title: Steroid 5β-Reductase from Leaves of Vitis vinifera: Molecular Cloning, Expression, and Modeling.
abstract: A steroid 5β-reductase gene corresponding to the hypothetical protein LOC100247199 from leaves of Vitis vinifera (var. 'Chardonnay') was cloned and overexpressed in Escherichia coli. The recombinant protein showed 5β-reductase activity when progesterone was used as a substrate. The reaction was stereoselective, producing only 5β-products such as 5β-pregnane-3,20-dione. Other small substrates (terpenoids and enones) were also accepted as substrates, indicating the highly promiscuous character of the enzyme class. Our results show that the steroid 5β-reductase gene, encoding an orthologous enzyme described as a key enzyme in cardenolide biosynthesis, is also expressed in leaves of the cardenolide-free plant V. vinifera. We emphasize the fact that, on some occasions, different reductases (e.g., progesterone 5β-reductase and monoterpenoid reductase) can also use molecules that are similar to the final products as a substrate. Therefore, in planta, the different reductases may contribute to the immense number of diverse small natural products finally leading to the flavor of wine.
authors.affiliation: Chair of Pharmaceutical Biology, Department Biology, Friedrich-Alexander-University Erlangen-Nürnberg , Staudtstrasse 5, 91058 Erlangen, Germany.
authors.forename: Mona
authors.initials: M
authors.lastname: Ernst
authors.affiliation: Chair of Pharmaceutical Biology, Department Biology, Friedrich-Alexander-University Erlangen-Nürnberg , Staudtstrasse 5, 91058 Erlangen, Germany.
authors.forename: Jennifer
authors.initials: J
authors.lastname: Munkert
authors.affiliation: Research and Innovation Centre, Fondazione Edmund Mach (FEM) , Via Mach 1, 38010 San Michele all'Adige (Trentino), Italy.
authors.forename: Manuela
authors.initials: M
authors.lastname: Campa
authors.affiliation: Research and Innovation Centre, Fondazione Edmund Mach (FEM) , Via Mach 1, 38010 San Michele all'Adige (Trentino), Italy.
authors.forename: Mickael
authors.initials: M
authors.lastname: Malnoy
authors.affiliation: Research and Innovation Centre, Fondazione Edmund Mach (FEM) , Via Mach 1, 38010 San Michele all'Adige (Trentino), Italy.
authors.forename: Stefan
authors.initials: S
authors.lastname: Martens
authors.affiliation: Chair of Pharmaceutical Biology, Department Biology, Friedrich-Alexander-University Erlangen-Nürnberg , Staudtstrasse 5, 91058 Erlangen, Germany.
authors.forename: Frieder
authors.initials: F
authors.lastname: Müller-Uri
mesh_terms: D013741:3-Oxo-5-alpha-Steroid 4-Dehydrogenase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002298:Cardenolides / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D015870:Gene Expression; D008958:Models, Molecular; D018515:Plant Leaves / Q000201:enzymology*; D011994:Recombinant Proteins / Q000378:metabolism; D013649:Taste; D027843:Vitis / Q000201:enzymology*; D014920:Wine / Q000032:analysis
chemical_list: D002298:Cardenolides; D011994:Recombinant Proteins; D013741:3-Oxo-5-alpha-Steroid 4-Dehydrogenase
keywords: Vitis vinifera; biosynthesis; enone 5β-reductase; gene evolution; secondary metabolites