Department of Plant and Environmental Sciences and Copenhagen Plant Science Center, University of Copenhagen, Thorvaldsensvej 40, 1871, Frederiksberg, Denmark.
Plant Mol Biol. 2018 May;97(1-2):37-55. doi: 10.1007/s11103-018-0723-z. Epub 2018 Mar 30.
This study identifies six UGT73Cs all able to glucosylate sapogenins at positions 3 and/or 28 which demonstrates that B. vulgaris has a much richer arsenal of UGTs involved in saponin biosynthesis than initially anticipated. The wild cruciferous plant Barbarea vulgaris is resistant to some insects due to accumulation of two monodesmosidic triterpenoid saponins, oleanolic acid 3-O-β-cellobioside and hederagenin 3-O-β-cellobioside. Insect resistance depends on the structure of the sapogenin aglycone and the glycosylation pattern. The B. vulgaris saponin profile is complex with at least 49 saponin-like metabolites, derived from eight sapogenins and including up to five monosaccharide units. Two B. vulgaris UDP-glycosyltransferases, UGT73C11 and UGT73C13, O-glucosylate sapogenins at positions 3 and 28, forming mainly 3-O-β-D-glucosides. The aim of this study was to identify UGTs responsible for the diverse saponin oligoglycoside moieties observed in B. vulgaris. Twenty UGT genes from the insect resistant genotype were selected and heterologously expressed in Nicotiana benthamiana and/or Escherichia coli. The extracts were screened for their ability to glycosylate sapogenins (oleanolic acid, hederagenin), the hormone 24-epibrassinolide and sapogenin monoglucosides (hederagenin and oleanolic acid 3-O-β-D-glucosides). Six UGTs from the UGT73C subfamily were able to glucosylate both sapogenins and both monoglucosides at positions 3 and/or 28. Some UGTs formed bisdesmosidic saponins efficiently. At least four UGT73C genes were localized in a tandem array with UGT73C11 and possibly UGT73C13. This organization most likely reflects duplication events followed by sub- and neofunctionalization. Indeed, signs of positive selection on several amino acid sites were identified and modelled to be localized on the UGT protein surface. This tandem array is proposed to initiate higher order bisdesmosidic glycosylation of B. vulgaris saponins, leading to the recently discovered saponin structural diversity, however, not directly to known cellobiosidic saponins.
本研究鉴定了六个 UGT73C 都能够在 3 位和/或 28 位糖基化甾体皂角苷，这表明 B. vulgaris 参与皂角苷生物合成的 UGT 种类比最初预期的要丰富得多。野生十字花科植物 Barbarea vulgaris 由于积累了两种单糖基三萜皂苷，齐墩果酸 3-O-β-棉子糖和常春藤皂苷元 3-O-β-棉子糖，对某些昆虫具有抗性。昆虫抗性取决于皂角苷元糖苷配基的结构和糖基化模式。B. vulgaris 皂苷的特征是复杂的，至少有 49 种皂苷类似代谢物，来源于八种甾体皂角苷，包括多达五个单糖单元。两个 B. vulgaris UDP-糖基转移酶 UGT73C11 和 UGT73C13 在 3 位和 28 位糖基化甾体皂角苷，主要形成 3-O-β-D-葡萄糖苷。本研究的目的是鉴定负责 B. vulgaris 中观察到的不同皂苷寡糖部分的 UGTs。从抗虫基因型中选择了 20 个 UGT 基因，并在 Nicotiana benthamiana 和/或 Escherichia coli 中异源表达。提取物被筛选以鉴定它们糖基化甾体皂角苷（齐墩果酸、常春藤皂苷元）、激素 24-表油菜素内酯和甾体皂角苷单葡萄糖苷（常春藤皂苷元和齐墩果酸 3-O-β-D-葡萄糖苷）的能力。UGT73C 亚家族的 6 个 UGT 能够在 3 位和/或 28 位糖基化甾体皂角苷和单葡萄糖苷。一些 UGT 有效地形成双糖基皂苷。至少有四个 UGT73C 基因定位于 UGT73C11 及其可能的 UGT73C13 的串联排列中。这种组织很可能反映了随后的亚功能化和新功能化的重复事件。事实上，在几个氨基酸位点上发现了正选择的迹象，并对其进行了建模，认为这些迹象位于 UGT 蛋白表面。这个串联排列可能启动了 B. vulgaris 皂苷的更高阶双糖基化，导致了最近发现的皂苷结构多样性，但与已知的纤维二糖皂苷无关。
