Optimization of a cultivation process for recombinant protein production by Escherichia coli.
A single-stage fed-batch bioprocess for the production of a recombinant protein beta-galactosidase, by E. coli has been developed. The XL1-blue strain of E. coli which harbors a multi-number foreign plasmid PT was cultured in a reformulated medium. Critical medium components were selected and their respective concentrations were optimized with the Orthogonal Table method. An exponential substrate feeding schedule was used to maintain optimum conditions. Inhibition of growth and protein expression caused by excessive concentrations of glucose and acetate was investigated and subsequently minimized with an incremental nutrient feeding schedule which limited the specific growth rate of a culture. The program necessary to facilitate the control of substrate addition is fully described. This program has been used with a 2.5 l bioreactor and a commercially available software package for optimization without on-line or off-line measurement of optical density (OD), CO2, glucose or acetate. The optimized fed-batch process limited the acetate concentration to less than 20 mM; maintained an exponential growth phase for 50 h; and produced a cell density of 51 g l-1 dry cell weight (DCW) or 154 OD600 with a beta-galactosidase activity of 990 U ml-1.
R&D Lab, New Brunswick Scientific Co. Inc., Edison, New Jersey 08818-4005.
X M
XM
Yang
D000085:Acetates / Q000378:metabolism; D001709:Biotechnology / Q000379:methods; D003470:Culture Media; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development* / Q000378:metabolism; D007700:Kinetics; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D001616:beta-Galactosidase / Q000235:genetics
D000085:Acetates; D003470:Culture Media; D011994:Recombinant Proteins; D001616:beta-Galactosidase