Systematic identification of H-2 Kd binding peptides and induction of peptide specific CTL

Most peptides with putative MHC I restricted sequence motifs do not bind to the corresponding MHC I nor induce cytolytic T cells. There exist additional constraints which limit peptide binding and immunogenicity. To identify immunogenic peptides in novel protein sequences, it will be necessary to first evaluate peptide binding to MHC I. In this study, a soluble single chain fusion protein SC-Kd was used to evaluate potential Kd binding peptides from the sequences of mouse mammary tumor virus gag and env proteins. A total of 27 peptides were identified which displayed the reported Kd restricted motif. Of the 27 peptides, six demonstrated strong to moderate binding to SC-Kd. The strongest binding peptides expressed tyrosine or phenylalanine at position 2 and leucine at the C-terminus. The capability of MMTV peptides to induce CTL corresponds to their SC-Kd binding activity. Of the six peptides that demonstrated moderate to strong binding, five induced CTL in BALB/c mice. These peptides induced CTL after 1-3 in vivo immunizations followed by 5 day in vitro stimulation. Furthermore, a single in vitro stimulation of naive lymphocytes with strong-binding G425 was sufficient to induce significant CTL activity. Weak or non-binding peptides did not induce CTL. Therefore, peptide binding to SC-Kd is a predictive indicator of CTL inducing activity.