RESUMO

The recombinant G glycoprotein from the surface of the rabies virus (RVGP) is a promising candidate as a rabies vaccine component and also for diagnostic purposes. In this study, RVGP production by transfected Drosophila melanogaster S2 cells cultivated in a serum-free medium (supplemented IPL-41 medium) was carried out. The effects of pH and pO(2) were evaluated in batch culture in parallel spinner flasks. The use of a pH equal to 6.3 and a pO(2) of 40% air saturation resulted in the highest RVGP content. These conditions were also used in fed-batch mode, yielding a RVGP content level of 98g/10(7) cells. The main nutrients consumed were glucose, glutamine, asparagine, serine and proline and the major metabolites produced were alanine and ammonia, according to the metabolism studies performed. Since RVGP is a transmembrane protein, two different methods for protein recovery were assessed and compared. Detergent-based cell disruption showed to be more effective than mechanical disruption with glass beads for glycoprotein recovery.

Assuntos

Linhagem Celular , Meios de Cultura Livres de Soro/farmacologia , Drosophila melanogaster , Proteínas Recombinantes/metabolismo , Proteínas Virais de Fusão/isolamento & purificação , Proteínas Virais de Fusão/metabolismo , Aminoácidos/metabolismo , Animais , Metabolismo dos Carboidratos/efeitos dos fármacos , Técnicas de Cultura de Células , Linhagem Celular/metabolismo , Linhagem Celular/fisiologia , Proliferação de Células/efeitos dos fármacos , Concentração de Íons de Hidrogênio , Ácido Láctico/metabolismo , Consumo de Oxigênio/efeitos dos fármacos , Vírus da Raiva/genética , Vírus da Raiva/metabolismo , Proteínas Recombinantes/genética , Fatores de Tempo