Wang Aixia, Tian Wenzhe, Cheng Lei, Xu Youqiang, Wang Xiuwen, Qin Jiayang, Yu Bo
College of Pharmacy, Binzhou Medical University, Yantai, China.
Beijing Engineering and Technology Research Center of Food Additives, Beijing Technology and Business University (BTBU), Beijing, China.
Front Bioeng Biotechnol. 2020 Apr 22;8:288. doi: 10.3389/fbioe.2020.00288. eCollection 2020.
ε-Poly-L-lysine (ε-PL) is a natural amino acid polymer produced by microbial fermentation. It has been mainly used as a preservative in the food and cosmetics industries, as a drug carrier in medicines, and as a gene carrier in gene therapy. ε-PL synthase is the key enzyme responsible for the polymerization of L-lysine to form ε-PL. In this study, the ε-PL synthase gene was overexpressed in CICC 11022 by using the p promoter and the ribosome binding site from the capsid protein of phage ϕC31, which resulted in a genetically engineered strain Q-PL2. The titers of ε-PL produced by Q-PL2 were 88.2% ± 8.3% higher than that produced by the wild strain in shake flask fermentation. With the synergistic effect of 2 g/L sodium citrate, the titers of ε-PL produced by Q-PL2 were 211.2% ± 17.4% higher than that produced by the wild strain. In fed-batch fermentations, 20.1 ± 1.3 g/L of ε-PL was produced by Q-PL2 in 72 h with a productivity of 6.7 ± 0.4 g/L/day, which was 3.2 ± 0.3-fold of that produced by the wild strain. These results indicate that ε-PL synthase is one of the rate-limiting enzymes in ε-PL synthesis pathway and lays a foundation for further improving the ε-PL production ability of by metabolic engineering.
ε-聚-L-赖氨酸（ε-PL）是一种通过微生物发酵产生的天然氨基酸聚合物。它主要用作食品和化妆品行业的防腐剂、药物中的药物载体以及基因治疗中的基因载体。ε-PL合酶是负责将L-赖氨酸聚合成ε-PL的关键酶。在本研究中，通过使用来自噬菌体ϕC31衣壳蛋白的p启动子和核糖体结合位点，在CICC 11022中过表达ε-PL合酶基因，从而得到了基因工程菌株Q-PL2。在摇瓶发酵中，Q-PL2产生的ε-PL滴度比野生菌株产生的高88.2%±8.3%。在2 g/L柠檬酸钠的协同作用下，Q-PL2产生的ε-PL滴度比野生菌株产生的高211.2%±17.4%。在分批补料发酵中，Q-PL2在72小时内产生了20.1±1.3 g/L的ε-PL，生产力为6.7±0.4 g/L/天，是野生菌株产生量 的3.2±0.3倍。这些结果表明，ε-PL合酶是ε-PL合成途径中的限速酶之一，为通过代谢工程进一步提高ε-PL的生产能力奠定了基础。
