Deletion of N-glycosylation sites of hepatitis C virus envelope protein E1 enhances specific cellular and humoral immune responses.

N-linked glycosylations of viral proteins have been implicated in immunogenicity. In this study, the effects of the N-linked glycosylation of the hepatitis C virus (HCV) E1 protein, a naturally poor immunogen, on the induction of specific immune response were examined. We constructed the plasmids containing the genes encoding both wild type and mutant E1 proteins in which N-linked glycosylation sites are mutated individually or in combination by site-directed mutagenesis. The immunogenicity of wild type E1 and six mutated E1 proteins was analyzed in BALB/C mice using a DNA-based vaccination approach. We found that E1-M2 mutant (at site of N209SS) significantly enhanced E1-specific CD8(+)T cells cytotoxic T lymphocytes (CTL) activities, expression of IFN-gamma producing T cells, and suppression of tumor growth. While E1-M4 mutant (at site of N305CS) induced the highest specific antibody response among all groups. Moreover, E1 wild-type vaccinated mice developed a mixture of IgG1 and Ig2a, but E1-M2 mutant induced only IgG2a isotype, and E1-M4 mutant dominantly developed IgG1 isotype. Our data showed that N-linked glycosylation can limit both cellular and antibody response to the HCV E1 protein and deletion of the N-glycosylation sites at N209SS and N305CS of hepatitis C virus envelope protein E1 provided potential applications for the development of DNA vaccine with enhanced immunogenicity.