Zhang Ye, Sun Qing, Liu Yu, Cen Xuecong, Liu Dehua, Chen Zhen
Key Laboratory of Industrial Biocatalysis (Ministry of Education), Department of Chemical Engineering, Tsinghua University, Beijing, 100084, China.
Tsinghua Innovation Center in Dongguan, Dongguan, 523808, China.
Bioresour Bioprocess. 2021 Dec 14;8(1):125. doi: 10.1186/s40643-021-00485-0.
Vibrio natriegens is a promising industrial chassis with a super-fast growth rate and high substrate uptake rates. V. natriegens was previously engineered to produce 1,3-propanediol (1,3-PDO) from glycerol by overexpressing the corresponding genes in a plasmid. However, antibiotic selection pressure for plasmid stability was not satisfactory and plasmid loss resulted in reduced productivity of the bioprocess. In this study, we developed an antibiotic-free plasmid stabilization system for V. natriegens. The system was achieved by shifting the glpD gene, one of the essential genes for glycerol degradation, from the chromosome to plasmid. With this system, engineered V. natriegens can stably maintain a large expression plasmid during the whole fed-batch fermentation and accumulated 69.5 g/L 1,3-PDO in 24 h, which was 23% higher than that based on antibiotic selection system. This system was also applied to engineering V. natriegens for the production of 3-hydroxypropionate (3-HP), enabling the engineered strain to accumulate 64.5 g/L 3-HP in 24 h, which was 30% higher than that based on antibiotic system. Overall, the developed strategy could be useful for engineering V. natriegens as a platform for the production of value-added chemicals from glycerol.
嗜盐栖热袍菌是一种很有前景的工业底盘细胞，具有超快的生长速率和高底物摄取率。此前已通过在质粒中过表达相应基因，对嗜盐栖热袍菌进行工程改造，使其能从甘油生产1,3 - 丙二醇（1,3 - PDO）。然而，用于质粒稳定性的抗生素选择压力并不理想，质粒丢失导致生物过程的生产力下降。在本研究中，我们为嗜盐栖热袍菌开发了一种无抗生素的质粒稳定系统。该系统是通过将甘油降解必需基因之一的glpD基因从染色体转移到质粒上来实现的。利用该系统，工程化的嗜盐栖热袍菌在整个补料分批发酵过程中能够稳定维持一个大表达质粒，并在24小时内积累了69.5 g/L的1,3 - PDO，比基于抗生素选择系统的产量高23%。该系统还应用于工程化嗜盐栖热袍菌以生产3 - 羟基丙酸（3 - HP），使工程菌株在24小时内积累了64.5 g/L的3 - HP，比基于抗生素系统的产量高30%。总体而言，所开发的策略对于将嗜盐栖热袍菌工程改造为从甘油生产增值化学品的平台可能是有用的。
