Malcherek G, Gnau V, Stevanovic S, Rammensee H G, Jung G, Melms A
Department of Neurology, University of Tübingen, Germany.
J Immunol. 1994 Aug 1;153(3):1141-9.
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.
与DR17天然相关的肽配体的序列基序已表明在相对位置P1 - P4 - P6 - P8、9或10处存在保守残基。合成了8种天然加工的DR17配体，以研究保守残基在DR17结合中的作用。它们中的大多数显示出与纯化的DR17分子结合的出色能力。用变体肽进行的结合实验证实天冬氨酸是P4处的DR17特异性接触位点。此外，P1和P9处的疏水或芳香族残基可能与NH2 - 和COOH - 末端口袋相互作用，P6处的赖氨酸或化学相关氨基酸对结合很重要。由末端接触位点界定的10个氨基酸的核心肽就足够了，尽管其结合能力与天然配体的结合能力相比降低了约10倍。在任一端侧翼延伸段的丙氨酸取代完全恢复了与天然配体相同的结合能力。这表明肽核心侧翼区域非特异性地有助于结合强度，即通过与MHC分子形成氢键。天然的DR1和DR12配体，如HLA - A2（103 - 117）和转铁蛋白受体（140 - 156）不能与DR17分子结合。然而，在P4处用亮氨酸取代天冬氨酸可将DR1和DR12配体转化为出色的DR17结合剂。这种由单个氨基酸取代实现的转化强调了天冬氨酸作为DR17特异性接触位点的重要性，并表明末端接触残基在DR1、DR12和DR17配体之间是共享的。相反，在P4处接触位点旁边引入额外的天冬氨酸会损害结合能力。鉴于天冬氨酸的这种特定作用，我们预计在与几种不同DR分子结合的“混杂”肽中很少会发现DR17特异性配体。
