Metabolic engineering of the flavone-C-glycoside pathway using polyprotein technology
Metabolic engineering of the flavone-C-glycoside pathway using polyprotein technology
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DOI：
10.1016/j.ymben.2012.11.004
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发表时间：
2013-03-01
影响因子：
8.4
通讯作者：
Edwards, Robert
中科院分区：
文献类型：
--
作者：
Brazier-Hicks, Melissa;Edwards, Robert
C-糖基化类黄酮是具有生物活性的植物天然产物，与膳食健康益处有关。我们已经使用多蛋白表达技术来重建烟草和酵母中各自的生物合成途径的一部分，使得二氢查耳酮和黄烷酮前体直接转化为C-糖苷。开发的多蛋白系统促进了在植物和酵母中需要不同亚细胞定位的途径酶的简单而有效的共表达。黄酮-C-葡糖苷的途径包括与C-葡糖基转移酶（CGT）共表达的黄烷酮2-羟化酶（F2 H）。虽然烟草中的途径工程化仅导致少量C-糖苷形成，但当用黄烷酮柚皮素进料时，用F2 H-CGT多蛋白构建体转化的酵母在培养基中产生高浓度的2-羟基柚皮素-C-糖苷。然后，这些发酵产物可以容易地化学转化为相应的黄酮-C-葡糖苷。当F2 H和CGT都从同一物种（水稻）获得时，生物合成的效率是最佳的。这些结果证实了F2 H和CGT在体内产生C-葡糖苷中的偶联作用，其中在酵母中使用多蛋白表达系统提供了一个有用的系统来优化这些天然产物的合成，其量适合于饮食研究。(c)2012 Elsevier Inc. All rights reserved.
C-Glycosylated flavonoids are biologically active plant natural products linked to dietary health benefits. We have used polyprotein expression technology to reconstruct part of the respective biosynthetic pathway in tobacco and yeast, such that dihydrochalcone and flavanone precursors are directly converted to C-glycosides. The polyprotein system developed facilitated the simple and efficient co-expression of pathway enzymes requiring different sub-cellular localization in both plants and yeast. The pathway to flavone-C-glucosides comprised a flavanone 2-hydroxylase (F2H), co-expressed with a C-glucosyltransferase (CGT). While pathway engineering in tobacco resulted in only minor C-glycoside formation, when fed with the flavanone naringenin, yeast transformed with the F2H-CGT polyprotein construct produced high concentrations of 2-hydroxynaringenin-C-glucoside in the medium. These fermentation products could then be readily chemically converted to the respective flavone-C-glucosides. The efficiency of the biosynthesis was optimal when both the F2H and CGT were obtained from the same species (rice). These results confirm the coupled roles of the F2H and CGT in producing C-glucosides in vivo, with the use of the polyprotein expression system in yeast offering a useful system to optimize the synthesis of these natural products in quantities suitable for dietary studies. (c) 2012 Elsevier Inc. All rights reserved.