title: C-Glycosyltransferases catalyzing the formation of di-C-glucosyl flavonoids in citrus plants.
abstract: Citrus plants accumulate many kinds of flavonoids, including di-C-glucosyl flavonoids, which have attracted considerable attention due to their health benefits. However, the biosynthesis of di-C-glucosyl flavonoids has not been elucidated at the molecular level. Here, we identified the C-glycosyltransferases (CGTs) FcCGT (UGT708G1) and CuCGT (UGT708G2) as the primary enzymes involved in the biosynthesis of di-C-glucosyl flavonoids in the citrus plants kumquat (Fortunella crassifolia) and satsuma mandarin (Citrus unshiu), respectively. The amino acid sequences of these CGTs were 98% identical, indicating that CGT genes are highly conserved in the citrus family. The recombinant enzymes FcCGT and CuCGT utilized 2-hydroxyflavanones, dihydrochalcone, and their mono-C-glucosides as sugar acceptors and produced corresponding di-C-glucosides. The Kmand kcatvalues of FcCGT toward phloretin were <0.5 μm and 12.0 sec-1, and those toward nothofagin (3'-C-glucosylphloretin) were 14.4 μm and 5.3 sec-1, respectively; these values are comparable with those of other glycosyltransferases reported to date. Transcripts of both CGT genes were found to concentrate in various plant organs, and particularly in leaves. Our results suggest that di-C-glucosyl flavonoid biosynthesis proceeds via a single enzyme using either 2-hydroxyflavanones or phloretin as a substrate in citrus plants. In addition, Escherichia coli cells expressing CGT genes were found to be capable of producing di-C-glucosyl flavonoids, which is promising for commercial production of these valuable compounds.
authors.affiliation: Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University, 3-15-1 Tokida, Ueda, 386-8567, Japan.
authors.forename: Takamitsu
authors.initials: T
authors.lastname: Ito
authors.affiliation: Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University, 3-15-1 Tokida, Ueda, 386-8567, Japan.
authors.forename: Shunsuke
authors.initials: S
authors.lastname: Fujimoto
authors.affiliation: Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University, 3-15-1 Tokida, Ueda, 386-8567, Japan.
authors.forename: Fumiaki
authors.initials: F
authors.lastname: Suito
authors.affiliation: Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University, 3-15-1 Tokida, Ueda, 386-8567, Japan.
authors.forename: Makoto
authors.initials: M
authors.lastname: Shimosaka
authors.affiliation: Department of Applied Biology, Faculty of Textile Science and Technology, Shinshu University, 3-15-1 Tokida, Ueda, 386-8567, Japan.
authors.forename: Goro
authors.initials: G
authors.lastname: Taguchi
mesh_terms: D002384:Catalysis; D047188:Chalcones / Q000378:metabolism; D002957:Citrus / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D044950:Flavanones / Q000378:metabolism; D005419:Flavonoids / Q000378:metabolism*; D005638:Fruit / Q000378:metabolism; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism*; D010693:Phloretin / Q000378:metabolism; D010802:Phylogeny; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: C520534:2'-hydroxyflavanone; C000625625:2-hydroxynaringenin; D047188:Chalcones; D044950:Flavanones; D005419:Flavonoids; D010940:Plant Proteins; D011994:Recombinant Proteins; C528728:nothofagin; D016695:Glycosyltransferases; D010693:Phloretin
keywords: Citrus unshiu; Fortunella crassifolia; di-C-glucosylation; flavonoid C-glycosyltransferase; phloretin di-C-glucoside; vicenin-2