National Engineering Laboratory for Cereal Fermentation Technology, Jiangnan University, Wuxi, China.
The Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, China.
J Appl Microbiol. 2019 Apr;126(4):1128-1139. doi: 10.1111/jam.14204. Epub 2019 Feb 27.
To establish the biotechnology platforms for production of bio-based chemicals in various micro-organisms is considered as a promising target to improve renewable production of isoprene.
In this study, we heterologously expressed the mevalonate (MVA) isoprene biosynthesis pathway, and explored three strategies of increasing isoprene production in Escherichia coli. We first manipulated the expression levels of the MVA pathway genes through changing the gene cassettes and promoters. To introduce cofactor engineering, we then overexpressed NADP-dependent glyceraldehyde-3-phosphate dehydrogenase gene from Clostridium acetobutylicum to supply available NADPH. To reduce the inhibitory by-product accumulation, we finally knocked out acetate-producing genes, phosphate acetyl transferase and pyruvate oxidase B in E. coliJM109 (DE3), decreasing acetate accumulation 89% and increasing isoprene production 39%. The strategies described here finally increased the isoprene titre to 92 mg l in two-gene deletion strain JMAB-4T7P1Trc, increasing 2·6-fold comparing to strain JM7T7.
The multimodularly engineering approaches including promoter engineering, cofactor engineering and by-product reducing could be used to improve isoprene production in E. coli.
The metabolic strategies in this study show us directions for further studies to promote transformation of renewable sources to isoprene.
在各种微生物中建立生产生物基化学品的生物技术平台被认为是提高异戊二烯可再生生产的有前途的目标。
在本研究中，我们异源表达了甲羟戊酸（MVA）异戊二烯生物合成途径，并探索了提高大肠杆菌中异戊二烯产量的三种策略。我们首先通过改变基因盒和启动子来操纵 MVA 途径基因的表达水平。为了引入辅因子工程，我们过量表达了来自丙酮丁醇梭菌的 NADP 依赖性甘油醛-3-磷酸脱氢酶基因，以提供可用的 NADPH。为了减少抑制性副产物的积累，我们最终敲除了大肠杆菌 JM109（DE3）中的乙酸产生基因磷酸乙酰转移酶和丙酮酸氧化酶 B，使乙酸积累减少 89%，异戊二烯产量增加 39%。这里描述的策略最终使两基因缺失菌株 JMAB-4T7P1Trc 的异戊二烯产量达到 92mg/L，比菌株 JM7T7 增加了 2.6 倍。
包括启动子工程、辅因子工程和减少副产物在内的多模块工程方法可用于提高大肠杆菌中的异戊二烯产量。
本研究中的代谢策略为进一步研究利用可再生资源转化为异戊二烯提供了方向。
