文献类型：期刊文献

英文题名：Engineering Escherichia coli for l-homoserine production

作者：Sun, Bing-Yao[1];Wang, Feng-Qing[1];Zhao, Jian[1];Tao, Xin-Yi[1];Liu, Min[1];Wei, Dong-Zhi[1]

机构：[1]East China Univ Sci & Technol, Newworld Inst Biotechnol, State Key Lab Bioreactor Engn, 130 Meilong Rd, Shanghai 200237, Peoples R China

年份：2023

卷号：63

期号：2

起止页码：168

外文期刊名：JOURNAL OF BASIC MICROBIOLOGY

收录：;WOS:【SCI-EXPANDED(收录号:WOS:000871788100001)】；

基金：National Natural Science Foundation of China, Grant/Award Number: 31500043; National Key Research and Development Program of China, Grant/Award Number: 2021YFC2100300; Natural Science Foundation of Shanghai, Grant/Award Number: 21ZR1417200; Open Funding Project of the State Key Laboratory of Bioreactor Engineering

语种：英文

外文关键词：Escherichia coli; hok; sok system; l-homoserine; metabolic engineering; plasmid stability

摘要：l-homoserine, a nonprotein amino acid, is used to synthesize many active substances in the industry. Here, to develop a robust l-homoserine-producing strain, Escherichia coli W3110 was used as a chassis to be engineered. Based on a previous construct with blocked competing routes for l-homoserine synthesis, five genes were overexpressed by promoter replacement strategy to increase the l-homoserine production, including enhancement of precursors for l-homoserine synthesis (ppc, thrA, and asd), reinforcement of the NADPH supply (pntAB) and efflux transporters (rhtA) to improve the l-homoserine production. However, the plasmid losing was to blame for the wildly fluctuating fermentation performance of engineered strains, ranging between 2.1 and 6.2 g/L. Then, a hok/sok toxin/antitoxin system was introduced into the free plasmid expression cassette to maintain the genetic stability of the episomal plasmid; consequently, the plasmid-losing rate sharply decreased, resulting in the engineered strain SHL17, which exhibited excellent stability in l-homoserine production, with 6.3 g/L in shake flasks and 44.4 g/L in a 5-L fermenter without antibiotic addition. This work verified the effective use of the hok/sok toxin/antitoxin system combined with promoter engineering to improve the genetic stability of E. coli episomal plasmids without antibiotics.