Li Jia, Li Zhaobo, Li Changfu, Gou Junbo, Zhang Yansheng
CAS Key Laboratory of Plant Germplasm Enhancement and Specialty Agriculture, Wuhan Botanical Garden, Chinese Academy of Sciences, Wuhan, 430074, China,
Plant Cell Rep. 2014 Jul;33(7):1173-85. doi: 10.1007/s00299-014-1606-7. Epub 2014 Apr 4.
A novel isoflavone 7- O -glucosyltransferase PlUGT1 was isolated from Pueraria lobata . PlUGT1 could convert daidzein to daidzin, genistein to genistin as well as formononetin to ononin. Pueraria lobata roots are traditionally consumed as a rich source of isoflavone glycosides that have various human health benefits. However, to date, the genes encoding isoflavone UDP-glycosyltransferases (UGTs) have only been isolated from the roots of soybean seedlings (GmIF7GT), soybean seeds (UGT73F2) and Glycyrrhiza echinata cell suspension cultures (GeIF7GT). To investigate the isoflavone metabolism in P. lobata, 40 types of partial UGT cDNAs were isolated from P. lobata, and seven full-length UGT candidates with preferential expression in roots were identified. Functional assays in yeast (Saccharomyces cerevisiae) revealed that one of these UGT candidates, designated PlUGT1 (official UGT designation UGT88E12), efficiently glycosylated isoflavone aglycones at the 7-hydroxy group. Recombinant PlUGT1 purified from Escherichia coli cells was characterized and shown to be relatively specific for isoflavone aglycones, while flavonoid substrates were poorly accepted. The biochemical results suggested that PlUGT1 was an isoflavone 7-O-glucosyltransferase. The deduced amino acid sequence of PlUGT1 shared only 26 % identity with GeIF7GT, 27 % with UGT73F2 and 63 % with GmIF7GT. The PlUGT1 gene was highly expressed in P. lobata roots relative to other organs and strongly induced by methyl jasmonate signal in P. lobata cell suspension culture. The transcript abundance of PlUGT1 was correlated with the accumulation pattern of isoflavone glycosides such as daidzin in P. lobata plants or in cell suspension culture. The biochemical properties and gene expression profile supported the idea that PlUGT1 could play a role in isoflavone glycosylation in P. lobata.
从野葛中分离出一种新型异黄酮7-O-葡萄糖基转移酶PlUGT1。PlUGT1能够将大豆苷元转化为大豆苷，染料木黄酮转化为染料木苷，以及将芒柄花素转化为芒柄花苷。传统上，野葛根被视为富含异黄酮糖苷的来源，这些异黄酮糖苷对人体健康有多种益处。然而，迄今为止，编码异黄酮UDP-糖基转移酶（UGT）的基因仅从大豆幼苗根（GmIF7GT）、大豆种子（UGT73F2）和光果甘草细胞悬浮培养物（GeIF7GT）中分离得到。为了研究野葛中的异黄酮代谢，从野葛中分离出40种UGT部分cDNA，并鉴定出7个在根中优先表达的全长UGT候选基因。在酵母（酿酒酵母）中的功能分析表明，这些UGT候选基因之一，命名为PlUGT1（官方UGT命名为UGT88E12），能够有效地将异黄酮苷元在7-羟基处糖基化。从大肠杆菌细胞中纯化的重组PlUGT1经鉴定对异黄酮苷元具有相对特异性，而对黄酮类底物的接受性较差。生化结果表明PlUGT1是一种异黄酮7-O-葡萄糖基转移酶。PlUGT1推导的氨基酸序列与GeIF7GT仅具有26%的同一性，与UGT73F2具有27%的同一性，与GmIF7GT具有63%的同一性。相对于其他器官，PlUGT1基因在野葛根中高度表达，并在野葛细胞悬浮培养中受到茉莉酸甲酯信号的强烈诱导。PlUGT1的转录丰度与野葛植物或细胞悬浮培养中大豆苷等异黄酮糖苷的积累模式相关。生化特性和基因表达谱支持了PlUGT1可能在野葛异黄酮糖基化中发挥作用的观点。
