title: Functional Characterization of Cucurbitadienol Synthase and Triterpene Glycosyltransferase Involved in Biosynthesis of Mogrosides from Siraitia grosvenorii.
abstract: Mogrosides, the major bioactive components isolated from the fruits of Siraitia grosvenorii, are a family of cucurbitane-type tetracyclic triterpenoid saponins that are used worldwide as high-potency sweeteners and possess a variety of notable pharmacological activities. Mogrosides are synthesized from 2,3-oxidosqualene via a series of reactions catalyzed by cucurbitadienol synthase (CbQ), Cyt P450s (P450s) and UDP glycosyltransferases (UGTs) in vivo. However, the relevant genes have not been characterized to date. In this study, we report successful identification of SgCbQ and UGT74AC1, which were previously predicted via RNA-sequencing (RNA-seq) and digital gene expression (DGE) profile analysis of the fruits of S. grosvenorii. SgCbQ was functionally characterized by expression in the lanosterol synthase-deficient yeast strain GIL77 and was found to accumulate cucurbitadienol as the sole product. UGT74AC1 was heterologously expressed in Escherichia coli as a His-tag protein and it showed specificity for mogrol by transfer of a glucose moiety to the C-3 hydroxyl to form mogroside IE by in vitro enzymatic activity assays. This study reports the identification of CbQ and glycosyltransferase from S. grosvenorii for the first time. The results also suggest that RNA-seq, combined with DGE profile analysis, is a promising approach for discovery of candidate genes involved in biosynthesis of triterpene saponins.
authors.affiliation: National Engineering Laboratory for Industrial Enzymes, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China.
authors.forename: Longhai
authors.initials: L
authors.lastname: Dai
authors.affiliation: National Engineering Laboratory for Industrial Enzymes, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China.
authors.forename: Can
authors.initials: C
authors.lastname: Liu
authors.affiliation: National Engineering Laboratory for Industrial Enzymes, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China.
authors.forename: Yueming
authors.initials: Y
authors.lastname: Zhu
authors.affiliation: National Engineering Laboratory for Industrial Enzymes, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China.
authors.forename: Jiangsheng
authors.initials: J
authors.lastname: Zhang
authors.affiliation: National Engineering Laboratory for Industrial Enzymes, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Men
authors.affiliation: National Engineering Laboratory for Industrial Enzymes, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zeng
authors.affiliation: National Engineering Laboratory for Industrial Enzymes, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China syx0430@hotmail.com.
authors.forename: Yuanxia
authors.initials: Y
authors.lastname: Sun
mesh_terms: D000595:Amino Acid Sequence; D053898:Biosynthetic Pathways; D002851:Chromatography, High Pressure Liquid; D019662:Cucurbitaceae / Q000201:enzymology*; D018076:DNA, Complementary / Q000235:genetics; D044950:Flavanones / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D016695:Glycosyltransferases / Q000378:metabolism*; D019751:Intramolecular Transferases / Q000737:chemistry / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D010802:Phylogeny; D011794:Quercetin / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D016415:Sequence Alignment; D014315:Triterpenes / Q000737:chemistry / Q000378:metabolism*
chemical_list: D018076:DNA, Complementary; D044950:Flavanones; D014315:Triterpenes; D011794:Quercetin; D016695:Glycosyltransferases; D019751:Intramolecular Transferases; C021924:lanosterol synthase; C005273:naringenin
keywords: Cucurbitadienol; Cucurbitadienol synthase; Glycosyltransferase; Mogrosides; Siraitia grosvenorii