Structural Relatedness of Distinct Determinants Recognized by Monoclonal Antibody TP25.99 on β2-Microglobulin-Associated and β2-Microglobulin-Free HLA Class I Heavy Chains

Abstract

The association of HLA class I heavy chains with β2-microglobulin (β2m) changes their antigenic profile. As a result, Abs react with either β2m-free or β2m-associated HLA class I heavy chains. An exception to this rule is the mAb TP25.99, which reacts with both β2m-associated and β2m-free HLA class I heavy chains. The reactivity with β2m-associated HLA class I heavy chains is mediated by a conformational determinant expressed on all HLA-A, -B, and -C Ags. This determinant has been mapped to amino acid residues 194–198 in the α3 domain. The reactivity with β2m-free HLA class I heavy chains is mediated by a linear determinant expressed on all HLA-B Ags except the HLA-B73 allospecificity and on <50% of HLA-A allospecificities. The latter determinant has been mapped to amino acid residues 239–242, 245, and 246 in the α3 domain. The conformational and the linear determinants share several structural features, but have no homology in their amino acid sequence. mAb TP25.99 represents the first example of a mAb recognizing two distinct and spatially distant determinants on a protein. The structural homology of a linear and a conformational determinant on an antigenic entity provides a molecular mechanism for the sharing of specificity by B and TCRs.

Switch ofHLA-G alternative splicing in a melanoma cell line causes loss of HLA-G1 expression and sensitivity to NK lysis

“…The following MAbs (monoclonal antibodies) were used: MEM-G/ 9, a mouse IgG1 specific for native HLA-G1 and HLA-G5 isoforms (Exbio, Prague, Czech Republic); 35 4H84, a mouse IgG1 anti-HLA-G free heavy chain (kindly provided by M. Mc Master, University of California, San Francisco, CA); 34,36 5A6G7, a mouse IgG1 specific for the intron 4-retaining part of both soluble HLA-G5 and HLA-G6; 37 TP25.99, a mouse IgG1 anti-HLA-A, -B, -C and -E but not anti-HLA-G (kindly provided by S. Ferrone, Roswell Park Cancer Institute, Buffalo, NY); 34,38 B1G6, a mouse IgG2a, anti-b2 microglobulin (Immunotech, France); B8.12.2, a mouse IgG2b anti-HLA-DR (Immunotech); GHI/75, a mouse IgG2b anti-ILT-2 (Pharmingen, San Diego, CA); Z199, a mouse IgG2b anti-NKG2A (Immunotech) and B-5-1-2 , a mouse IgG1 anti-a tubulin (Sigma Chemical Co.).…”

β2-Microglobulin-Free HLA Class I Heavy Chain Epitope Mimicry by Monoclonal Antibody HC-10-Specific Peptide

“…HLA-A, B, C, E, and G HC (15,16); the mAb TP25.99 (IgG1) which recognizes distinct determinants expressed on all ␤ 2 m-associated HLA-A, B, C, and E HC and on all ␤ 2 m-free HLA-A, B, C HC except A2, A10, A29, A31, 32, 33, A68.1, A69, and B73 (4,17); and the mAb L31 (9,12) which recognizes a determinant preferentially expressed on ␤ 2 m-free HLA-B HC were produced and characterized as previously described. The BALB/c mouse mAb 655 (IgG1) and its corresponding Ag, the human V␤-17 TCR-derived peptide GYSVSREKKES (p-V␤17), were used as controls.…”

Differential Immunogenicity of Two Peptides Isolated by High Molecular Weight-Melanoma-Associated Antigen-Specific Monoclonal Antibodies with Different Affinities

“…Anti-HLA class I Ag mAb TP25.99 was developed as previously described (23). Affinity-purified rabbit anti-mouse IgG Abs and HRP-conjugated goat anti-mouse Ig Abs were purchased from Jackson ImmunoResearch Laboratories.…”

“…The binding assays to assess the reactivity of mAb with peptides and of sera with peptides and cells were performed as previously described (23,27). The Ab titer was defined as the highest serum dilution that resulted in 50% of maximal binding of serum to peptides or cells.…”