A 2-STAGE BIOREACTOR SYSTEM FOR THE PRODUCTION OF RECOMBINANT PROTEINS USING A GENETICALLY-ENGINEERED BACULOVIRUS-INSECT CELL SYSTEM

A two-stage bioreactor scheme was developed for the large-scale produc tion of recombinant proteins using a genetically engineered baculoviru s/insect cell system. The first bioreactor was employed for cell growt h and the second for cell infection. Silkworm Bm5 cells were infected with a recombinant baculovirus, BmNPV/P5.cat, containing a bacterial c hloramphenicol acetyltransferase (CAT) gene under the control of the p olyhedrin gene promoter of Bombyx mori nuclear polyhedrosis virus (BmN PV). This recombinant baculovirus has been used as an expression vecto r for the production of recombinant CAT enzyme. A specific productivit y of 82 to 90 mug CAT/(10(6) cells) was obtained using the BmNPV/Bm5 e xpression system, a yield similar to that achieved using the AcNPV/Sf expression system. Repeated infection of high-density cell cultures di d not reduce the specific productivity of the CAT enzyme. Most importa ntly, the problems associated with the infection of high-density cell cultures were resolved by means of controlled infection conditions and appropriate replenishment of spent culture medium following infection . The glucose uptake rate by the cells following infection was 50% hig her than that by the cells before infection. Not only did the infectio n of high-density cell cultures result in consistent yields of 250 mg/ L of CAT enzyme, but also the two-stage bioreactor system was proven t o be reliable for a long-term operation beyond 600 h.