磷酸盐精确控制策略提升大肠杆菌生长和产物合成效率

Precise control strategy of phosphate to enhance the cell growth and the efficiency of product synthesis in Escherichia coli

磷酸盐是微生物生长代谢关键物质之一,研究发酵进程中的磷酸盐变化与变化规律及其与特定目标产物合成效率间相互关系,有助于提升微生物的生长和产物合成效率,建立新的发酵控制模式。本文以工业上重要的大肠杆菌为研究对象,研究了发酵过程中磷酸盐含量变化规律,解析了大肠杆菌生长和产酶过程与磷酸盐消耗间的关系,并通过调控磷酸盐浓度提高产酶水平。通过摇瓶发酵实验,确定了大肠杆菌的磷酸盐耐受上限为磷酸氢二铵24.0 g/L、磷酸二氢钾81.0 g/L,磷酸氢二铵和磷酸二氢钾最适初始含量分别为1.0 g/L、10.12 g/L,补加方式为:发酵培养基中补加总量的一半,剩余部分发酵4 h后补加。在此基础上,大肠杆菌细胞密度和赖氨酸脱羧酶的比酶活分别提高了58.42%和14.09%。研究结果证实通过动态监控并精确控制发酵过程中磷酸盐的存在量可以有效的提升大肠杆菌的生长代谢和产物合成效率。

Phosphate is one of the key materials for microbial growth and product synthesis.Thus,systematic studies of the changes of phosphate,its change rule and the corresponding relationships between the change rule and the synthetic efficiency of target products help to improve the growth of microorganisms and the efficiency of product synthesis.This will also establish a novel control mode of fermentation processes.In this paper,industrially important Escherichia coli was used as a research object to investigate the change rule of phosphate in the fermentation process as well as analyze the corresponding relationships between the growth of Escherichia coli,enzyme production and phosphate consumption.Subsequently,the production level of lysine decarboxylase was increased by controlling the concentrations of phosphate. Through shake flask fermentation,the highest concentrations of diammonium hydrogen phosphate and potassium dihydrogen phosphate that Escherichia coli cells could tolerate were determined to be 24.00 g / L and 81.00 g / L,respectively.The optimum initial contents of diammonium hydrogen phosphate and potassium dihydrogen phosphate were 1.00 g / L and 10.12 g / L,respectively. Besides,only 1 /2 of the total amounts of diammonium hydrogen phosphate and potassium dihydrogen phosphate was added at the beginning,while the remaining 1 /2 was complemented after 4 hours of fermentation. After cultivated under the optimized conditions,the cell density of recombinant Escherichia coli and the specific activity of lysine decarboxylase were increased by 58.42% and 14.09%,respectively. According to the result,the cell growth and products synthesis of E. coli can be improved by dynamic determination and precise control of the phosphate concentration during the fermentation process.