Biochemical Characterization of a Recombinant UDP-glucosyltransferase from Rice and Enzymatic Production of Deoxynivalenol-3-O-β-D-glucoside.
Biochemical Characterization of a Recombinant UDP-glucosyltransferase from Rice and Enzymatic Production of Deoxynivalenol-3-O-β-D-glucoside.
复制标题
从水稻中重组UDP-葡萄糖基转移酶的生化表征以及脱氧烯醇-3-O-β-D-葡萄糖苷的酶促产生。
DOI：
10.3390/toxins7072685
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发表时间：
2015-07-21
期刊：
影响因子：
4.2
通讯作者：
Adam G
中科院分区：
文献类型：
--
作者：
Michlmayr H;Malachová A;Varga E;Kleinová J;Lemmens M;Newmister S;Rayment I;Berthiller F;Adam G
糖基化是一种重要的植物防御机制，镰刀菌毒素的偶联物经常与其母体化合物共存于谷类食品和饲料中。脱氧雪腐镰刀菌醇（DON）中，脱氧雪腐镰刀菌醇-3- o -β-d-葡萄糖苷（D3G）是最重要的掩蔽真菌毒素。D3G的毒理学意义尚未完全了解，因此获得纯度和足够数量的这种化合物用于毒理学风险评估和作为分析标准是至关重要的。本研究的目的是对水稻中一种don失活的udp -葡萄糖基转移酶（OsUGT79）进行生化鉴定，并探讨其用于制备D3G的适用性。重组OsUGT79的表观米歇利常数（Km）为0.23 mM DON和2.2 mM UDP-glucose。当DON浓度大于2 mM （Ki = 24 mM DON）时，底物抑制作用发生，UDP强烈抑制酶。Cu2+和Zn2+ (1 mM)完全抑制酶。蔗糖合酶AtSUS1在糖基化反应中再生udp -葡萄糖。使用这种方法，在有限浓度的昂贵的辅助因子udp -葡萄糖下可以获得最佳转化率。现在可以合成足够数量和纯度的D3G。类似的策略可能对生产其他毒素的β-糖苷感兴趣。
Glycosylation is an important plant defense mechanism and conjugates of Fusarium mycotoxins often co-occur with their parent compounds in cereal-based food and feed. In case of deoxynivalenol (DON), deoxynivalenol-3-O-β-d-glucoside (D3G) is the most important masked mycotoxin. The toxicological significance of D3G is not yet fully understood so that it is crucial to obtain this compound in pure and sufficient quantities for toxicological risk assessment and for use as an analytical standard. The aim of this study was the biochemical characterization of a DON-inactivating UDP-glucosyltransferase from rice (OsUGT79) and to investigate its suitability for preparative D3G synthesis. Apparent Michaelis constants (Km) of recombinant OsUGT79 were 0.23 mM DON and 2.2 mM UDP-glucose. Substrate inhibition occurred at DON concentrations above 2 mM (Ki = 24 mM DON), and UDP strongly inhibited the enzyme. Cu2+ and Zn2+ (1 mM) inhibited the enzyme completely. Sucrose synthase AtSUS1 was employed to regenerate UDP-glucose during the glucosylation reaction. With this approach, optimal conversion rates can be obtained at limited concentrations of the costly co-factor UDP-glucose. D3G can now be synthesized in sufficient quantity and purity. Similar strategies may be of interest to produce β-glucosides of other toxins.