Inducibility of Protein-Reactive Antibodies by Peptide Immunization: Comparison of Three Epitope Peptides of Hen Egg-White Lysozyme

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Three epitope peptides of hen egg-white lysozyme (HEL) were tested for ability to induce antibodies reactive with native HEL. Each peptide was coupled to bovine γ-globulin (BγG) and 4 rabbits were immunized with each peptide-BγG conjugate in complete Freund's adjuvant. The mean association constants (K_os) of HEL-reactive antibodies (HEL-R-Abs) from each immunizing group to [³H]acetyl HEL or to [³H]acetyl-peptide were measured in solution by a double antibody method. Only peptide loop I•II (sequences 57-107 containing Cys64-Cys80 and Cys76-Cys94) induced high-affinity antibodies to HEL (K_o=2.5×10⁶-2.3×10⁷M^-1) among the three epitope peptides tested. The association constants of antipeptide loop I•II to [³H] acetyl peptide loop I•II were always one to two orders of magnitude higher than those to HEL. In addition, 50 to 80% of the anti-peptide loop I•II antibodies were reactive with native HEL. The specificity of anti-peptide loop I•II was directed to a conformational feature of the peptide rather than to native HEL and reactivity of the antibody to HEL was interpreted as a kind of cross-reaction. The HEL-R-Abs from anti-Ploop I•II antisera also manifested neutralizing activities against the enzymic activity of HEL when Micrococcus luteus was used as the substrate.