Functional identification and characterization of two flavonoid glycosyltransferases ZmUGT84A3 and ZmUGT84A4 from maize

Luteolin is an important secondary metabolite of maize, which can be glycosylated through uridine diphosphate glycosyltransferase (UGT). Extensive research on UGT has been centered in Arabidopsis thaliana, but little in maize. We cloned two maize glycosyltransferase genes, ZmUGT84A3 and ZmUGT84A4. In vitro experiments revealed that ZmUGT84A3 and ZmUGT84A4 glycosylated luteolin to both luteolin-7-O-glucoside and luteolin-4′,7-di-O-glucoside. Notably, both ZmUGT84A3 and ZmUGT84A4 catalyzed glycosylation of many flavonoids including apigenin, naringenin, eriodyctiol, kaempferol, and quercetin. Increased temperature enhanced activities of both ZmUGT84A3 and ZmUGT84A4, and elevated a production of luteolin-4′,7-di-O-glucoside. ZmUGT84A3 activity was optimal under acidic conditions but ZmUGT84A4 preferred alkaline environments. ZmUGT84A4 exhibited higher catalytic activities (1.3 to 2.6-fold) at various substrate concentrations than did ZmUGT84A3; the latter enzyme evidenced a unique activity pattern. Enzyme-catalyzed reaction processes of ZmUGT84A3 and ZmUGT84A4 in vitro for luteolin were depicted. This study unveiled two novel multifunctional glycosyltransferases identified in maize, which exhibited extensive substrate specificity, possessed multiple catalytic sites, and offered valuable insights for studying their enzymatic characteristics.