Identification of a Flavanone 2-Hydroxylase Involved in Flavone -Glycoside Biosynthesis from .

Tea contains a variety of flavone -glycosides, which are important compounds that distinguish tea cultivars and tea categories. However, the biosynthesis pathway of flavone -glycosides in tea plant remains unknown, and the key enzymes involved have not been characterized. In this study, a liquid chromatography-mass spectrometry method to determine 9 flavone -glycosides was developed, and the accumulation patterns of 9 flavone -glycosides in tea plants were examined first. Then, an entry enzyme F2H for flavone -glycoside biosynthesis was identified, which had four cytochrome P450-specific conserved motifs and was targeted to the endoplasmic reticulum. Correlation analysis indicated that the expression level of was positively correlated with all contents of 9 flavone -glycosides. The recombinant F2H could convert flavanone (naringenin) into the corresponding 2-hydroxyflavonone (2-hydroxynaringenin), rather than into flavone (apigenin). Heterologous coexpression of F2H and CGT1 in yeast revealed that the substrate naringenin could be enzymatically converted to flavone mono--glycosides vitexin and isovitexin under the catalytic control of F2H and CGT1 following dehydration. Gene-specific antisense oligonucleotide analysis suggested that suppressing significantly reduced the levels of 9 flavone -glycosides. Together, F2H is the first key enzyme that generates flavone -glycosides through the 2-hydroxyflavanone biosynthesis pathway in tea plants.