title: Molecular cloning and biochemical characterization of the UDP-glucose: flavonoid 3-O-glucosyltransferase from Concord grape (Vitis labrusca).
abstract: Glucosylation of anthocyanidin substrates at the 3-O-position is crucial for the red pigmentation of grape berries and wine. The gene that encodes the enzyme involved in this reaction has been cloned from Vitis labrusca cv. Concord, heterologously expressed, and the recombinant enzyme (rVL3GT) was characterized. VL3GT has 96% amino acid sequence identity with Vitis vinifera VV3GT and groups phylogenetically with several other flavonoid 3-O-glycosyltransferases. In vitro substrate specificity studies and kinetic analyses of rVL3GT indicate that this enzyme preferentially glucosylates cyanidin as compared with quercetin. Crude protein extracts from several Concord grape tissues were assayed for glucosyltransferase activity with cyanidin and quercetin as acceptor substrates. A comparison of the VL3GT activities toward with these substrates showed that the 3GT enzyme activity is consistent with the expression of VL3GT in these tissues and is coincident with the biosynthesis of anthocyanins in both location and developmental stages. Enzyme activities in grape mesocarp, pre-veraison exocarp, leaf, flower bud, and flower tissues glucosylated quercetin but not cyanidin at high rates, suggesting the presence of additional enzymes which are able to glucosylate the 3-O-position of flavonols with higher specificity than anthocyanidins.
authors.affiliation: Department of Biological Sciences, Brock University, St. Catharines, ON, Canada L2S 3A1.
authors.forename: Dawn
authors.initials: D
authors.lastname: Hall
authors.forename: Xiao Xin
authors.initials: XX
authors.lastname: Yuan
authors.forename: Jun
authors.initials: J
authors.lastname: Murata
authors.forename: Vincenzo
authors.initials: V
authors.lastname: De Luca
mesh_terms: D000595:Amino Acid Sequence; D000872:Anthocyanins / Q000378:metabolism*; D003001:Cloning, Molecular; D005638:Fruit / Q000201:enzymology; D015870:Gene Expression; D017343:Genes, Plant; D005964:Glucosyltransferases / Q000235:genetics / Q000378:metabolism*; D006031:Glycosylation; D008969:Molecular Sequence Data; D010802:Phylogeny; D010860:Pigments, Biological; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D018514:Plant Structures; D011794:Quercetin / Q000378:metabolism*; D011994:Recombinant Proteins; D013045:Species Specificity; D013379:Substrate Specificity; D014532:Uridine Diphosphate Glucose / Q000378:metabolism*; D027843:Vitis / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D014920:Wine
chemical_list: D000872:Anthocyanins; D010860:Pigments, Biological; D010940:Plant Proteins; D011994:Recombinant Proteins; C017154:cyanidin; D011794:Quercetin; D005964:Glucosyltransferases; C023249:flavonol 3-O-glucosyltransferase; D014532:Uridine Diphosphate Glucose