Enhancement of human erythropoietin production in Chinese hamster ovary cells through supplementation of 30Kc19-30Kc6 fusion protein

Abstract

The importance of anti-apoptosis in mammalian cell culture has been widely recognized. We reported previously that expression of Bombyx mori 30K genes in Chinese hamster ovary (CHO) cells increases recombinant protein production by inhibiting apoptosis and enhancing specific productivity. However, previous studies have shown expression of the anti-apoptotic 30Kc6 protein as inclusion bodies in Escherichia coli. 30Kc19 protein, another silkworm hemolymph protein, has cell-penetrating and recombinant protein productivity-improving properties, and we found that it improves soluble expression of its partner. In this study, we fused 30Kc6 with 30Kc19 as an expression partner to express a soluble fused protein and to deliver the protein to cells. Supplementing the recombinant 30Kc19-30Kc6 fusion protein in cell culture medium increased viability, effectively penetrated the cells, and inhibited CHO cell apoptosis by 68%. Moreover, the mitochondrial membrane potential and ATP generation also increased by 50% and 33%, respectively. Erythropoietin (EPO) productivity increased by>30% because of the anti-apoptotic effect and increased specific productivity. These results demonstrate the potential use of this fusion protein as a supplement in mammalian cell culture during production of biopharmaceutical proteins.