Flavonol glycosides are health-promoting phytochemicals important for human nutrition and plant defense against environmental stresses. Glycosylation modification greatly enriches the diversity of flavonols. Morella rubra, a member of the Myricaceae, contains high amounts of myricetin 3-O-rhamnoside (M3Rha), quercetin 3-O-rhamnoside (Q3Rha), and quercetin 3-O-galactoside (Q3Gal). In the present study, MrUGT78R1 and MrUGT78R2 were identified as two functional UDP-rhamnosyltransferases, while MrUGT78W1 was identified as a UDP-galactosyltransferase. Site-directed mutagenesis identified Pro143 and Asn386 as important residues for rhamnosyl transfer activity of MrUGT78R1, while the two corresponding positions in MrUGT78W1 (i.e. Ser147 and Asn370) also play important roles in galactosyl transfer activity. Transient expression data for these three MrUGTs in Nicotiana benthamiana tested the function of MrUGT78R1 and MrUGT78R2 as rhamnosyltransferases and MrUGT78W1 as a galactosyltransferase in glycosylation of flavonols. This work enriches knowledge of the diversity of UDP-rhamnosyltransferase in planta and identifies two amino acid positions important for both rhamnosyltransferase and galactosyltransferase.

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