Conformational display of two neutralizing epitopes of HIV-1 gp41 on the Flock House virus capsid protein.
We have developed an antigen presenting system based on the capsid protein of the Flock House virus (FHV) and used it to display, in different positions on its external surface, two neutralizing epitopes found at residues 735-752 of HIV-1 gp160. We have compared the immunoreactivity of these FHV chimeric proteins and of the corresponding synthetic peptide using a panel of neutralizing mouse monoclonal antibodies (mAbs) directed against two distinct sequences (IEEE and ERDRD) contained in this epitope of the gp41 region. We have observed that both the FHV chimeric protein and the synthetic peptide are clearly detected in ELISA procedures by the mAbs recognizing the sequence IEEE. The denaturation of these recombinant proteins had little effect on the recognition pattern of this group of monoclonals, suggesting minor conformational requirements for the display of this epitope. The FHV chimeric proteins were also recognized by the mAbs directed against the ERDRD epitope, whereas the corresponding synthetic peptide was not recognized. In this case, denaturation of these recombinant proteins completely abolished the reactivity of the second group of mAbs, arguing for the existence of strong conformational constraints. Additionally, we have investigated whether an isolated loop structure from the FHV protein was sufficient to provide the conformational requirements for the presentation of these epitopes. These experiments have shown that the stabilized loop structure, although improving the presentation of both epitopes, is not as efficient as the native loop in the intact FHV protein. The data obtained with these mAbs support the recently observed limitations in the use of synthetic peptides for the screening of the immune response against conformational epitopes. The establishment of appropriate tools able to present epitope sequences in a structure resembling the native conformation will be useful for accurate epidemiological studies and for the design of new epitope-specific vaccines.
Buratti
E
E
International Centre for Genetic Engineering and Biotechnology, Trieste, Italy.
Tisminetzky
S G
SG
Scodeller
E S
ES
Baralle
F E
FE
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001483:Base Sequence; D001667:Binding, Competitive; D018604:Epitope Mapping; D000939:Epitopes / Q000737:chemistry; D015483:HIV Antibodies / Q000276:immunology*; D015488:HIV Antigens / Q000276:immunology*; D015700:HIV Envelope Protein gp41 / Q000276:immunology*; D007304:Insect Viruses / Q000235:genetics / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins
D000911:Antibodies, Monoclonal; D000939:Epitopes; D015483:HIV Antibodies; D015488:HIV Antigens; D015700:HIV Envelope Protein gp41; D011993:Recombinant Fusion Proteins