Functional Characterization of a Highly Efficient UDP-Glucosyltransferase CitUGT72AZ4 Involved in the Biosynthesis of Flavonoid Glycosides in Citrus

Citrus is an important dietary source of flavonoid glycosides, and UDP-glycosyltransferases (UGTs) are the key enzymes responsible for their glycosylation. In this study, a genome-wide analysis of the CitUGT gene family was conducted to identify CitUGTs that contribute to flavonoid 4′-O-glucosides biosynthesis. Our analysis identified 136 CitUGTs in the Citrus clementina genome, classifying them into 18 phylogenetic groups (A–R) and 25 families. This classification was strongly supported by consistent gene structures and motif patterns. Moreover, we identified a CitUGT gene (Ciclev10025462m, designated CitUGT72AZ4) that encodes flavonoid 4′-O-glucosyltransferase for the first time in citrus. This enzyme preferentially glycosylated the 4′-OH group of multiple flavonoids, exhibiting higher catalytic efficiency toward quercetin and three flavones in vitro. Virus-induced gene silencing of CitUGT72AZ4 significantly decreased the accumulation of flavonoid 4′-O-glucosides. These results indicated that CitUGT72AZ4 participated in the biosynthesis of flavonoid 4′-O-glucoside in citrus. Overall, our findings provide valuable insights into the CitUGT gene family and contribute to its functional characterization.