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-2Db-restricted CD8 cytotoxic T lymphocyte (CTL) clone which is specific for the 60-kDa mycobacterial heat shock protein (hsp) and cross-reacts with stressed host cells were characterized. None of the nonapeptides from hsp60 conforming to the H-2Db binding motif were able to sensitize target cells for lysis by this CTL clone. Sequence analysis of the stimulatory fraction from a trypsin digest of hsp60, together with synthetic peptide studies, defined a cluster of overlapping epitopes. Carboxy-terminal extension by at least one amino acid of the nonamer predicted to bind best to H-2Db was 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 the natural epitope. In contrast, the 11-mer was inactive. The stimulatory 10-mer bound to H-2Db with an efficacy similar to that of the nonapeptide corresponding to the H-2Db motif, as revealed by peptide induced major histocompatibility complex (MHC) surface expression on RMA-S cells and competitive blocking of epitope recognition by the nonamer. Binding of these carboxy-terminally extended peptides to the MHC groove can be explained by anchoring through the amino acid residue Asn in position 5 of the peptide and by intrusion of the hydrophobic carboxy-terminal Ala (10-mer) or Leu (12-mer), but not Gly (11-mer), into the hydrophobic pocket of the H-2Db cleft. Because the carboxy-terminal 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 specificity for a single T cell receptor. This flexibility not only promotes recognition of several overlapping peptides from a single antigen, but may also increase the chance of cross-reaction with similar peptides from unrelated proteins, including autoantigens. Consistent with this latter assumption, the T cell clone cross-recognizes mycobacterial hsp60 and stressed host cells.
Schoel
B
B
Department of Immunology, University of Ulm, FRG.
Zügel
U
U
Ruppert
T
T
Kaufmann
S H
SH
D000595:Amino Acid Sequence; D000818:Animals; D000942:Antigens, Bacterial / Q000737:chemistry; D002478:Cells, Cultured; D018834:Chaperonin 60 / Q000276:immunology; D003602:Cytotoxicity, Immunologic; D018604:Epitope Mapping; D006183:H-2 Antigens / Q000276:immunology*; D007111:Immunity, Cellular; D066298:In Vitro Techniques; D051379:Mice; D008969:Molecular Sequence Data; D009161:Mycobacterium / Q000276:immunology; D010455:Peptides / Q000737:chemistry / Q000276:immunology*; D013602:T-Lymphocytes, Cytotoxic / Q000276:immunology*
D000942:Antigens, Bacterial; D018834:Chaperonin 60; D006183:H-2 Antigens; D010455:Peptides