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 two Escherichia coli carrier proteins.
A peptide comprising residues glu293 to ser334 from the principal neutralization determinant (V3 loop) of the envelope of human immunodeficiency virus type 1 (HIV1 LAVBRU isolate) has been inserted within internal permissive sites of either LamB or MalE, two envelope proteins from Escherichia coli K12. The MalE hybrid protein (MalE133-V3 loop) was stably expressed 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 between the two cysteines of the loop was formed. In contrast, genetic coupling to the outer membrane protein LamB did not allow the expression of a stable hybrid protein, and major proteolytic cleavage products of the LamB153-V3 loop were detected by mAb 110-A. The two plasmid-encoded hybrid genes were transferred to an aroA mutant of Salmonella typhimurium. Constitutive expression of the MalE133-V3 loop had no detectable effect on cell growth and on the survival in vivo of the recipient strain. The LamB153-V3 loop was not stably expressed in Salmonella, either in vitro or in vivo. Live recombinant salmonellas expressing MalE-V3 and LamB-V3 loop hybrids were used to immunize mice. The MalE-V3 loop hybrid induced anti-HIV1 envelope antibodies detectable by Western blot and ELISA, while the anti-HIV1 envelope antibodies induced by the LamB-V3 loop hybrid were only detectable by Western blot. In addition, purified MalE-V3 loop hybrid protein was able to stimulate in vitro and induce in vivo a V3 loop-specific T-cell proliferative response.
Charbit
A
A
Programmation Moleculaire et Toxicologie Génétique (CNRS URA 1444), Institut Pasteur, Paris, France.
Martineau
P
P
Ronco
J
J
Leclerc
C
C
Lo-Man
R
R
Michel
V
V
O'Callaghan
D
D
Hofnung
M
M
D016915:AIDS Vaccines / Q000235:genetics / Q000276:immunology / Q000494:pharmacology; D018528:ATP-Binding Cassette Transporters; D000595:Amino Acid Sequence; D000818:Animals; D000917:Antibody Formation / Q000187:drug effects / Q000276:immunology; D001425:Bacterial Outer Membrane Proteins; D001483:Base Sequence; D002352:Carrier Proteins / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D015686:Gene Products, env / Q000235:genetics* / Q000276:immunology*; D019302:HIV Envelope Protein gp160; D015497:HIV-1 / Q000235:genetics* / Q000276:immunology*; D008213:Lymphocyte Activation / Q000187:drug effects / Q000276:immunology; D058619:Maltose-Binding Proteins; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins; D033902:Periplasmic Binding Proteins; D018272:Porins; D011498:Protein Precursors / Q000235:genetics* / Q000276:immunology*; D011991:Receptors, Virus / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D012486:Salmonella typhimurium / Q000235:genetics*; D012680:Sensitivity and Specificity; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology
D016915:AIDS Vaccines; D018528:ATP-Binding Cassette Transporters; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D015686:Gene Products, env; D019302:HIV Envelope Protein gp160; C461619:MalE protein, E coli; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D033902:Periplasmic Binding Proteins; D018272:Porins; D011498:Protein Precursors; D011991:Receptors, Virus; D011993:Recombinant Fusion Proteins; C016532:maltoporins; C461599:maltose transport system, E coli