Molecular characterization of serologic recognition sites in the human HLA-A2 molecule.
The localization of the amino acid residues involved in the serologic specificity of the HLA-A2 molecule has been investigated using a combination of site-directed mutagenesis, DNA-mediated gene transfer, indirect immunofluorescence and flow cytometry techniques. Synthetic oligonucleotides were designed to introduce individual and combined amino acid substitutions in both the alpha 1 (positions 9, 43, and the highly polymorphic cluster of residues from aa 62 to 83) and alpha 2 (positions 107, 152, and 156) domains to investigate the effect of the specific mutation on the recognition of the molecule at the surface of transfected human and mouse cell lines by a panel of mAb that recognize monomorphic or polymorphic determinants in MHC class I molecules. At least three non-overlapping serologic epitopes were identified. Mutations in the highly polymorphic region at aa 62 to 66 completely eliminated binding of mAb MA2.1 (A2/B17 cross-reactive). Mutation at position 107 resulted in complete loss of binding of the A2/Aw69-specific mAb PA2.1 and MA2.2 and partial loss of mAb BB7.2 binding. The recognition by other allotypic mAbs was not affected by these mutations and they therefore represent at least a third serologic epitope. Mutations at positions 152 and 156, known to be important for T cell recognition, did not affect serologic recognition. Introduction of residues of HLA-B7 origin in the polymorphic segment spanning aa 70 to 80 created a molecule carrying the -Bw6 supertypic determinant as demonstrated by mAb SFR8-B6 binding.
Santos-Aguado
J
J
Division of Tumor Virology, Dana-Farber Cancer Institute, Boston, MA 02115.
Barbosa
J A
JA
Biro
P A
PA
Strominger
J L
JL
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal; D000937:Antigen-Antibody Reactions; D001483:Base Sequence; D000939:Epitopes / Q000235:genetics* / Q000302:isolation & purification; D005796:Genes; D015234:HLA-A Antigens / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D015789:HLA-A2 Antigen; D006650:Histocompatibility Testing; D006801:Humans; D007739:L Cells; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009710:Nucleotide Mapping; D015345:Oligonucleotide Probes; D012208:Rhabdomyosarcoma
D000911:Antibodies, Monoclonal; D000939:Epitopes; D015234:HLA-A Antigens; D015789:HLA-A2 Antigen; D015345:Oligonucleotide Probes