Influence of controlled-pH and uncontrolled-pH operations on recombinant benzaldehyde lyase production by Escherichia coli

To select the host microorganism having the highest benzaldehyde lyase (BAL) production capacity, pUC18∷bal gene was transferred into four Escherichia coli strains. As the highest enzyme activity was obtained with E. coli K12 (ATCC 10798) carrying pUC18∷bal gene, BAL production medium was designed for K12. Using the designed medium containing 8.0 kg m−3 glucose, 5.0 kg m−3 (NH4)2HPO4 and the salt solution, the effects of uncontrolled-pH and controlled-pH operations were investigated at uncontrolled-pH pHUC 7.2 and controlled-pH values pHC 5.0, 6.4, 6.7, 7.0, 7.2 and 7.8 in 3.0 dm3 bioreactor systems with a VR = 1.65 dm3 working volume at the air inlet rate of Qo/VR = 0.5 vvm and agitation rate of N = 500 min−1. The uncontrolled-pH (pHUC 7.2) operation produced the highest cell concentration and BAL activity as CX = 2.3 kg m−3 and A = 860 U cm−3, respectively. Among the controlled-pH operations, the highest cell concentration and enzyme activity were obtained at pHC 7.0 operation, respectively, as CX = 2.1 kg m−3 and A = 775 U cm−3. The accumulation of the metabolic by-product acetic acid and the total organic acid concentrations were the highest at pHC 7.2 and 7.8 operations. On the other hand, there was no significant difference in the acetic acid concentration profiles of pHC 5.0, 6.4, 6.7 and 7.0 operations; however, the highest total organic acid concentration was attained at pHC 5.0 because of the lactic acid excretion, and the lowest total organic acid was obtained at pHC 6.7. KLa values varied between 0.01 and 0.03 s−1. To compare the mass transfer and biochemical reaction rates, the maximum possible oxygen utilization rate, possible mass transfer rate, Damköhler number, and effectiveness factor, were also calculated. Damköhler number increased with the cultivation time indicating that mass transfer resistances were becoming more effective than biochemical reaction resistances. The yield and maintenance coefficients as well as the kinetic constants for BAL production process were also reported.