B Cell Responses to a Peptide Epitope. X. Epitope Selection in a Primary Response Is Thermodynamically Regulated

Abstract

We examine the etiological basis of hierarchical immunodominance of B cell epitopes on a multideterminant Ag. A model T-dependant immunogen, containing a single immunodominant B cell epitope, was used. The primary IgM response to this peptide included Abs directed against diverse determinants presented by the peptide. Interestingly, affinity of individual monomeric IgM Abs segregated around epitope recognized and was independent of their clonal origins. Furthermore, affinity of Abs directed against the immunodominant epitope were markedly higher than that of the alternate specificities. These studies suggested that the affinity of an epitope-specific primary response, and variations therein, may be determined by the chemical composition of epitope. This inference was supported by thermodynamic analyses of monomer IgM binding to Ag, which revealed that this interaction occurs at the expense of unfavorable entropy changes. Permissible binding required compensation by net enthalpic changes. Finally, the correlation between chemical composition of an epitope, the resultant affinity of the early primary humoral response, and its eventual influence on relative immunogenicity could be experimentally verified. This was achieved by examining the effect of various amino-terminal substitutions on immunogenicity of a, hitherto cryptic, amino-terminal determinant. Such experiments permitted delineation of a hierarchy of individual amino acid residues based on their influence; which correlated well with calculated Gibbs-free energy changes that individual residue side chains were expected to contribute in a binding interaction. Thus, maturation of a T-dependant humoral response is initiated by a step that is under thermodynamic control.

The Primary Antibody Repertoire Represents a Linked Network of Degenerate Antigen Specificities

“…The anti-(4-hydroxy-3-nitrophenyl)-acetyl (anti-NP) 3 hybridomas BBE6.12H3 and Bg53-5, and the anti-p-azophenylarsonate (anti-Ars) hybridomas 36-65 and 36-71 were a kind gift from Dr. T. Manser (Kimmel Cancer Institute, Philadelphia, PA), whereas the anti-peptide hybridoma PC7bM has been described earlier (12). All of these mAbs were first obtained as ascites and then purified by affinity chromatography as described earlier (5).…”

“…All of these mAbs were first obtained as ascites and then purified by affinity chromatography as described earlier (5). When necessary, the IgM class mAb PC7bM was subsequently converted to its monomeric form by reductive alkylation as described earlier (12). For mAb 36-65, the amino acid sequences of the complementarity-determining region 3 of the H chain (HCDR3) and complementarity-determining region 3 of the L chain (LCDR3) are SVYYGGSYY-FDY and QQGNTLPRT, respectively.…”

“…For this, biotinylated Ab was used, and the immune complexes were separated using streptavidin-agarose. Bound phage was eluted by incubation with a 10-fold excess of the homologous peptide Ag, PS1CT3 (12). For binding analysis, all individual clones were amplified to titers of between 10 13 and 3 ϫ 10 17 PFU/ml, and then they were either biotinylated for surface plasmon resonance studies or derivatized with FITC for analysis of binding to M12 cells.…”

The Fourth Surface-Exposed Region of the Outer Membrane Protein P5-Homologous Adhesin of Nontypable Haemophilus influenzae Is an Immunodominant But Nonprotective Decoying Epitope

“…more conserved loop 6 were capable of eliciting bactericidal antisera capable of binding 14 of the 15 NTHI strains assayed (35). Hence, by locking the immune response onto a dominant decoying epitope of a multideterminant Ag, this group of microorganisms could ensure that, via the mechanisms of deceptive imprinting, the host would continue to mount and expand an ineffective immune response to that epitope even when reinfected with a different but antigenically related member of the same microbial family (22).…”

“…These selective, epitope-specific primary responses also influence the hierarchy of immunodominance observed in later, secondary responses (22,23). Whereas in the past we have described the ability of several immunogens derived from surface-exposed region 3 of the P5-homologous adhesin to confer augmented bacterial clearance or protection from development of experimental OM, we have not extensively characterized the quality of the induced protective Ab response in those animal models, nor have we compared that response to the nonprotective response induced by either experimental OM in chinchillas or natural OM in children.…”