Analysis of allele-specific contact sites of natural HLA-DR17 ligands.
The sequence motif of peptide ligands naturally associated with DR17 has indicated conserved residues at the relative positions P1-P4-P6-P 8.9 or 10. Eight naturally processed DR17 ligands were synthesized to study the role of conserved residues in DR17 binding. In their majority, they showed an excellent ability to bind to purified DR17 molecules. Binding experiments with variant peptides confirmed aspartate as the DR17-specific contact site at P4. In addition, hydrophobic or aromatic residues at P1 and P9, probably interacting with the NH2- and COOH-terminal pockets, and lysine or chemically related amino acids at P6 were important for binding. A core peptide of 10 amino acids, bordered by the terminal contact sites, is sufficient, although the ability to bind is reduced approximately 10-fold compared with the binding capacity of the natural ligand. Ala substitution of flanking stretches at either end completely restores the binding capacity to that of the natural ligand. This suggests that regions flanking the peptide core contribute to the binding strength nonspecifically, i.e., by forming H-bonds to MHC molecules. Natural DR1 and DR12 ligands like HLA-A2 (103-117) and transferrin receptor (140-156) failed to bind to DR17 molecules. However, substituting leucine for aspartate at P4 transformed DR1 and DR12 ligands into excellent DR17 binders. This conversion, enabled by a single amino acid substitution, emphasizes the importance of aspartate as the DR17-specific contact site and suggests that terminal contact residues are shared among DR1, DR12, and DR17 ligands. In contrast, additional aspartates introduced next to the contact site at P4 impaired the binding capacity. Regarding this specific role of asparate we expect that DR17-specific ligands will be rarely found among "promiscuous" peptides binding to several different DR molecules.
Department of Neurology, University of Tübingen, Germany.
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G
Malcherek
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Gnau
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Stevanovic
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Rammensee
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Jung
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Melms
D000483:Alleles; D000595:Amino Acid Sequence; D000941:Antigens / Q000737:chemistry* / Q000378:metabolism; D001055:Apolipoproteins B / Q000737:chemistry / Q000276:immunology / Q000378:metabolism; D001665:Binding Sites; D006684:HLA-DR Antigens / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008024:Ligands; D008969:Molecular Sequence Data; D013329:Structure-Activity Relationship; D000515:alpha 1-Antitrypsin / Q000737:chemistry / Q000276:immunology
D000941:Antigens; D001055:Apolipoproteins B; D006684:HLA-DR Antigens; D008024:Ligands; D000515:alpha 1-Antitrypsin