A cellulose synthase-derived enzyme catalyses 3-O-glucuronosylation in saponin biosynthesis

Susumu Hiraga,Kazuki Saito,Toshiya Muranaka,Masao Ishimoto,Yuhta Nomura,Soo Yeon Chung,Hikaru Seki,Yukiko Fujisawa,Yoshikazu Shimoda

doi:10.1038/s41467-020-19399-0

Abstract

Triterpenoid saponins are specialised metabolites distributed widely in the plant kingdom that consist of one or more sugar moieties attached to triterpenoid aglycones. Despite the widely accepted view that glycosylation is catalysed by UDP-dependent glycosyltransferase (UGT), the UGT which catalyses the transfer of the conserved glucuronic acid moiety at the C-3 position of glycyrrhizin and various soyasaponins has not been determined. Here, we report that a cellulose synthase superfamily-derived glycosyltransferase (CSyGT) catalyses 3-O-glucuronosylation of triterpenoid aglycones. Gene co-expression analyses of three legume species (Glycyrrhiza uralensis, Glycine max, and Lotus japonicus) reveal the involvement of CSyGTs in saponin biosynthesis, and we characterise CSyGTs in vivo using Saccharomyces cerevisiae. CSyGT mutants of L. japonicus do not accumulate soyasaponin, but the ectopic expression of endoplasmic reticulum membrane–localised CSyGTs in a L. japonicus mutant background successfully complement soyasaponin biosynthesis. Finally, we produced glycyrrhizin de novo in yeast, paving the way for sustainable production of high-value saponins.

Highlights

Triterpenoid saponins are specialised metabolites distributed widely in the plant kingdom that consist of one or more sugar moieties attached to triterpenoid aglycones
We observed that Glyma.06G324300 (GmCSyGT1), a member of the cellulose-synthase superfamily, had an expression pattern strongly correlated with that of genes involved in soyasaponin biosynthesis (Fig. 2a)
GuCSyGT (Glyur003152s00037491) and LjCSyGT (Lj3g3v1981230) had an amino acid sequence identity of >80%, and their expression patterns were highly correlated with those of genes involved in saponin biosynthesis (Fig. 2b, c)

Summary

Introduction

Triterpenoid saponins are specialised metabolites distributed widely in the plant kingdom that consist of one or more sugar moieties attached to triterpenoid aglycones. Despite the widely accepted view that glycosylation is catalysed by UDP-dependent glycosyltransferase (UGT), the UGT which catalyses the transfer of the conserved glucuronic acid moiety at the C-3 position of glycyrrhizin and various soyasaponins has not been determined. Triterpenoid saponin biosynthesis begins with cyclisation of the common precursor 2,3-oxidosqualene by oxidosqualene cyclases (OSCs)[11] into various triterpene scaffolds These triterpene scaffolds undergo site-specific oxidation catalysed by cytochrome P450 monooxygenase (P450s), forming diverse triterpenoid aglycones or non-glycosylated forms. We demonstrate that a cellulose-synthase superfamily-derived glycosyltransferase (CSyGT) catalyses the transfer of glucuronic acid from UDP-glucuronic acid to the C-3 position of oleanane-type triterpenoid aglycones. Our study challenges the conventional theory that specialised plant metabolites are glycosylated by UGTs and provides a foundation for the microbial production of glycyrrhizin, in a stable, costefficient manner

Methods

Results

Conclusion