Functional Characterization of Cucurbitadienol Synthase and Triterpene Glycosyltransferase Involved in Biosynthesis of Mogrosides from Siraitia grosvenorii
Functional Characterization of Cucurbitadienol Synthase and Triterpene Glycosyltransferase Involved in Biosynthesis of Mogrosides from Siraitia grosvenorii
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DOI：
10.1093/pcp/pcv043
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发表时间：
2015-06-01
影响因子：
4.9
通讯作者：
Sun, Yuanxia
中科院分区：
文献类型：
--
作者：
Dai, Longhai;Liu, Can;Sun, Yuanxia
罗汉果皂苷是从罗汉果果实中分离得到的主要生物活性成分，是葫芦烷型四环三萜皂苷家族，具有多种显著的药理活性，被广泛用作高效甜味剂。罗汉果苷是由2，3-氧化角鲨烯在体内通过一系列由葫芦二烯醇合酶（CbQ）、Cyt P450 s（P450 s）和UDP糖基转移酶（UGT）催化的反应合成的。然而，迄今为止，相关基因尚未被表征。在这项研究中，我们报告了成功的鉴定SgCbQ和UGT 74 AC 1，这是以前预测通过RNA测序（RNA-seq）和数字基因表达（DGE）分析的果实S。罗汉果。SgCbQ的功能特征在于在羊毛甾醇脱氢酶缺陷型酵母菌株GIL 77中表达，并发现其积累葫芦二烯醇作为唯一产物。UGT 74 AC 1在大肠杆菌中异源表达为His-标签蛋白，并且通过体外酶活性测定，其通过将葡萄糖部分转移到C-3羟基以形成罗汉果苷IE而显示出对罗汉果醇的特异性。本研究报道了从S.格罗夫纳第一次。结果还表明，RNA-seq，结合DGE图谱分析，是一个有前途的方法，发现参与三萜皂苷生物合成的候选基因。
Mogrosides, the major bioactive components isolated from the fruits of Siraitia grosvenorii, are a family of cucurbitane-type tetracyclic triterpenoid saponins that are used worldwide as high-potency sweeteners and possess a variety of notable pharmacological activities. Mogrosides are synthesized from 2,3-oxidosqualene via a series of reactions catalyzed by cucurbitadienol synthase (CbQ), Cyt P450s (P450s) and UDP glycosyltransferases (UGTs) in vivo. However, the relevant genes have not been characterized to date. In this study, we report successful identification of SgCbQ and UGT74AC1, which were previously predicted via RNA-sequencing (RNA-seq) and digital gene expression (DGE) profile analysis of the fruits of S. grosvenorii. SgCbQ was functionally characterized by expression in the lanosterol synthase-deficient yeast strain GIL77 and was found to accumulate cucurbitadienol as the sole product. UGT74AC1 was heterologously expressed in Escherichia coli as a His-tag protein and it showed specificity for mogrol by transfer of a glucose moiety to the C-3 hydroxyl to form mogroside IE by in vitro enzymatic activity assays. This study reports the identification of CbQ and glycosyltransferase from S. grosvenorii for the first time. The results also suggest that RNA-seq, combined with DGE profile analysis, is a promising approach for discovery of candidate genes involved in biosynthesis of triterpene saponins.