PDF(Pubmed)
Abstract：
Aloesone is a bioactive natural product and biosynthetic precursor of rare glucosides found in rhubarb and some aloe plants including Aloe vera. This study aimed to investigate biocatalytic aloesone glycosylation and more than 400 uridine diphosphate-dependent glycosyltransferase (UGT) candidates, including multifunctional and promiscuous enzymes from a variety of plant species were assayed. As a result, 137 selective aloesone UGTs were discovered, including four from the natural producer rhubarb. Rhubarb UGT72B49 was further studied and its catalytic constants (kcat = 0.00092 ± 0.00003 s-1, KM = 30 ± 2.5 μM) as well as temperature and pH optima (50 °C and pH 7, respectively) were determined. We further aimed to find an efficient aloesone glycosylating enzyme with potential application for biocatalytic production of the glucoside. We discovered UGT71C1 from Arabidopsis thaliana as an efficient aloesone UGT showing a 167-fold higher catalytic efficiency compared to that of UGT72B49. Interestingly, sequence analysis of all the 137 newly identified aloesone UGTs showed that they belong to different phylogenetic groups, with the highest representation in groups B, D, E, F and L. Finally, our study indicates that aloesone C-glycosylation is highly specific and rare, since it was not possible to achieve in an efficient manner with any of the 422 UGTs assayed, including multifunctional GTs and 28 known C-UGTs.
摘要：
Aloesone是一种生物活性天然产物，是在大黄和包括芦荟在内的一些芦荟植物中发现的稀有葡糖苷的生物合成前体。这项研究旨在研究生物催化的芦荟酮糖基化和400多种尿苷二磷酸依赖性糖基转移酶（UGT）候选物，包括来自多种植物物种的多功能和混杂酶。因此,发现了137种选择性芦荟酮UGTs，包括四个来自天然生产者大黄。进一步研究了大黄UGT72B49，并确定了其催化常数（kcat=0.00092±0.00003s-1，KM=30±2.5μM）以及温度和最佳pH值（分别为50°C和pH7）。我们进一步旨在寻找一种有效的芦荟酮糖基化酶，具有潜在的生物催化生产葡糖苷的应用。我们发现拟南芥的UGT71C1是一种有效的芦荟酮UGT，与UGT72B49相比，其催化效率提高了167倍。有趣的是,对所有137个新鉴定的芦荟UGTs的序列分析表明，它们属于不同的系统发育群体，B组中代表性最高，D,E,F和L最后，我们的研究表明，芦荟酮C-糖基化是高度特异性和罕见的，由于不可能以有效的方式实现422个UGT中的任何一个，包括多功能GTs和28个已知的C-UGTs。
