MODELING ASSEMBLY, AGGREGATION, AND CHAPERONING OF IMMUNOGLOBULIN-G PRODUCTION IN INSECT CELLS

A model for immunoglobulin G (IgG) production in the baculovirus-insec t cell system was developed that incorporates polypeptide synthesis, o ligomer assembly, protein aggregation, and protein secretion. In addit ion, the capacity of a chaperone to protect heavy and light chain poly peptides from protein aggregation was considered by including in vitro chaperone-peptide binding and dissociation kinetic constants from the literature. Model predictions were then compared to experiments in wh ich the chaperone immunoglobulin heavy chain binding protein, BiP, was coexpressed by coinfecting insect cells with BiP-containing baculovir us. The model predicted a nearly twofold increase in intracellular and secreted IgG that was similar to the behavior observed experimentally after approximately 3 days of coexpressing heterologous IgG and BiP. However, immunoglobulin aggregation was still significant in both the model simulation and experiments, so the model was then used to predic t the effect of strategies for improving IgG production even further. Increasing expression of the chaperone BiP by 10-fold over current exp erimental levels provided a 2.5-fold increase in secreted IgG producti on over IgG assembly without BiP. Alternatively, the expression of BiP earlier in the baculovirus infection cycle achieved a twofold increas e in protein secretion without requiring excessive BiP production. The potential effect of cochaperones on BiP activity was considered by va rying the BiP binding and release constants. The utilization of lower binding and release kinetic constants led to a severalfold increase in IgG secretion because the polypeptides were protected from aggregatio n for greater periods. An optimized strategy for chaperone action woul d include the rapid peptide binding of a BiP-ATP conformation along wi th the slow peptide release of a BiP-ligand conformation. However, eve n with an optimized chaperoning system, limitations in the secretion k inetics can result in the accumulation of intracellular IgG. Thus, the entire secretor/pathway must be considered when enhanced secretion of heterologous proteins is desired. (C) 1997 John Wiley & Sons, Inc.