【作者】 梁山泉； 江正强；

【Author】 Liang Shanquan;Jiang Zhengqiang;College of Food Science and Nutritional Engineering,China Agricultural University;Food Laboratory of Zhongyuan;

【机构】 中国农业大学食品科学与营养工程学院； 中原食品实验室；

【摘要】 二岩藻糖基乳糖(difucosyllactose,DFL)是一种含量丰富的人乳寡糖(human milk oligosaccharides,HMOs),具有促进婴幼儿生长和大脑发育等重要生理功能。首先在大肠杆菌BL21star(DE3)中构建了 DFL前体2’-岩藻糖基乳糖(2’-fucosyllactose,2’-FL)的从头合成通路。接着,优化关键酶α-1,2-岩藻糖基转移酶FutC表达和增加辅因子GDP和NADPH的供给提高2’-FL产量。随后,幽门螺杆菌(Helicobacter pylori,H.pylori)来源的α-1,3/4-岩藻糖基转移酶Hp3/4FT导入到重组菌株从而实现从头合成DFL。通过增加重组菌GDP-L-岩藻糖的积累和调控通路基因的表达水平增加DFL的产量。进一步强化2’-FL到DFL的转化和减少2’-FL的分泌减少中间产物的积累。最后,重组菌在5 L发酵罐中DFL的产量达55.3 g/L(中间产物仅为2.59 g/L),是迄今为止报道的DFL最高产量。本研究为高效合成DFL及其他人乳寡糖提供了重要的研究基础。更多还原

【Abstract】 Difucosyllactose(DFL) is an abundant human milk oligosaccharides(HMOs) and has significant benefits for the growth and brain development of infants.Firstly,the de novo pathway for the precursor 2’-fucosyllactose(2’-FL) of DFL was constructed in E.coli BL21star(DE3).Next,the biosynthesis of 2’-FL was increased by optimizing the expression of key α-1,2-fucosyltransferase FutCand increasing the supply of cofactors GDP and NADPH.Then,the α-1,3/4-fucosyltransferase Hp3/4FT from Helicobacter pylori was then introduced into the recombinant strain to achieve de novo biosynthesis of DFL.The biosynthesis of DFL was improved by increasing the accumulation of GDP-L-fucose and regulating the expression levels of pathway genes.Furthermore,the residual intermediate products were decreased by enhancing the conversion of2’-FL to DFL and reducing the efflux of 2’-FL.Finally,the recombinant strain produced 55.3 g/L(only 2.59 g/L of intermediate products) of DFL in a 5 L fermenter,which is the highest reported DFL titer so far.This study provided an important research foundation for the efficient synthesis of DFL and other HMOs.更多还原