[High cell density culture of phosphotransacetylase mutants of Escherichia coli BL21(DE3)]

Abstract

Cell culture, organic acid production and foreign protein (TNF) expression of E. coli BL21(DE3) and its pta mutant were investigated. Under shaking conditions, TNF expression in pta mutant increased by 23%. During the fed-batch culture without limitation of specific growth rates, the mutant reached a cell density as high as 32.5 g(DCW)/L and total TNF expression at 2.8 g/L, while the parental strain only obtained 19.5 g(DCW)/L and 0.84 g/L. The results indicate that utility of pta mutant as a host is advantageous in foreign protein expression and high cell density culture. Meanwhile, the analysis data of organic acids accumulated during fed-batch culture showed that as the decrease of acetate production(42% of the parental strain), the accumulation of other organic acids(mainly pyruvate, lactate and succinate) obviously increased. As a result, the amount of total organic acids increased by 123% over its parent. The lactate production may be the main obstacle in further growth of the cells.