Optimal IgG production in mammalian cells requires a heavy chain translation efficiency of approximately 50% relative to light chain translation.
Immunoglobulin G (IgG) is a heterotetrameric protein assembled from two identical heavy chain (HC) and two identical light chain (LC) polypeptides. The HC and LC folding and assembly are a crucial step for IgG production. It is affected by the ratio of HC to LC expression (HC:LC). To date, the HC:LC ratio was analysed mainly by cotransfection of different amounts of two monocistronic HC and LC expression plasmids, an approach biased by different transfection efficiencies. To circumvent this problem, a series of Encephalomyocarditis virus internal ribosome entry site (EMCV IRES) variants with different translation efficiencies were created and used to mediate HC translation in bicistronic constructs. HC and LC were translated from the same mRNA, which provides a more accurate method for the evaluation of the optimal ratio of HC:LC. The results show that the IgG optimal expression levels were obtained when the IRES mediated translation efficiency of the HC was about 50% compared to the cap-dependent translation of the LC. A surprisingly sharp transition to low production was shown when the ratios were below 40%. This study provides a new method to investigate the production of heterodimeric proteins in mammalian cells and adds understanding to the mechanisms of IgG folding and assembly.
Li et al. (Sat,) studied this question.