AIM： To study the baculovirus/mammalian cell system for efficient expression of functional large hepatitis delta antigen （L-HDAg）. METHODS： A recombinant baculovirus expressing histidine-tagged L-HDAg （L-HDAgH） ...AIM： To study the baculovirus/mammalian cell system for efficient expression of functional large hepatitis delta antigen （L-HDAg）. METHODS： A recombinant baculovirus expressing histidine-tagged L-HDAg （L-HDAgH） was constructed to transduce baby hamster kidney （BHK） cells by a simplified transduction protocol. RESULTS： The recombinant baculovirus transduced BHK cells with efficiencies higher than 90% as determined by flow cytometry. The expression level was significantly higher than that obtained by plasmid transfection and was further enhanced 3-fold to around 19 pg/cell by the addition of 10 mmol/L sodium butyrate. Importantly, the expressed L-HDAgH was localized to the cell nucleus and correctly isoprenylated as determined by immunofluorescence labeling and confocal microscopy. Moreover, L-HDAgH interacted with hepatitis B surface antigen to form virus-like particles. CONCLUSION： The fusion with histidine tags as well as overexpression of L-HDAgH in the baculovirus-transduced BHK cells does not impair the biological functions. Taken together, the baculovirus/mammalian cell system offers an attractive alternative for high level expression of L-HDAgH or other proteins that require extensive posttranslational modifications.展开更多
基金Supported by National Health Research Institutes (NHRI-EX94-9412EI) VTY Joint Research Program, Tsou's Foundation (VGHUST94-P6-32)
文摘AIM： To study the baculovirus/mammalian cell system for efficient expression of functional large hepatitis delta antigen （L-HDAg）. METHODS： A recombinant baculovirus expressing histidine-tagged L-HDAg （L-HDAgH） was constructed to transduce baby hamster kidney （BHK） cells by a simplified transduction protocol. RESULTS： The recombinant baculovirus transduced BHK cells with efficiencies higher than 90% as determined by flow cytometry. The expression level was significantly higher than that obtained by plasmid transfection and was further enhanced 3-fold to around 19 pg/cell by the addition of 10 mmol/L sodium butyrate. Importantly, the expressed L-HDAgH was localized to the cell nucleus and correctly isoprenylated as determined by immunofluorescence labeling and confocal microscopy. Moreover, L-HDAgH interacted with hepatitis B surface antigen to form virus-like particles. CONCLUSION： The fusion with histidine tags as well as overexpression of L-HDAgH in the baculovirus-transduced BHK cells does not impair the biological functions. Taken together, the baculovirus/mammalian cell system offers an attractive alternative for high level expression of L-HDAgH or other proteins that require extensive posttranslational modifications.
