Expression of a Dianthus flavonoid glucosyltransferase in Saccharomyces cerevisiae for whole-cell biocatalysis

• Published in: Journal of biotechnology Vol. 142; no. 3; pp. 233 - 241
• Main Authors: Werner, Sean R., Morgan, John A.
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 15.07.2009; Elsevier
• Subjects: Bioconversions. Hemisynthesis; Biological and medical sciences; Biotechnology; Cell Culture Techniques - methods; Chromatography, High Pressure Liquid; Dianthus; Dianthus - enzymology; Dianthus - genetics; Dianthus caryophyllus; Eugenia; Flavanones - metabolism; Fundamental and applied biological sciences. Psychology; Galactose inducible promoter; Glucosides - metabolism; Glucosyltransferases - biosynthesis; Glucosyltransferases - genetics; Glucosyltransferases - metabolism; Glyceraldehyde-3-phosphate dehydrogenase promoter; Glycosylation; Methods. Procedures. Technologies; Naringenin glycoside; Nuclear Magnetic Resonance, Biomolecular; Phloretin - metabolism; Phlorhizin - metabolism; Phlorizin; Plant Proteins - biosynthesis; Plant Proteins - genetics; Plant Proteins - metabolism; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Saccharomyces cerevisiae; Saccharomyces cerevisiae - enzymology; Saccharomyces cerevisiae - genetics; UDP-glucose; Phlorizin; Glyceraldehyde-3-phosphate dehydrogenase promoter; Naringenin glycoside; Galactose inducible promoter; UDP-glucose; Ascomycota; Phosphates; Yeast; Biocatalysis; Enzyme; Glucose; Glyceraldehyde-3-phosphate; Dehydrogenase; Dianthus; Flavonoid; Caryophyllaceae; dehydrogenase promoter; Fungi; Dicotyledones; Angiospermae; Spermatophyta; Glycoside; Oxidoreductases; Galactose; Saccharomyces cerevisiae
• ISSN: 0168-1656; 1873-4863
• DOI: 10.1016/j.jbiotec.2009.05.008

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.