Identification of T- and B-cell epitopes in synthetic peptides derived from a Streptococcus mutans protein and characterization of their antigenicity and immunogenicity

Abstract

Natural immunity to synthetic peptides (SP) derived from the sequences of a 3800 M r Streptococcus mutans antigen was found in human subjects. Significant serum IgG antibodies were detected both to the native streptococcal antigen and to the SP17, containing essentially residues 1–15. A series of short peptides with deletions at the amino- and carboxy-termini were then tested to identify the B-cell epitopes. Residues 8–13 and 1–6 bound significant serum IgG antibodies but only the former consistently inhibited human antibodies, suggesting that residues 8–13 constitute a major B-cell epitope. The human CD4 subset of T-cells was then examined and this showed a significant uptake of [ 3H]-thymidine when stimulated with both the native streptococcal antigen and the SP17. The series of short peptides was then used to stimulate CD4 cells, in order to determine the T-cell epitope. The synthetic peptide with residues 6–15 was the shortest peptide that stimulated significant [ 3H]-thymidine uptake and this peptide was designated as a T-cell epitope. The immunogenicity and antigenicity of SP17 was also investigated in macaques. Immunization of monkeys with the free SP17 failed to elicit serum antibodies or T-cell responses. However, immunization with SP17 linked to tetanus toxoid as a carrier elicited serum antibodies and proliferative responses of lymphocytes, not only to the synthetic peptide but also to the native streptococcal antigen. As in the human studies a B-cell epitope was found in residues 8–13, whereas an overlapping T-cell epitope was located in residues 7–15. In spite of the B- and T-cell epitopes being expressed in SP17 (residues 1–15), the synthetic peptide without a carrier failed to elicit serum antibodies or proliferative responses of lymphocytes in monkeys. However, immunization with a dimer of SP17 induced both serum antibodies and proliferative responses of lymphocytes. The findings suggest that the monomeric SP17 is not immunogenic and needs to be dimerized in order to elicit antibodies and T-cell responses, both to the synthetic peptide and to the native streptococcal antigen. A comparison of the T- and B-cell epitopes in humans and in non-human primates revealed that a B-cell epitope (residues 8–13) was identical in both primates. The T-cell epitope in the two primates differed only by 1 amino acid, in that residues 7–15 in the non-human primate required an additional isoleucine (residue 6) to stimulate human T-cells. Hence, the B-cell epitope and, with the exception of 1 residue, the T-cell epitope share immunogenic peptide sequences in primates, although the humans were exposed to natural immunity and the non-human primates were immunized artificially.