Protection of hCTLA4Ig from Plant Proteases Using Anionic Polyelectrolytes in Transgenic Rice Cell Cultures

초록

Production of recombinant biopharmaceuticals in plant cells has emerged as an alternative to conventional platforms, and transgenic rice cell cultures provide an effective bridge between these platforms for the production of antibodies. Human cytotoxic T-lymphocyte antigen 4-immunoglobulin (hCTLA4Ig) was expressed using RAmy3D promoter. This system is regulated by sugar starvation and causes cell death. The cell death motivates the release of proteases which degrade the target protein. Polyelectrolytes as precipitants might be useful for selective partitioning of the protein and impurities. Isoelectric point, pH, polyelectrolyte concentration and molecular weight are key factors in precipitation. Poly vinylsulfonic acid (PVS), an anionic polyelectrolyte, was selected for acceptable precipitation of impurities. Various conditions of medium were evaluated at pH 3, 4, 5, 6 and 7, and PVS concentrations at 0.01%, 0.05%, 0.10%, 0.15%, and 0.20%. At pH 4, 0.01% PVS and pH 5, 0.10% PVS, protease activity was decreased by the precipitation of protease. Therefore, utilization of polyelectrolytes could be an efficient integrated process in cell culture platforms and enhance the productivity of target protein.