Functional Analysis of a UDP-Glucosyltransferase Gene Contributing to Biosynthesis of the Flavonol Tri-glycoside in Tea Plants

Flavonol glycosides have many prominent benefits to human health and significant contributions to the growth and development of tea plant as well as the color and taste of tea infusion. In this study, a gene has been isolated from tea plant and encodes a 52.2 kDa protein located on the plasma membrane and in cytoplasm with an activity of flavonol glycosyltransferase(CsFGT). The prokaryotically expressed recombinant CsFGT (rCsFGT) exhibits its main glucosyl-transfer activity towards rutin to produce quercetin 3-O-β-D-glucopyranosyl-(1→3)-α-L-rhamnopyranosy-(1→6)-β-D-glucopyranoside(Q-g-r-g), and shows a minor galactosyl-transfer activity towards delphinidin to produce delphinidin 3-O galactoside. The maximum activity of the rCsFGT has been observed at 30 °C and pH 8.0. Main function of the rCsFGT seems to catalyze the biosynthesis of Q-g-r-g rather than delphinidin 3-O galactoside since its affinity and catalytic efficiency are much higher towards rutin than towards delphinidin. The molecular docking and site-directed mutation reveal amino acid residues G290, E292, R319 and Q352 play important roles in catalytic specificity of the CsFGT. The Q-g-r-g content in leaves of different tea cultivars is significantly correlated with the CsFGT expression level. Injection of antisense oligodeoxyribonucleotides remarkably downregulates the endogenous CsFGT expression and consequently reduces the Q-g-r-g content significantly. These findings will help elucidate the differential accumulation mechanism of flavonol glycosides in different tea germplasms.