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Transient expression and enzymatic assay identified uridine-diphosphate glucosyltransferases related to flavonoid glycosylation in Vernonia amygdalina leaves
Industrial Crops and Products ( IF 5.9 ) Pub Date : 2021-09-04 , DOI: 10.1016/j.indcrop.2021.114005
Kaisen Huo 1 , Yan Chen 1 , Lanya Sui 1 , Yu Wang 1 , Yijun Fu 1 , Xingjie Pei 1 , Jun Niu 1
Affiliation  

Although previous metabolomics analysis revealed that flavonoid glycosylation commonly occurred at 3−OH and 7−OH positions in Vernonia amygdalina leaves, the biosynthetic mechanisms of flavonoid glycosides are still largely unknown. Based on the previous transcriptome data for V. amygdalina leaves, a total of 62 uridine-diphosphate glycosyltransferase (UGT) genes were acquired. We conducted phylogenetic analyses and tested for expression profiles to further select the potential V. amygdalina UGT (VaUGT) related to flavonoid glycosylation. Functional analyses were conducted on the following 6 VaUGT genes: VaUGT9 and VaUGT40 from Clade I, VaUGT3 and VaUGT62 from Clade Ⅳ, and VaUGT38 and VaUGT49 from Clade Ⅵ. Transient expression analysis indicated that VaUGT38 might have substrate specificity involved in the 3−OH and 7−OH positions of kaempferol glycosylation, while VaUGT40 was related to the 7−OH position of kaempferol and apigenin glycosylation. In addition, VaUGT9 and VaUGT3 displayed glycosyltransferase activities at apigenin 7−OH and 7-O-glucoside 2″−OH positions, respectively. Recombinant expression analysis verified that the VaUGT9 and VaUGT38 proteins exhibit a substrate preference to apigenin and kaempferol, respectively. The recombinant VaUGT3, VaUGT9 and VaUGT40 proteins could use apigetrin as substrate to generate apigenin, in particular, VaUGT9 and VaUGT40 were the most efficient. Recombinant VaUGT3, VaUGT38, VaUGT40, VaUGT49, and VaUGT62 proteins exhibited glycosyltransferase activity toward kaempferol, among which VaUGT38 and VaUGT40 were the most efficient. These results improve our understanding of flavonoid glycosylation in V. amygdalina leaves.



中文翻译:

瞬时表达和酶促测定鉴定了与斑鸠菊叶中黄酮类糖基化相关的尿苷二磷酸葡萄糖基转移酶

尽管之前的代谢组学分析表明黄酮糖基化通常发生在斑鸠菊叶的3-OH 和 7-OH 位置,但黄酮苷的生物合成机制仍然在很大程度上未知。根据先前的杏仁核转录组数据,共获得了 62 个尿苷二磷酸糖基转移酶 ( UGT ) 基因。我们进行了系统发育分析并测试了表达谱,以进一步选择与类黄酮糖基化相关的潜在V. amygdalina UGT ( VaUGT )。对以下 6 个VaUGT基因进行功能分析:VaUGT9VaUGT40从克拉德我,VaUGT3VaUGT62分化单位Ⅳ和VaUGT38VaUGT49来自进化枝Ⅵ。瞬时表达分析表明,VaUGT38 可能具有参与山奈酚糖基化的 3-OH 和 7-OH 位置的底物特异性,而 VaUGT40 与山奈酚和芹菜素糖基化的 7-OH 位置有关。此外,VaUGT9 和 VaUGT3 分别在芹菜素 7-OH 和 7-O-葡萄糖苷 2"-OH 位置显示出糖基转移酶活性。重组表达分析证实 VaUGT9 和 VaUGT38 蛋白分别表现出对芹菜素和山奈酚的底物偏好。重组VaUGT3、VaUGT9和VaUGT40蛋白可以以芹菜素为底物生成芹菜素,其中以VaUGT9和VaUGT40最为有效。重组 VaUGT3、VaUGT38、VaUGT40、VaUGT49 和 VaUGT62 蛋白表现出对山奈酚的糖基转移酶活性,其中 VaUGT38 和 VaUGT40 的效率最高。这些结果提高了我们对类黄酮糖基化的理解V.杏仁核叶。

更新日期:2021-09-04
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