title: Expression of a Dianthus flavonoid glucosyltransferase in Saccharomyces cerevisiae for whole-cell biocatalysis.
abstract: Glycosyltransferases are promising biocatalysts for the synthesis of small molecule glycosides. In this study, Saccharomyces cerevisiae expressing a flavonoid glucosyltransferase (GT) from Dianthus caryophyllus (carnation) was investigated as a whole-cell biocatalyst. Two yeast expression systems were compared using the flavonoid naringenin as a model substrate. Under in vitro conditions, naringenin-7-O-glucoside was formed and a higher specific glucosyl transfer activity was found using a galactose inducible expression system compared to a constitutive expression system. However, S. cerevisiae expressing the GT constitutively was significantly more productive than the galactose inducible system under in vivo conditions. Interestingly, the glycosides were recovered directly from the culture broth and did not accumulate intracellularly. A previously uncharacterized naringenin glycoside formed using the D. caryophyllus GT was identified as naringenin-4'-O-glucoside. It was found that S. cerevisiae cells hydrolyze naringenin-7-O-glucoside during whole-cell biocatalysis, resulting in a low final glycoside titer. When phloretin was added as a substrate to the yeast strain expressing the GT constitutively, the natural product phlorizin was formed. This study demonstrates S. cerevisiae is a promising whole-cell biocatalyst host for the production of valuable glycosides.
authors.affiliation: School of Chemical Engineering, Purdue University, West Lafayette, IN 47907, USA.
authors.forename: Sean R
authors.initials: SR
authors.lastname: Werner
authors.forename: John A
authors.initials: JA
authors.lastname: Morgan
mesh_terms: D018929:Cell Culture Techniques / Q000379:methods; D002851:Chromatography, High Pressure Liquid; D029749:Dianthus / Q000201:enzymology* / Q000235:genetics; D044950:Flavanones / Q000378:metabolism; D005960:Glucosides / Q000378:metabolism; D005964:Glucosyltransferases / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D006031:Glycosylation; D019906:Nuclear Magnetic Resonance, Biomolecular; D010693:Phloretin / Q000378:metabolism; D010695:Phlorhizin / Q000378:metabolism; D010940:Plant Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics
chemical_list: D044950:Flavanones; D005960:Glucosides; D010940:Plant Proteins; D011994:Recombinant Proteins; C529068:naringenin-7-O-glucoside; D010695:Phlorhizin; D005964:Glucosyltransferases; C005273:naringenin; D010693:Phloretin