title: Oxidation of Cucurbitadienol Catalyzed by CYP87D18 in the Biosynthesis of Mogrosides from Siraitia grosvenorii.
abstract: Mogrosides, the principally bioactive compounds extracted from the fruits of Siraitia grosvenorii, are a group of glycosylated cucurbitane-type tetracyclic triterpenoid saponins that exhibit a wide range of notable biological activities and are commercially available worldwide as natural sweeteners. The biosynthesis of mogrosides involves initial cyclization of 2,3-oxidosqualene to the triterpenoid skeleton of cucurbitadienol, followed by a series of oxidation reactions catalyzed by Cyt P450s (P450s) and then glycosylation reactions catalyzed by UDP glycosyltransferases (UGTs). We previously reported the identification of a cucurbitadienol synthase (SgCbQ) and a mogrol C-3 hydroxyl glycosyltransferase (UGT74AC1). However, molecular characterization of further transformation of cucurbitadienol to mogrol by P450s remains unavailable. In this study, we report the successful identification of a multifunctional P450 (CYP87D18) as being involved in C-11 oxidation of cucurbitadienol. In vitro enzymatic activity assays showed that CYP87D18 catalyzed the oxidation of cucurbitadienol at C-11 to produce 11-oxo cucurbitadienol and 11-hydroxy cucurbitadienol. Furthermore, 11-oxo-24,25-epoxy cucurbitadienol as well as 11-oxo cucurbitadienol and 11-hydroxy cucurbitadienol were produced when CYP87D18 was co-expressed with SgCbQ in genetic yeast, and their structures were confirmed by liquid chromatography-solid-phase extraction-nuclear magnetic resonance-mass spectrometry coupling (LC-SPE-NMR-MS). Taken together, these results suggest a role for CYP87D18 as a multifunctional cucurbitadienol oxidase in the mogrosides pathway.
authors.affiliation: National Engineering Laboratory for Industrial Enzymes, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin, 300308, China These authors contributed equally to this work.
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 These authors contributed equally to this work.
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: Jiangang
authors.initials: J
authors.lastname: Yang
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: 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.
authors.forename: Yi
authors.initials: Y
authors.lastname: Cai
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 syx0430@hotmail.com.
authors.forename: Yuanxia
authors.initials: Y
authors.lastname: Sun
mesh_terms: D053898:Biosynthetic Pathways; D002384:Catalysis; D019662:Cucurbitaceae / Q000201:enzymology* / Q000235:genetics; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D005638:Fruit / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression; D006027:Glycosides / Q000378:metabolism*; D016695:Glycosyltransferases / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D012503:Saponins / Q000378:metabolism; D013185:Squalene / Q000031:analogs & derivatives / Q000378:metabolism; D014315:Triterpenes / Q000378:metabolism*
chemical_list: C002821:2,3-oxidosqualene; D006027:Glycosides; D010940:Plant Proteins; D012503:Saponins; D014315:Triterpenes; C514277:cucurbitane; D013185:Squalene; D003577:Cytochrome P-450 Enzyme System; D016695:Glycosyltransferases
keywords: CYP87D18; Cucurbitadienol; Cytochrome P450; Mogrosides; Siraitia grosvenorii