Photocontrol of Itaconic Acid Synthesis in Escherichia coli .

Metabolic engineering aims to control cellular metabolic flow and maximize the production of a product of interest. Photocontrol of the activities of proteins is an effective method for accurately regulating metabolic pathways. In this study, we inserted the photosensor light-oxygen-voltage-sensing domain 2 of Avena sativa (AsLOV2) into selected sites of isocitrate dehydrogenase (IDH), the key enzyme in the competitive pathway of itaconic acid (ITA) synthesis, to construct photoswitchable IDH-AsLOV2 (ILOVs). These engineered light-sensitive proteins were used to regulate the metabolic flux of the tricarboxylic acid (TCA) cycle in Escherichia coli to improve ITA production. The engineered fusion proteins ILOV2, ILOV3, ILOV6, and ILOV7 exhibited effective reversibility under the oscillation of darkness and blue light illumination in vitro . The efficacies of the intracellular photoswitches were evaluated, and an optimal photocontrol strategy was established in vivo . The ITA titer was significantly enhanced to 3.30 g/L for strain ITAΔ43, which displayed superior photoswitchable potency for ITA production compared with the strains that completely deleted the icd gene. The photocontrol strategy developed here can be extended for process optimization and titer improvement of other high-value bioengineering chemicals.