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202204004.pdf
文章编号： 1672-6987（2022）04-0027-06； DOI： 10.16351/j.1672-6987.2022.04.004
韩雪影a, 李露b*(青岛科技大学 a.海洋科学与生物工程学院；b.化工学院，山东 青岛 266042）
摘要： 柠檬酸因其广泛的应用而具有很高的经济潜力，它主要是以黑曲霉为宿主进行发酵生产，但黑曲霉生长缓慢，发酵时间长，需富氧发酵等特点为工业生产带来了许多不便。为了获得生长快、培养简便的高产柠檬酸菌株，以大肠杆菌为生产宿主，在大肠杆菌W(ATCC 9637)中过表达编码柠檬酸合酶的基因gltA和编码顺乌头酸脱羧酶的基因cad，所得的重组菌株使用M9培养基摇瓶发酵96 h产生245 mg·L-1的柠檬酸。通过单因素实验对碳源、初始糖含量、温度、pH、接种量和外源添加酵母膏等发酵条件进行优化。单因素实验结果表明，在蔗糖8 g·L-1、温度37 ℃、初始培养基pH 70、接种量4%和不添加外源酵母膏的条件下获得最高柠檬酸产量为1 443 mg·L-1。
关键词： 柠檬酸； 大肠杆菌； 代谢工程； 发酵条件优化
中图分类号： TQ 921文献标志码： A
引用格式： 韩雪影, 李露. 代谢工程改造大肠杆菌生产柠檬酸［J］. 青岛科技大学学报(自然科学版), 2022, 43(4): 27-32.
HAN Xueying，LI Lu. Metabolic engineering of Escherichia coli to produce citric acid［J］. Journal of Qingdao University of Science and Technology（Natural Science Edition), 2022， 43（4）： 27-32.
Metabolic Engineering of Escherichia Coli to Produce Citric Acid
HAN Xueyinga， LI Lub
(a. College of Mairine Science and Biological Engineering; b. College of Chemical Engineering,
Qingdao University of Science and Technology, Qingdao 266042, China)
Abstract: Citric acid has a high economic potential due to its wide application. It is mainly produced by Aspergillus niger as a host for fermentation. However, the characteristics of Aspergillus niger, such as slow growth, long fermentation time, and oxygen-enriched fermentation, have brought a lot of inconvenient for industrial production. In order to obtain a high-yield citrate strain with fast-growing and easy-to-cultivation, E. coli was used as the production host, overexpression of the gene gltA encoding citrate synthase and the gene cad encoding cis-aconitic acid decarboxylase in Escherichia coli W (ATCC 9637). The obtained recombinant strain was fermented with M9 medium in shake flask for 96 h to produce 245 mg·L-1 citric acid. The fermentation conditions, such as carbon source, initial sugar content, temperature, pH, inoculum amount, and exogenous addition of yeast extract, were optimized through single factor experiments. The results showed that the highest yield of citric acid was 1 443 mg·L-1 under the conditions of 8 g·L-1 sucrose, temperature 35 ℃ , pH 70, inoculum amount 4% and no exogenous yeast extract.
Key words: citric acid; Escherichia coli; metabolic engineering; optimization of fermentation conditions
收稿日期： 2021-08-04
作者简介： 韩雪影（1997—），女，硕士研究生.*通信联系人.
