PROBING IMMUNOGENICITY OF A T-CELL EPITOPE BY L-ALANINE AND D-AMINO-ACID SCANNING

All residues of the I-E(d) restricted fragment 24-36 of a snake toxin were individually changed into L-alanine and the corresponding D-enant iomer. Four analogs substituted with L-Ala at positions 25, 30, 31 and 33, and nine analogs substituted with a D-residue along the stretch 2 5-33 lost most (position 28) or all their capacity to stimulate a toxi n-specific T hybridoma. None of these analogs stimulated splenocytes f rom mice immunized with the peptide 24-36. Only the L-A(31) and D-W-29 modified analogs could prime a T cell response which, however, showed no cross-reactivity with the native peptide, demonstrating that T cel l response selectivity can be deeply modified by mutation or configura tion inversion of a single residue. Our data suggest that (i) the regi on 25-33 is the core of the T epitope that binds to I-E(d), and (ii) Y -25 R(30) and R(33) contribute to the peptide binding by anchoring int o pockets of I-E(d). In agreement with T cell priming observations, on ly the L-A(31) and D-W-29 modified analogs elicited strong antibody re sponses, just like the peptide 24-36, whereas nearly all other analogs were less immunogenic. All but the L-Ala(30) and L-Ala(33) modified a nalogs were recognized by a 24-36 specific antiserum as well as the na tive peptide. Altogether, our results show that susbstitution by D-ami no acid in a peptide could be particulary well-suited for either minim izing the risk of hypersensitivity or designing peptidic vaccines.