Optimising fed-batch production of recombinant proteins using the baculovirus expression vector system.
Fed-batch culture can offer significant improvement in recombinant protein production compared to batch culture in the baculovirus expression vector system (BEVS), as shown by Nguyen et al. (1993) and Bedard et al. (1994) among others. However, a thorough analysis of fed-batch culture to determine its limits in improving recombinant protein production over batch culture has yet to be performed. In this work, this issue is addressed by the optimisation of single-addition fed-batch culture. This type of fed-batch culture involves the manual addition of a multi-component nutrient feed to batch culture before infection with the baculovirus. The nutrient feed consists of yeastolate ultrafiltrate, lipids, amino acids, vitamins, trace elements, and glucose, which were added to batch cultures of Spodoptera frugiperda (Sf9) cells before infection with a recombinant Autographa californica nuclear polyhedrosis virus (AcNPV) expressing beta-galactosidase (beta-Gal). The fed-batch production of beta-Gal was optimised using response surface methods (RSM). The optimisation was performed in two stages, starting with a screening procedure to determine the most important variables and ending with a central-composite experiment to obtain a response surface model of volumetric beta-Gal production. The predicted optimum volumetric yield of beta-Gal in fed-batch culture was 2.4-fold that of the best yields in batch culture. This result was confirmed by a statistical analysis of the best fed-batch and batch data (with average beta-Gal yields of 1.2 and 0.5 g/L, respectively) obtained from this laboratory. The response surface model generated can be used to design a more economical fed-batch operation, in which nutrient feed volumes are minimised while maintaining acceptable improvements in beta-Gal yield.
Chan
L C
LC
Department of Chemical Engineering, The University of Queensland, St Lucia, Qld 4072, Australia. e1lchan@mailbox.up.edu.au
Greenfield
P F
PF
Reid
S
S
D000818:Animals; D001709:Biotechnology / Q000379:methods; D018929:Cell Culture Techniques / Q000379:methods; D002460:Cell Line; D003470:Culture Media; D017924:Nucleopolyhedroviruses; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000378:metabolism; D018411:Spodoptera; D014162:Transfection; D001616:beta-Galactosidase / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism
D003470:Culture Media; D011994:Recombinant Proteins; D001616:beta-Galactosidase