CHARACTERIZATION OF T-CELL EPITOPES RESTRICTED BY HLA-DP9 IN STREPTOCOCCAL M12 PROTEIN

Interaction of the HLA-DP9 (DPA10201/DPB1*0901) molecule and M protei n of serotype 12 (SS95/12) streptococci, a main component of the strep tococcal cell wall Ag, has been investigated to decipher peptide-bindi ng capacity and T cell activation in the context of the HLA-DP molecul e. Seven antigenic peptides (amino acids 19-25) restricted by the HLA- DP9 molecule were identified in M12 protein, using M12 protein- or pep tide-specific T cell lines from naturally exposed individuals. The bin ding affinity of each peptide to the HLA-DP9 molecule was measured by fluorescence intensity of biotinylated peptides bound to L cell transf ectants expressing HLA-DP9, followed by treatment with advom-fluoresce nce. Binding of biotinylated peptides to the HLA-DP9 molecule was inhi bited by an excess amount of corresponding nonbiotinylated peptides an d other nonbiotinylated peptides, indicating that the peptides were bo und to the HLA-DP9 molecule at a single binding site. Seven synthetic peptides containing the T cell epitopes restricted by the HLA-DP9 mole cule had high binding affinity to the HLA-DP9 molecule. Comparison of the amino acid sequences of truncated analogues that could bind to the HLA-DP9 molecule and/or activate T cells suggested an HLA-DP9-specifi c binding motif, composed of a positively charged residue (R or K) at position 1, a hydrophobic residue (A, G, or L) at position 6, and anot her hydrophobic residue (L or V) at position 9. Analysis of single ami no acid-substituted analogues suggested that the positively charged am ino acid in the motif served as a key anchor residue for binding to th e HLA-DPS molecule, which differs from the binding motif to the HLA-DR molecules.