Hanah Margalit

Protein Antigenic Structures Recognized by T Cells; Potential Applications to Vaccine Design

In summary, our results using the model protein antigen myoglobin indicated, in concordance with others, that helper T lymphocytes recognize a limited number of immunodominant antigenic sites of any given protein. Such immunodominant sites are the focus of a polyclonal response of a number of different T cells specific for distinct but overlapping epitopes. Therefore, the immunodominance does not depend on the fine specificity of any given clone of T cells, but rather on other factors, either intrinsic or extrinsic to the structure of the antigen. A major extrinsic factor is the MHC of the responding individual, probably due to a requirement for the immunodominant peptides to bind to the MHC of presenting cells in that individual. In looking for intrinsic factors, we noted that both immunodominant sites of myoglobin were amphipathic helices, i.e., helices having hydrophilic and hydrophobic residues on opposite sides. Studies with synthetic peptides indicated that residues on the hydrophilic side were necessary for T-cell recognition. However, unfolding of the native protein was shown to be the apparent goal of processing of antigen, presumably to expose something not already exposed on the native molecule, such as the hydrophobic sides of these helices. We propose that such exposure is necessary to interact with something on the presenting cell, such as MHC or membrane, where we have demonstrated the presence of antigenic peptides by blocking of presentation of biotinylated peptide with avidin. The membrane may serve as a short-term memory of peptides from antigens encountered by the presenting cell, for dynamic sampling by MHC molecules to be available for presentation to T cells. These ideas, together with the knowledge that T-cell recognition required only short peptides and therefore had to be based only on primary or secondary structure, not tertiary folding of the native protein, led us to propose that T-cell immunodominant epitopes may tend to be amphipathic structures. An algorithm to search for potential amphipathic helices from sequence information identified 18 of 23 known immunodominant T-cell epitopes from 12 proteins (p less than 0.001). Another statistical approach confirmed the importance of amphipathicity and also supported the importance of helical structure that had been proposed by others. It suggested that peptides able to form a stable secondary structure, especially a helix, more commonly formed immunodominant epitopes. We used this approach to predict potential immunodominant epitopes for induction of T-cell immunity in proteins of clinical relevance, such as the malarial circumsporozoite protein and the AIDS viral envelope.(ABSTRACT TRUNCATED AT 400 WORDS)

Repeated determinants within the retinal interphotoreceptor retinoid-binding protein (IRBP): Immunological properties of the repeats of an immunodominant determinant

Interphotoreceptor retinoid-binding protein (IRBP), a glycoprotein which localizes in the retina and pineal gland, induces inflammatory changes in these organs (EAU and EAP, respectively) when injected into various mammals. We have previously identified a determinant (residues 1169-1191) in bovine IRBP which is immunodominant and highly immunogenic and immunopathogenic in Lewis rats. IRBP exhibits a fourfold repeat structure and we report here on the comparison between the active sequence 1179-1191 and its three repeat peptides. Only one of the repeats, 271-283, cross-reacted with 1179-1191 and exhibited immunodominance, albeit of a low level. Peptide 271-283 was also immunogenic and immunopathogenic in Lewis rats, but with a minimal dose approximately 100 times higher than that of 1179-1191. Peptide 880-892, a nondominant determinant, resembled 271-283 in its immunogenicity, but was markedly less immunopathogenic. No immunological activity was detected in the fourth repeat peptide, 579-591. Peptide 1179-1191 was superior to the other repeats also in its antigenicity, i.e., the capacity to stimulate presensitized lymphocytes in culture: the minimal stimulatory concentrations of 1179-1191 was greater than 1000 times lower than those of 271-283 or 880-892. Furthermore, 1179-1191 was stimulatory at concentrations lower than those of 271-283 even when tested with lymphocytes sensitized against 271-283. A correlation was also found between the immunological activities of the repeat peptides and their amphipathicity. This study thus identifies two new immunopathogenic determinants of IRBP and provides additional data to show the association between immunodominance of peptides and their various immunological activities.

Molecular Features of Class II MHC-Restricted T-Cell Recognition of Protein and Peptide Antigens: The Importance of Amphipathic Structures

Much work has been done on the use of peptides as immunogens, but most has concentrated on antibody production (reviewed in LERNER 1984; ARNON 1984) rather than T-cell responses. There are certain potential problems with their use for antibody production that do not apply to T cells. Antibodies raised against peptides as immunogens generally cross-react with the native protein from which the peptides were derived with affinities several Orders of magnitude lower than those for the peptide (reviewed in BERZOFSKY 1985 a). Conversely, antibodies raised against the native protein generally cross-react with peptides derived from it with a lower affinity, although exceptions have been noted (LANDO and REICHLIN 1982). In both directions, cross-reactions seem to be most easily measured when the peptide corresponds to a more mobile portion of the protein (TAINER et al. 1984, 1985; WESTHOF et al. 1984); perhaps such portions are better able to share some conformational states with the peptide so that one can achieve an induced fit with antibodies made against the other (BERZOFSKY 1985 a). In addition, there are antibodies made against the native protein that react with assembled topographic determinants consisting of amino acid residues far apart in the primary sequence that are brought together on the surface of the native molecule as it folds in its native conformation (BENJAMIN et al. 1984; BERZOFSKY 1985a). Such antigenic sites exist only on the surface topography of the native protein, and thus do not exist in any peptide segment of the protein.

Immunodominance of Amphipathic Peptides and Their Localization on the Cell Surface for Antigen Presentation to Helper T Cells

We have identified two immunodominant antigenic sites of sperm whale myoglobin using sequence variants and synthetic peptides. One is shown to be a region on which a polyclonal response is focused. Both sites are amphipathic helices, and the results suggest roles for both the hydrophilic and hydrophobic surfaces. This idea is generalized by the observation that 18 of 23 known immunodominant T cell sites from 12 proteins have the ability to fold as an amphipathic α- or 310-helix. This result implies a general feature fundamental to the chemistry of antigen presentation and T cell recognition and may be useful in the rational design of synthetic and recombinant vaccines. We further show, using biotin-conjugated peptide, that avidin can block presentation at the presenting cell surface. We conclude that immunologically relevant peptide antigen is present on the presenting cell surface in a form accessible to macromolecules in solution, despite previous difficulties blocking with antibodies to the antigen. The amphipathic peptide may be bound to Ia, or amphipathic structures may be selected by their ability to concentrate in the membrane to push the equilibrium toward a sufficient number of peptide–Ia complexes despite the low affinity of Ia for peptide.