Two structural states of complexes of peptide and class II major histocompatibility complex revealed by photoaffinity-labeled peptides.
The complex of the murine class II histocompatibility molecules I-A(k) with high affinity binding peptides were resistant to denaturation when examined by SDS-polyacrylamide gel electrophoresis at various pH levels. In contrast, complexes made with low affinity binding peptides were highly sensitive to denaturation by SDS. This effect was more pronounced at low pH. Placing a photoactivatable probe at the amino terminus of the peptides resulted in their covalent linkage to soluble I-A(k) molecules. We found an inverse relationship between the capacity of peptides to form SDS-stable complexes with I-A(k) and their extent of covalent association with either the alpha or beta chain. The relationship held true for three different peptides in which the main anchor residues were changed so as to affect their binding affinity for I-A(k) molecules. Thus, high affinity peptides generate a complex in which the motion of their amino termini was restricted, whereas complexes of low affinity peptides are more flexible. In agreement with this observation, complexes of I-A(k) with high affinity peptides were highly resistant to proteolysis, in contrast to those formed with weakly binding peptides, which were more likely to be cleaved. Complexes with low affinity peptides generate a structure with enhanced flexibility as compared with complexes with high affinity peptides.
Carrasco-Marín
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Department of Pathology, Center for Immunology, Washington University School of Medicine, St. Louis, Missouri 63110, USA.
Petzold
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Unanue
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ER
D000595:Amino Acid Sequence; D000818:Animals; D017951:Antigen Presentation; D003432:Cross-Linking Reagents; D000949:Histocompatibility Antigens Class II / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D016056:Immunodominant Epitopes / Q000378:metabolism*; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D009113:Muramidase / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D019920:Photoaffinity Labels; D011485:Protein Binding; D011487:Protein Conformation; D011489:Protein Denaturation; D012967:Sodium Dodecyl Sulfate
D003432:Cross-Linking Reagents; D000949:Histocompatibility Antigens Class II; C111388:I-Ak antigen; D016056:Immunodominant Epitopes; D046911:Macromolecular Substances; D010446:Peptide Fragments; D019920:Photoaffinity Labels; C400524:hen egg lysozyme peptide (48-61); D012967:Sodium Dodecyl Sulfate; D009113:Muramidase