Strain Engineering for Robust E. coli Fermentation Processes

Since the beginning of the biotechnology era in the late 70s, the pharmaceutical industry has extensively exploited E. coli for the production of recombinant proteins. High cell density (HCD) fermentations with E. coli can accumulate acetate, which impacts growth as well as recombinant protein production.  Consequently, processes accumulating high levels of acetate may not be robust. There have been numerous examples in recent years of engineering E. coli for useful applications owing to the advancements in the genetic tools and also the breadth of knowledge available on this organism. In this work, E. coli production strains were engineered by introducing mutations into the genes involved in the glycolysis and TCA cycle pathways to reduce the formation of acetate and, thus, improve process robustness and also product titer under certain “stress-test” HCD fermentation conditions, as well as more typical target fermentation conditions. In addition, the study results indicate that the strains engineered for decreased acetate accumulation might be more robust under multivariate (e.g., Design-Of-Experiment) conditions evaluated during process characterization studies. Consequently, processes utilizing these strains could potentially have wider acceptable process parameter ranges.