Biosynthesis of rare 20(R)-protopanaxadiol/protopanaxatriol type ginsenosides through Escherichia coli engineered with uridine diphosphate glycosyltransferase genes

Abstract

BackgroundGinsenosides are known as the principal pharmacological active constituents in Panax medicinal plants such as Asian ginseng, American ginseng, and Notoginseng. Some ginsenosides, especially the 20(R) isomers, are found in trace amounts in natural sources and are difficult to chemically synthesize. The present study provides an approach to produce such trace ginsenosides applying biotransformation through Escherichia coli modified with relevant genes.MethodsSeven uridine diphosphate glycosyltransferase (UGT) genes originating from Panax notoginseng, Medicago sativa, and Bacillus subtilis were synthesized or cloned and constructed into pETM6, an ePathBrick vector, which were then introduced into E. coli BL21star (DE3) separately. 20(R)-Protopanaxadiol (PPD), 20(R)-protopanaxatriol (PPT), and 20(R)-type ginsenosides were used as substrates for biotransformation with recombinant E. coli modified with those UGT genes.ResultsE. coli engineered with GT95syn selectively transfers a glucose moiety to the C20 hydroxyl of 20(R)-PPD and 20(R)-PPT to produce 20(R)-CK and 20(R)-F1, respectively. GTK1- and GTC1-modified E. coli glycosylated the C3—OH of 20(R)-PPD to form 20(R)-Rh2. Moreover, E. coli containing p2GT95synK1, a recreated two-step glycosylation pathway via the ePathBrich, implemented the successive glycosylation at C20—OH and C3—OH of 20(R)-PPD and yielded 20(R)-F2 in the biotransformation broth.ConclusionThis study demonstrates that rare 20(R)-ginsenosides can be produced through E. coli engineered with UTG genes.

Efficacy of Panax ginseng Meyer Herbal Preparation HRG80 in Preventing and Mitigating Stress-Induced Failure of Cognitive Functions in Healthy Subjects: A Pilot, Randomized, Double-Blind, Placebo-Controlled Crossover Trial

“…To date, 112 saponins have been reported as components of P. ginseng, also known as Korean ginseng; >80 of them are isolated from raw or processed ginseng, and the others are acid/base hydrolysates, semisynthetic saponins, or metabolites [38,39]. A total of 39 ginsenosides were identified and quantified in ginseng roots, leaves, stems, and berries, where the roots' major ginsenosides are M-Rb1 (0.47%), Rb1 (0.38%), Rf (0.46%), Ra2 (0.32%), Rg1 (0.31%), Re (0.3%), and Rg2 (0.17%), whereas the content of other minor ginsenosides is less than 0.1% [40].…”

“…Major ginsenosides Rg1, Re, Rb1, Rb2, Rc, Rd, and Rf have poor bioavailability [ 41 , 42 ], but can be metabolized by intestinal microflora (gut bacteria) or gastric juice [ 41 , 43 , 44 , 45 , 46 , 47 , 48 , 49 , 50 ] into more bioavailable and active [ 3 , 38 , 49 , 50 , 51 ], minor, so-called “rare” ginsenosides Rg2, Rg3, Rg6, Rh2, Rh3, Rh4, compound K, Rk1, and PPD [ 7 , 38 , 39 , 41 , 46 , 52 ], comprising 0.8% as determined by mass of the root powder ( Table 3 ). The content of more bioavailable rare ginsenosides in the P. ginseng HRG80 preparation, obtained by hydroponic cultivation in controlled stressful conditions, followed by traditional cooking of roots, was 7.8-fold higher ( Table 3 ), suggesting the superior pharmacological activity of the HRG80 preparation compared with traditionally harvested six-year-old red P. ginseng.…”

miR-452 Reverses Abnormal Glycosylation Modification of ERα and Estrogen Resistance in TNBC (Triple-Negative Breast Cancer) Through Targeting UGT1A1

“…Therefore, UGT1A1 may be a potential target for research on the pathogenesis of TNBC. Previous studies have shown that UGT1A1-catalyzed glucuronidation is related to the metabolism and detoxification process of estrogen ( 40 , 41 ). Recently, studies have shown that UGT is closely related to estrogen metabolism.…”

The chromosome-level reference genome assembly for Panax notoginseng and insights into ginsenoside biosynthesis

“…Third, 2,3-oxidosqualene undergoes cyclization and hydroxylation ( Han et al., 2011 , 2012 ) to form the core structures PPD and PPT ( Luo et al., 2011 ; Lu et al., 2018 ). Finally, the formation of various saponins is catalyzed by a number of glycosyltransferases (GTs) ( Yu et al., 2019 ). The genetic and functional diversity of GTs gives rise to a variety of structurally diverse saponins.…”