optimal construct to improve the instantaneous rate of lactate production by over 50%, motivating the use of these systems in broad metabolic engineering applications.