Production of plant-specific flavones baicalein and scutellarein in an engineered E. coli from available phenylalanine and tyrosine

Baicalein and scutellarein are bioactive flavones found in the medicinal plant Scutellaria baicalensis Georgi, used in traditional Chinese medicine. Extensive previous work has demonstrated the broad biological activity of these flavonoids, such as antifibrotic, antiviral and anticancer properties. However, their supply from plant material is insufficient to meet demand. Here, to provide an alternative production source and increase production levels of these flavones, we engineered an artificial pathway in an Escherichia coli cell factory for the first time. By first reconstructing the plant flavonoid biosynthetic pathway genes from five different species: phenylalanine ammonia lyase from Rhodotorula toruloides (PAL), 4-coumarate-coenzyme A ligase from Petroselinum crispum (4CL), chalcone synthase from Petunia hybrida (CHS), chalcone isomerase from Medicago sativa (CHI) and an oxidoreductase flavone synthase I from P. crispum (FNSI), production of the intermediates chrysin and apigenin was achieved by feeding phenylalanine and tyrosine as precursors. By comparative analysis of various versions of P450s, a construction expressing 2B1 incorporated with a 22-aa N-terminal truncated flavone C-6 hydroxylase from S. baicalensis (F6H) and partner P450 reductase from Arabidopsis thaliana (AtCPR) was found most effective for production of both baicalein (8.5 mg/L) and scutellarein (47.1 mg/L) upon supplementation with 0.5 g/L phenylalanine and tyrosine in 48 h of fermentation. Finally, optimization of malonyl-CoA availability further increased the production of baicalein to 23.6 mg/L and scutellarein to 106.5 mg/L in a flask culture. This report presents a significant advancement of flavone synthetic production and provides foundation for production of other flavones in microbial hosts.