Enhancing indican production in Escherichia coli through a multi-faceted experimental strategy

Indican, abundant in plant stems and leaves, serves as a precursor for indigo and indirubin, possessing application potential in many industries. In this study, a multi-faceted experimental strategy for producing indican utilizing tryptophan as the starting material in E. coli was established. This system incorporated the synergistic action of TnaA, MeFMO, and PtUGT2^M91A/S145A, a mutant with a 20 % increase in activity of glycosyltransferase PtUGT2, all co-expressed within the bacterial host. Both a UDP-glucose recycling system and an NADPH regeneration system were incorporated for donor regeneration. Finally, a co-culture system aimed at reducing metabolic burdens was designed. After optimization, a maximum yield of 1043 mg/L was achieved, with a molar conversion rate of 48.1 %, representing an almost 14-fold improvement over the performance of a single strain. This study offers a plant-independent synthesis process for indican, paving a new path for year-round indican production.