Multiepitope polypeptide of the HIV-1 envelope induces neutralizing monoclonal antibodies against V3 loop.
A gene encoding a multiepitope polypeptide (MEP) has been synthesized. It contains the information for (1) an 11-amino acid (aa) epitope from the C1 region of gp120 of HIV-1 and (2) 3 epitopes of 15 amino acids each, from the central part of the V3 loop of isolates MN, SC, and WMJII. These four segments are linked by the short spacer peptide AGGGA. This gene was cloned in a plasmid vector and expressed in Escherichia coli as a fusion product with a 62-aa fragment of human IL-2. The recombinant protein TAB1 was purified by washed pellet procedures and reversed-phase HPLC. TAB1 was recognized in ELISAs by 25 of 27 sera from seropositive individuals. Mice were immunized and several hybridomas were obtained. Two of them secrete monoclonal antibodies that react with synthetic peptides from isolates MN, WMJI, WMJIII, and SC with an affinity constant in the range of 10(8) M-1. They also recognized peptides from isolates SF2 and WMJII, but at much lower affinity. The results obtained from peptide ELISAs indicate that the putative epitope recognized by these MAbs lies within the sequence IHIGPGRAFYT. Classic neutralization assays demonstrated that MAb 2C4 neutralizes 50% of the MN isolate at 0.6 micrograms/ml but fails to neutralize IIIB and SF2 strains. The presence of antibodies directed against every one of the component peptides in the sera of rabbits immunized with TAB1 was also documented.
Duarte
C A
CA
Center for Genetic Engineering and Biotechnology, Havana, Cuba.
Montero
M
M
Seralena
A
A
Valdés
R
R
Jiménez
V
V
Benítez
J
J
Narciandi
E
E
Madrazo
J
J
Padrón
G
G
Sánchez
G
G
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology*; D002460:Cell Line; D003001:Cloning, Molecular; D000939:Epitopes / Q000235:genetics / Q000276:immunology*; D005813:Genes, Synthetic; D015483:HIV Antibodies / Q000096:biosynthesis / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000235:genetics / Q000276:immunology*; D015497:HIV-1 / Q000235:genetics / Q000276:immunology*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D010452:Peptide Biosynthesis; D010446:Peptide Fragments / Q000235:genetics / Q000276:immunology*; D010455:Peptides / Q000276:immunology; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology
D000911:Antibodies, Monoclonal; D000939:Epitopes; D015483:HIV Antibodies; D015699:HIV Envelope Protein gp120; C059762:HIV envelope protein gp120 (305-321); D010446:Peptide Fragments; D010455:Peptides; D011993:Recombinant Fusion Proteins