Characterization of UGT716A1 as a Multi-substrate UDP:Flavonoid Glucosyltransferase Gene in Ginkgo biloba

Abstract

Ginkgo biloba L., a “living fossil” and medicinal plant, is a well-known rich source of bioactive flavonoids. The molecular mechanism underlying the biosynthesis of flavonoid glucosides, the predominant flavonoids in G. biloba, remains unclear. To better understand flavonoid glucosylation in G. biloba, we generated a transcriptomic dataset of G. biloba leaf tissue by high-throughput RNA sequencing. We identified 25 putative UDP-glycosyltransferase (UGT) unigenes that are potentially involved in the flavonoid glycosylation. Among them, we successfully isolated and expressed eight UGT genes in Escherichia coli, and found that recombinant UGT716A1 protein was active toward broad range of flavonoid/phenylpropanoid substrates. In particular, we discovered the first recombinant UGT protein, UGT716A1 from G. biloba, possessing unique activity toward flavanol gallates that have been extensively documented to have significant bioactivity relating to human health. UGT716A1 expression level paralleled the flavonoid distribution pattern in G. biloba. Ectopic over-expression of UGT716A1 in Arabidopsis thaliana led to increased accumulation of several flavonol glucosides. Identification and comparison of the in vitro enzymatic activity of UGT716A1 homologs revealed a UGT from the primitive land species Physcomitrella patens also showed broader substrate spectrum than those from higher plants A. thaliana, Vitis vinifera, and Medicago truncatula. The characterization of UGT716A1 from G. biloba bridges a gap in the evolutionary history of UGTs in gymnosperms. We also discuss the implication of UGT716A1 for biosynthesis, evolution, and bioengineering of diverse glucosylated flavonoids.

Characterization of a heat responsive UDP: Flavonoid glucosyltransferase gene in tea plant (Camellia sinensis)

“…In the initial study of rUGT73A17 protein, we found it was capable of glucosylating epicatechin gallate as GbUGT716A1 [ 29 ], which is distinct from the other characterized UGTs. Therefore, we further identified the other flavonoid substrates of the recombinant UGT73A17 protein.…”

“…Instead, UGT73A17 is highly expressed in mature leaves, which is possibly responsible for the production of rutin. Although CsUGT73A17 formed 7- O -glucosides as major products and 3- O -glucosides as minor products, it is still possible that it is responsible for the biosynthesis of rutin, since in vitro activity doesn't always reflect in vivo activity as for UGT in several previous studies [ 29 , 31 , 36 , 37 ]. Taken together, these CsUGTs are most likely responsible for the accumulation of different flavonoid glucosides in different organs of tea plant.…”

“…In comparison, CsUGT73A17 has a broader substrate profile than the other UGTs that were identified in tea plants so far. In particular, CsUGT73A17 also exhibited activity toward epicatechin gallate, which is the same as UGT716A1 from Ginkgo biloba that was active toward catechin galloylates [ 29 ], although the identity between them is only 30%. The glycosylated product of epigallocatechin gallate could increase water solubility, rapid metabolism and ready degradation in aqueous solutions, with significant bioactivity relating to human health [ 32 – 34 ].…”

Phylogenomic analysis of UDP‐dependent glycosyltransferases provides insights into the evolutionary landscape of glycosylation in plant metabolism

“…Association of sequential enzymes and structural proteins into metabolons to facilitate metabolic channeling has been reported in plant core and specialized metabolism (Knudsen et al ., ). Group Q UGTs have shown broad activities towards phenolic substrates, and a strong preference of flavones and flavonols in vitro ( Ginkgo biloba , P. granatum and Pilosella officinarum ; Witte et al ., ; Su et al ., ; Wilson et al ., ). The physical interactions between group Q UGTs and other proteins could be explored in the future to determine the function of the surface regions of these UGTs in forming metabolons and producing flavonoid glycosides.…”

N-glucosyltransferase GbNGT1 fromGinkgocomplement auxin metabolic pathway

“…Ginkgo flavonoids and ginkgolides had been widely used as important drug and dietary supplements in the world (Su et al, 2017). IAA-Asp-N-glucoside was mentioned as the main pharmacological compound in ginkgo for inhibiting cough, antiasthmatic and eliminating phlegm (Liu et al, 2018);furthermore, there were considerable pharmacological studies on IAA derivates or indole alkaloids, for instance, IAA induced cell death in combination with UV-B irradiation by increasing apoptosis in PC-3 prostate cancer cells (Kim et al, 2010); indole-N-glucosides have the potential to serve as a novel SGLT2-selective inhibitors (Sodium-Glucose cotransporter) to cure type Ⅱ diabetes (Nomura et al, 2013).…”