Synthesis of Mono- and Di-Glucosides of Zearalenone and α-/β-Zearalenol by Recombinant Barley Glucosyltransferase HvUGT14077

Toxins

Alexandra Malachová¹  Franz Berthiller¹  Francesca Lupi¹  Elisabeth Varga¹  Gerhard Adam²  Herbert Michlmayr²  Christian Hametner³ 

[1] Center for Analytical Chemistry and Christian Doppler Laboratory for Mycotoxin Metabolism, Department of Agrobiotechnology (IFA-Tulln), BOKU, Konrad Lorenz Straße 20, 3430 Tulln, Austria;Department of Applied Genetics and Cell Biology, University of Natural Resources and Life Sciences, Vienna (BOKU), Konrad Lorenz Straße 24, 3430 Tulln, Austria;Institute of Applied Synthetic Chemistry, Vienna University of Technology, Getreidemarkt 9/163, 1060 Vienna, Austria;

关键词: zearalenone-14-glucoside;    zearalenone-16-glucoside;    masked mycotoxin;    glycosylation;    sucrose synthase;    secondary metabolite;   

DOI  :  10.3390/toxins9020058

来源: DOAJ

【 摘 要 】

Zearalenone (ZEN) is an estrogenic mycotoxin occurring in Fusarium-infected cereals. Glucosylation is an important plant defense mechanism and generally reduces the acute toxicity of mycotoxins to humans and animals. Toxicological information about ZEN-glucosides is limited due to the unavailability of larger amounts required for animal studies. HvUGT14077, a recently-validated ZEN-conjugating barley UDP-glucosyltransferase was expressed in Escherichia coli, affinity purified, and characterized. HvUGT14077 possesses high affinity (Km = 3 µM) and catalytic efficiency (kcat/Km = 190 s−1·mM−1) with ZEN. It also efficiently glucosylates the phase-I ZEN-metabolites α-zearalenol and β-zearalenol, with kcat/Km of 40 and 74 s−1·mM−1, respectively. HvUGT14077 catalyzes O-glucosylation at C-14 and C-16 with preference of 14-glucoside synthesis. Furthermore, relatively slow consecutive formation of 14,16-di-glucosides was observed; their structures were tentatively identified by mass spectrometry and for ZEN-14,16-di-glucoside confirmed by nuclear magnetic resonance spectroscopy. Recombinant HvUGT14077 allowed efficient preparative synthesis of ZEN-glucosides, yielding about 90% ZEN-14-glucoside and 10% ZEN-16-glucoside. The yield of ZEN-16-glucoside could be increased to 85% by co-incubation with a β-glucosidase highly selective for ZEN-14-glucoside. Depletion of the co-substrate UDP-glucose was counteracted by a sucrose synthase based regeneration system. This strategy could also be of interest to increase the yield of minor glucosides synthesized by other glucosyltransferases.

【 授权许可】

Unknown