Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia, V6T 1Z4, Canada.
Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia, V6T 1Z4, Canada
Plant Physiol. 2019 Jul;180(3):1277-1290. doi: 10.1104/pp.19.00254. Epub 2019 Apr 19.
The plant metabolite montbretin A (MbA) and its precursor mini-MbA are potential new drugs for treating type 2 diabetes. These complex acylated flavonol glycosides only occur in small amounts in the corms of the ornamental plant montbretia ( × ). Our goal is to metabolically engineer using montbretia genes to achieve increased production of mini-MbA and MbA. Two montbretia UDP-dependent glycosyltransferases (UGTs), CcUGT1 and CcUGT2, catalyze the formation of the first two pathway-specific intermediates in MbA biosynthesis, myricetin 3--rhamnoside and myricetin 3--glucosyl rhamnoside. In previous work, expression of these UGTs in resulted in small amounts of kaempferol glycosides but not myricetin glycosides, suggesting that myricetin was limiting. Here, we investigated montbretia genes and enzymes of flavonol biosynthesis to enhance myricetin formation in We characterized two flavanone hydroxylases, a flavonol synthase, a flavonoid 3'-hydroxylase (F3'H), and a flavonoid 3'5'-hydroxylase (F3'5'H). Montbretia flavonol synthase converted dihydromyricetin into myricetin. Unexpectedly, montbretia F3'5'H shared higher sequence relatedness with F3'Hs in the CYP75B subfamily of cytochromes P450 than with those with known F3'5'H activity. Transient expression of combinations of montbretia flavonol biosynthesis genes and a montbretia MYB transcription factor in resulted in availability of myricetin for MbA biosynthesis. Transient coexpression of montbretia flavonol biosynthesis genes combined with and in resulted in 2 mg g fresh weight of the MbA pathway-specific compound myricetin 3--glucosyl rhamnoside. Additional expression of the montbretia acyltransferase led to detectable levels of mini-MbA in .
植物代谢产物蒙布特林 A（MbA）及其前体迷你-MbA 是治疗 2 型糖尿病的潜在新药。这些复杂的酰化黄酮醇糖苷仅在观赏植物萱草的球茎中以少量存在。我们的目标是利用萱草基因对 进行代谢工程改造，以实现迷你-MbA 和 MbA 的产量增加。两种萱草 UDP 依赖性糖基转移酶（UGTs），CcUGT1 和 CcUGT2，催化 MbA 生物合成中前两个途径特异性中间体的形成，即杨梅素 3--鼠李糖苷和杨梅素 3--葡萄糖基鼠李糖苷。在之前的工作中，这些 UGT 在 中的表达导致了少量的山奈酚糖苷，但没有杨梅素糖苷，这表明杨梅素是有限的。在这里，我们研究了萱草类黄酮生物合成的基因和酶，以增强 中杨梅素的形成。我们鉴定了两种黄烷酮羟化酶、一种类黄酮合酶、一种类黄酮 3'-羟化酶（F3'H）和一种类黄酮 3'5'-羟化酶（F3'5'H）。萱草类黄酮合酶将二氢杨梅素转化为杨梅素。出乎意料的是，萱草 F3'5'H 与细胞色素 P450 CYP75B 亚家族中的 F3'Hs 的序列相关性高于与已知具有 F3'5'H 活性的 F3'Hs 的序列相关性。在 中瞬时表达萱草类黄酮生物合成基因和一个萱草 MYB 转录因子的组合导致 MbA 生物合成中杨梅素的可用性。在 中瞬时共表达萱草类黄酮生物合成基因以及 和 导致 MbA 途径特异性化合物杨梅素 3--葡萄糖基鼠李糖苷的含量达到 2 mg g 鲜重。额外表达萱草酰基转移酶 导致 在 中可检测到迷你-MbA 的水平。
