金龙胆草3-O-葡萄糖基转移酶基因的克隆及功能预测

徐小英; 吴谦; 孙蓉; 刘姗; 郑毅

金龙胆草3-O-葡萄糖基转移酶基因的克隆及功能预测

Cloning and Functional Prediction of the 3-O-Glucosyltransferase Gene from Conyza blinii

摘要

摘要:
目的克隆金龙胆草（Conyza blinii H. Lév）3-O-葡萄糖基转移酶（3-O-glucosyltransferase, 3GT）基因，为其生物学功能分析与研究提供材料基础。
方法基于课题组前期测序的金龙胆草转录组数据设计引物序列，利用PCR技术克隆3GT基因，通过生物信息学工具分析其分子特性，利用分子对接技术预测3GT底物特异性。
结果金龙胆草3GT基因（Conyza blinii 3-O-glucosyltransferase, CbUGT）cDNA全长1470 bp，编码489个氨基酸，理论相对分子质量54.88 kDa，等电点5.50，不稳定指数（II）为43.12，亲水性系数−0.185，表明CbUGT为不稳定亲水性蛋白；BLAST比对表明CbUGT含有糖基转移酶家族1和具有GTB拓扑结构的相关蛋白结构域；进化树分析表明其与拟南芥D类UGT73C1聚为一类；亚细胞预测其定位于细胞膜。分子对接预测CbUGT对葡萄糖（glucose,Glc）和齐墩果酸（oleanolic acid, OA）2种分子均具有结合能力，与结构分析相吻合。
结论 CbUGT基因编码一个典型的植物3-O-葡萄糖基转移酶基因，可能与金龙胆草皂苷C3位上的葡萄糖转移有关。

Abstract:
Objective 3-O-glucosyltransferase gene (3GT) was cloned from Conyza blinii H. Lév for a biological function analysis.
Methods Based on the previously sequenced C. blinii transcriptome, a primer was designed to clone 3GT using PCR for the bioinformatic analysis. Molecular dockings of the protein were determined.
Results The cloned 3GT was named CbUGT that had a cDNA with the full-length of 1 470 bp encoding 489 amino acids. The unstable hydrophilic protein had a theoretical molecular weight of 54.88 kDa, an isoelectric point of 5.50, an instability index (II) of 43.12, and a hydrophilicity coefficient of −0.185. The BLAST alignment revealed that it contained a glycosyltransferase family 1 domain and a GTB topology-related protein domain. Phylogenetically, it was clustered with Arabidopsis thaliana D-class UGT73C1 and localized on plasma membrane. Consistent with the prediction by a structure analysis, the molecular docking of the protein showed a binding affinity to glucose and oleanolic acid.
Conclusion CbUGT encoded a typical plant 3-O-glucosyltransferase and might be involved in the glucosyl transfer at C-3 position of saponins in C. blinii.

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