title: Construction and characterization of a Salmonella typhi-based human immunodeficiency virus type 1 vector vaccine.
abstract: Since the human immunodeficiency virus type 1 (HIV-1) is transmitted either parenterally or sexually, both systemic and mucosal immune responses might be required to provide protective immunity. One option is to express HIV proteins in attenuated Salmonella vectors that elicit immune responses in both compartments. The first step to constructing such a strain was achieved by integrating a gene expression cassette encoding recombinant HIV-1 gp120 (rgp120) into the aroC locus of an attenuated vaccine strain of S. typhi. This rgp120 expression cassette utilizes the strong constitutive promoter, P1pp/lacUV5, and produces rgp120 to 0.05-01% of the total bacterial cell protein. Immunoblot analysis shows that the S. typhi strains containing the integrated cassette express a protein that is both recognized by anti-gp120 monoclonal antibodies (mAbs) and is the appropriate size for nonglycosylated full-length gp120 (52 kDa). Immunoblot analysis also demonstrates that the recombinant S. typhi strains express the rgp120 as monomers and multimers found predominantly in the insoluble fraction of the bacteria. Antigen-capture ELISA, using antibodies specific for continuous epitopes on gp120, revealed that the exposure of these epitopes on S. typhi-expressed rgp120 differs from exposure of these epitopes on baculovirus-expressed rgp120 that binds CD4. Epitopes in the first conserved region (109-113) and the third conserved/fourth variable regions (376-380, 382-384, 395-400) are more "surface-exposed", while one epitope in the third variable region (313-324) is more "buried" relative to the corresponding epitopes of baculovirus expressed gp120. Antibodies recognizing discontinuous epitopes of the CD4 binding domain do not react with the S. typhi expressed rgp120.(ABSTRACT TRUNCATED AT 250 WORDS)
mesh_terms: D016915:AIDS Vaccines / Q000235:genetics* / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology; D001483:Base Sequence; D003001:Cloning, Molecular; D000939:Epitopes / Q000276:immunology; D005822:Genetic Vectors; D015699:HIV Envelope Protein gp120 / Q000235:genetics* / Q000276:immunology*; D015497:HIV-1 / Q000276:immunology*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology; D012485:Salmonella typhi / Q000235:genetics* / Q000378:metabolism; D014613:Vaccines, Attenuated / Q000235:genetics / Q000276:immunology
chemical_list: D016915:AIDS Vaccines; D000939:Epitopes; D015699:HIV Envelope Protein gp120; D011994:Recombinant Proteins; D014613:Vaccines, Attenuated