Glucose feeding strategy accounting for the decreasing oxidative capacity of recombinant Escherichia coli in fed-batch cultivation for phenylalanine production

Abstract

The glucose feed rate is known to be a key factor in the control of various fed-batch processes. It plays a special role in the cultivation of Escherichia coli strains because of their inherent ability to readily excrete inhibitory metabolites, particularly acetic acid, when an inappropriate feeding strategy is adopted. Here we present the results of our study on the peculiarities of acetic acid excretion by recombinant Escherichia coli AT2471 in fed-batch cultivation for phenylalanine production. In addition to confirming that under aerobic conditions acetic acid excretion is a function of the glucose feeding strategy, we found that the excretion is closely related to the decreasing oxidative capacity of the cell population. This is clearly expressed by a declining dynamical profile of the so called “critical specific glucose uptake rate”, which discriminates the process field into distinct regions. We have concluded that the actual specific feed rate should always be dynamically adjusted below the critical limits, otherwise the oxidative capabilities of the population will be exceeded and acetate will be excreted. To provide the required time-course of the specific glucose feed rate, a closed-loop control algorithm was developed which utilizes a useful empirical relationship. Its application resulted in a suitable profile of the specific glucose feed rate, prevention of acetic acid excretion and improved phenylalanine synthesis, the final concentration of which reached 46 g/ l.