Effects of Aeration and Agitation on the Stability of hCTLA4Ig in Transgenic Rice Cell Cultures

초록

Human cytotoxic T-lymphocyte-associated antigen 4-immunoglobulin (hCTLA4Ig), an immunosuppressive agent, was expressed in transgenic rice cells using RAmy3D promoter for the inducible production and secretion into culture media by sugar depletion. Aeration and agitation have been known as key parameters that affect productivity and quality of the secreted proteins in bioreactor. On the other hand, several reports have demonstrated that the degradation of foreign protein occurs in hydrodynamic environment related to agitation and aeration. In plant cell cultures, degradation of foreign protein by proteases has been widely known. However, the clear mechanism for the degradation of secreted protein in the medium of plant cell cultures remains unclear. In this study, we investigated the effects of aeration and agitation on the degradation of recombinant protein in scale-down model. In a previous study, degradation of target protein was confirmed when the culture conditions were 200 rpm and 0.3 vvm in 3-L bioreactor. To test the stability in a 100-mL bottle, hCTLA4Ig (20 mg/L) was added and working volume was 50 mL (AA medium and PBS). The rate of aeration and agitation were determined on the basis of the constant scale-up criteria such as tip speed and volumetric aeration.