EXPRESSION AND IMMUNOGENICITY OF THE V3 LOOP FROM THE ENVELOPE OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 IN AN ATTENUATED AROA STRAIN OF SALMONELLA-TYPHIMURIUM UPON GENETIC COUPLING TO 2 ESCHERICHIA-COLI CARRIER PROTEINS

A peptide comprising residues glu293 to ser334 from the principal neut ralization determinant (V3 loop) of the envelope of human immunodefici ency virus type 1 (HIV1 LAV(BRU) isolate) has been inserted within int ernal permissive sites of either LamB or MalE, two envelope proteins f rom Escherichia coli K12. The MalE hybrid protein (MalE133-V3 loop) wa s stably expresssed in the periplasm of Escherichia coli K12, and the V3 loop peptide was detectable on the surface of the native protein by an anti-gp160 monoclonal antibody (mAb 110-A). The disulfide bridge b etween the two cysteines of the loop was formed. In contrast, genetic coupling to the outer membrane protein LamB did not allow the expressi on of a stable hybrid protein, and major proteolytic cleavage products of the LamB153-V3 loop were detected by mAb 110-A. The two plasmid-en coded hybrid genes were transferred to an aroA mutant of Salmonella ty phimurium. Constitutive expression of the MalE133-V3 loop had no detec table effect on cell growth and on the survival in vivo of the recipie nt strain. The LamB153-V3 loop was not stably expressed in Salmonella, either in vitro or in vivo. Live recombinant salmonellas expressing M alE-V3 and LamB-V3 loop hybrids were used to immunize mice. The MalE-V 3 loop hybrid induced anti-HIV1 envelope antibodies detectable by West ern blot and ELISA, while the anti-HIV1 envelope antibodies induced by the LamB- V3 loop hybrid were only detectable by Western blot. In add ition, purified MalE-V3 loop hybrid protein was able to stimulate in v itro and induce in vivo a V3 loop-specific T-cell proliferative respon se.