Engineering nonphosphorylative metabolism to synthesize mesaconate from lignocellulosic sugars in Escherichia coli
Engineering nonphosphorylative metabolism to synthesize mesaconate from lignocellulosic sugars in Escherichia coli
复制标题
DOI：
10.1016/j.ymben.2016.09.007
复制
发表时间：
2016-11-01
影响因子：
8.4
通讯作者：
Zhang, Kechun
中科院分区：
文献类型：
--
作者：
Bai, Wenqin;Tai, Yi-Shu;Zhang, Kechun
二羧酸是有吸引力的生物合成目标，由于其广泛的应用和其具有挑战性的制造过程从化石燃料原料。中康酸酯是一种支链的不饱和二羧酸，可用作共聚单体以生产水凝胶和阻燃材料。在这项研究中，我们设计了非磷酸化代谢，从D-木糖和L-阿拉伯糖生产中康酸。这种非磷酸化代谢与大肠杆菌中的固有戊糖代谢正交，并且具有比戊糖磷酸途径更少的酶促步骤和更高的TCA循环中间体的理论产率。在此，能够从D-木糖途径和L-阿拉伯糖途径产生中康酸盐。为了增强D-木糖和L-阿拉伯糖的转运，检查了戊糖转运蛋白。我们确定戊糖/质子同向转运体AraE是中康酸生产过程中D-木糖和L-阿拉伯糖最有效的转运体。通过操纵子筛选和代谢工程实现了进一步的生产优化。这些努力产生了在48小时后分别从20 g/l D-木糖和L-阿拉伯糖产生12.5 g/l和13.2 g/l中康酸的工程菌株。最后，过表达L-阿拉伯糖和D-木糖操纵子的工程菌株从1：1的D-木糖和L-阿拉伯糖混合物产生14.7g/l中康酸，产率为理论最大值的85%。(0.87 g/g）。这项工作证明了一个有效的系统，将戊糖转化为增值的化学品，mesaconate，有前途的滴度，速率和产量。
Dicarboxylic acids are attractive biosynthetic targets due to their broad applications and their challenging manufacturing process from fossil fuel feedstock. Mesaconate is a branched, unsaturated dicarboxylic acid that can be used as a co-monomer to produce hydrogels and fire-retardant materials. In this study, we engineered nonphosphorylative metabolism to produce mesaconate from D-xylose and L-arabinose. This nonphosphorylative metabolism is orthogonal to the intrinsic pentose metabolism in Escherichia call and has fewer enzymatic steps and a higher theoretical yield to TCA cycle intermediates than the pentose phosphate pathway. Here mesaconate production was enabled from the D-xylose pathway and the L-arabinose pathway. To enhance the transportation of D-xylose and L-arabinose, pentose transporters were examined. We identified the pentose/proton symporter, AraE, as the most effective transporter for both D-xylose and L-arabinose in mesaconate production process. Further production optimization was achieved by operon screening and metabolic engineering. These efforts led to the engineered strains that produced 12.5 g/l and 13.2 g/l mesaconate after 48 h from 20 g/l of D-xylose and L-arabinose, respectively. Finally, the engineered strain overexpressing both L-arabinose and D-xylose operons produced 14.7 g/l mesaconate from a 1:1 D-xylose and L-arabinose mixture with a yield of 85% of the theoretical maximum. (0.87 g/g). This work demonstrates an effective system that converts pentoses into a value-added chemical, mesaconate, with promising titer, rate, and yield.