期刊
TOXINS
卷 9, 期 2, 页码 -出版社
MDPI
DOI: 10.3390/toxins9020058
关键词
zearalenone-14-glucoside; zearalenone-16-glucoside; masked mycotoxin; glycosylation; sucrose synthase; secondary metabolite
资金
- Vienna Science and Technology Fund [WWTF LS12-021]
- Austrian Science Fund (FWF) [SFB F3706, F3708, F3715]
- Austrian Federal Ministry of Science, Research and Economy
- National Foundation of Research, Technology and Development
- BIOMIN Holding GmbH
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 (K-m = 3 mu M) and catalytic efficiency (k(cat)/K-m = 190 s(-1.)mM(-1)) with ZEN. It also efficiently glucosylates the phase-I ZEN-metabolites alpha-zearalenol and beta-zearalenol, with k(cat)/K-m 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 beta-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.
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