NADPH regeneration for efficient biosynthesis of indigo by flavin-containing monooxygenase and formate dehydrogenase.

Indigo is an important blue pigment widely used in textile, food, and medicine industries. Biological production of indigo attracts increasing attention recently. Cell factory production of indigo encounters the problem of the toxicity of the precursor indole. Enzymatic production is the alternative biological approach, however, NADPH regeneration should be solved. In this study, flavin-containing monooxygenase from Methylophaga aminisulfidivorans was used for enzymatic production of indigo from indole and formate dehydrogenase from Pseudomonas sp. 101 was co-expressed for NADPH regeneration. Indigo production was enhanced by combination of molecular modification, promoter engineering, and translation initiation region engineering. Finally, 0.183 g/L of indigo was produced from 0.5 g/L of indole and 0.5 mM of sodium formate, with the conversion ratio of 32.5 %. This study demonstrates a feasible and effective strategy for enzymatic production of indigo.