Diabetes and obesity are affecting human health worldwide. Their occurrence is increasing with lifestyle choices, globalization of food systems, and economic development. The specialized plant metabolite montbretin A (MbA) is being developed as an antidiabetes and antiobesity treatment due to its potent and specific inhibition of the human pancreatic α-amylase. MbA is a complex acylated flavonol glycoside formed in small amounts in montbretia (Crocosmia x crocosmiiflora) corms during the early summer. The spatial and temporal patterns of MbA accumulation limit its supply for drug development and application. We are exploring MbA biosynthesis to enable metabolic engineering of this rare and valuable compound. Genes and enzymes for the first four steps of MbA biosynthesis, starting from the flavonol precursor myricetin, have recently been identified. Here, we describe the gene discovery and functional characterization of the final two enzymes of MbA biosynthesis. The UDP-glycosyltransferases, CcUGT4 and CcUGT5, catalyze consecutive reactions in the formation of the disaccharide moiety at the 4’-hydroxy position of the MbA flavonol core. CcUGT4 is a flavonol glycoside 4’-O-xylosyltransferase that acts on the second to last intermediate (MbA-XR²) in the pathway. CcUGT5 is a flavonol glycoside 1,4-rhamnosyltransferase that converts the final intermediate (MbA-R²) to complete the MbA molecule. Both enzymes belong to the UGT family d-clade and are specific for flavonol glycosides and their respective sugar donors. This study concludes the discovery of the MbA biosynthetic pathway and provides the complete set of genes to engineer MbA biosynthesis. We demonstrate successful reconstruction of MbA biosynthesis in Nicotiana benthamiana.

糖尿病和肥胖正在影响全世界人类健康。随着生活方式的选择、食品系统的全球化和经济的发展，它们的发生率正在增加。特殊的植物代谢物 montbretin A (MbA) 因其对人类胰腺 α-淀粉酶的有效和特异性抑制而被开发为抗糖尿病和抗肥胖疗法。 MbA 是一种复杂的酰化黄酮醇糖苷，在初夏期间在 montbretia ( Crocosmia x crocosmiiflora ) 球茎中少量形成。 MbA 积累的空间和时间模式限制了其对药物开发和应用的供应。我们正在探索 MbA 生物合成，以实现这种稀有且有价值的化合物的代谢工程。最近，从黄酮醇前体杨梅素开始，MbA 生物合成前四个步骤的基因和酶已被鉴定。在这里，我们描述了 MbA 生物合成的最后两种酶的基因发现和功能表征。 UDP-糖基转移酶 CcUGT4 和 CcUGT5 催化 MbA 黄酮醇核心 4'-羟基位置上的二糖部分形成的连续反应。 CcUGT4 是一种黄酮醇糖苷 4'- O-木糖基转移酶，作用于该途径中倒数第二个中间体 (MbA-XR ² )。 CcUGT5 是一种黄酮醇糖苷 1,4-鼠李糖基转移酶，可转化最终中间体 (MbA-R ² ) 以完成 MbA 分子。两种酶均属于 UGT 家族d -进化枝并且对黄酮醇糖苷及其各自的糖供体具有特异性。这项研究总结了 MbA 生物合成途径的发现，并提供了设计 MbA 生物合成的全套基因。 我们展示了本塞姆氏烟草中 MbA 生物合成的成功重建。