C-terminal domain of hepatitis C virus core protein is essential for secretion

Figure 1 Core proteins are secreted into the culture medium in insect cells. A: Expression of HCV core protein in insect cells. cDNA corresponding to the HCV core was subcloned into the BamHI site of the transfer vector pVL941 behind polyhedrin promoter. Recombinant baculoviruses expressing HCV core protein were produced as described in Materials and methods. Sf9 cells were infected with either wild type (AcNPV) or recombinant baculoviruses expressing SHDAg, or recombinant baculoviruses expressing HCV core protein and were harvested at d 3 postinfection. Cell lysates were separated by SDS-containing polyacrylamide gel electrophoresis, transferred to a nitrocellulose membrane. Proteins were detected by Western blotting using HCV patient sera. Lane 1, Sf9 cells infected with wild type baculoviruses; lane 2, Sf9 cells infected with recombinant baculovirus expressing SHDAg; lane 3, Sf9 cells infected with recombinant baculovirus expressing HCV core; B: The culture medium from (A) was centrifuged at 3 500 r/min for 15 min to remove cell debris. Supernatant was further centrifuged at 12 000 g for 30 min to remove baculoviruses. The resultant supernatant was then pelleted through a 300 g/L sucrose cushion for 90 min at 27 000 g. The pellet was dissolved in sample buffer and analyzed by Western blotting using HCV patient sera.