Variable binding affinities of listeriolysin O peptides for the H‐2K^d class I molecule

Previously we used the peptide‐binding motif for the murine class I major histocompatibility complex molecule H‐2K^d to identify a nonamer peptide of the Listeria monocytogenes listeriolysin (LLO) protein that was recognized by cytotoxic T lymphocytes (CTL) in association with H‐2K^d. Eleven nonamer peptides contained in the LLO sequence were synthesized and one, LLO 91‐99, proved to be a CTL target. Using peptide binding competition assays with H‐2K^d‐restricted CTL, we show that 3 out of the 11 LLO peptides, including the CTL epitope, have a high binding affinity for H‐2K^d; 2 of 11 peptides have approximately 10‐fold lower affinity, while the remaining 6 peptides have no or very low affinity for H‐2K^d. Single residue changes were made in the LLO 91‐99 peptide and two other LLO peptides to identify non‐anchor amino acids that might interfere with peptide binding. In addition, we used the LLO peptides which bound well to H‐2K^d to attempt to restimulate a secondary CTL response from L. monocytogenes‐primed spleen cells. Only LLO 91‐99 was able to induce such a response. Thus only a fraction of nonamer peptides which fit the original binding motif have a high affinity for the H‐2K^d class I molecule, and only a fraction of these serve as CTL epitopes.