ELONGATED PEPTIDES, NOT THE PREDICTED NONAPEPTIDE STIMULATE A MAJOR HISTOCOMPATIBILITY COMPLEX CLASS I-RESTRICTED CYTOTOXIC T-LYMPHOCYTE CLONE WITH SPECIFICITY FOR A BACTERIAL HEAT-SHOCK PROTEIN

The peptides recognized by an H-2D(b)-restricted CD8 cytotoxic T lymph ocyte (CTL) clone which is specific for the 60-kDa mycobacterial heat shock protein (hsp) and cross-reacts with stressed host cells were cha racterized. None of the nonapeptides from hsp60 conforming to the H-2D (b) binding motif were able to sensitize target cells for lysis by thi s CTL clone. Sequence analysis of the stimulatory fraction from a tryp sin digest of hsp60, together with synthetic peptide studies, defined a cluster of overlapping epitopes. Carboxy-terminal extension by at le ast one amino acid of the nonamer predicted to bind best to H-2D(b) wa s essential for CTL recognition. Two such elongated peptides, a 10-mer and a 12-mer stimulated the clone at similarly low concentrations in the 100 pM range. We assume that these two peptides comply best with t he natural epitope. In contrast, the 11-mer was inactive. The stimulat ory 10-mer bound to H-2D(b) with an efficacy similar to that of the no napeptide corresponding to the H-2D(b) motif, as revealed by peptide i nduced major histocompatibility complex (MHC) surface expression on RM A-S cells and competitive blocking of epitope recognition by the nonam er. Binding of these carboxy-terminally extended peptides to the MHC g roove can be explained by anchoring through the amino acid residue Asn in position 5 of the peptide and by intrusion of the hydrophobic carb oxy-terminal Ala (10-mer) or Leu (12-mer), but not Gly (11-mer), into the hydrophobic pocket of the H-2D(b) cleft. Because the carboxy-termi nal part is thus larger than predicted, this region of the peptide may arch up from the binding groove. We assume that recognition of steric components of the MHC/peptide complex broaden the range of epitope sp ecificity for a single T cell receptor. This flexibility not only prom otes recognition of several overlapping peptides from a single antigen , but may also increase the chance of cross-reaction with similar pept ides from unrelated proteins, including autoantigens. Consistent with this latter assumption, the T cell clone cross-recognizes mycobacteria l hsp60 and stressed host cells.