ABSTRACT
A soluble, recombinant form of the human T cell receptor (TCR) beta-chain containing the V beta 3.1 sequence has been constructed, expressed in Chinese hamster ovary cells, amplified by dihydrofolate reductase selection, and purified in quantities appropriate for the generation of monoclonal antibodies (mAb). The V beta 3 sequence was chosen because of its reported elevated usage in the synovial T cells of rheumatoid arthritis patients but the approach described should be applicable to other known human V beta gene sequences. By this method, two mAb were prepared which reacted with up to 10% of normal, live peripheral blood T cells but with reactivity varying greatly among individual donors. Both mAb specifically bound to a murine T cell line transfected with a human TCR V beta 3.1 and immunoprecipitated a protein of the expected molecular weight for the TCR beta-chain. Both antibodies were mitogenic for T cells and analysis of peripheral blood lymphocyte cultures stimulated with the mAb suggested that both were specific for the V beta 3.1 subfamily and not D beta or J beta. Clones expressing V beta 3, which were derived from mAb-stimulated peripheral blood lymphocytes of a single individual, preferentially (8/13), but not exclusively, utilized the J beta 2.7 gene segment. The V beta 3.1 usage showed no preference for the CD8+ or CD4+ subpopulations of normal peripheral blood T cells.
Subject(s)
Antibodies, Monoclonal/immunology , Receptors, Antigen, T-Cell, alpha-beta/immunology , T-Lymphocytes/immunology , Animals , Base Sequence , Female , Humans , Lymphocyte Activation , Mice , Mice, Inbred BALB C , Molecular Sequence Data , Receptors, Antigen, T-Cell, alpha-beta/chemistry , Recombinant Proteins/immunology , SolubilityABSTRACT
A mAb, OT145, recognizes a TCR allotype encoded by one of two alleles of the V beta 6.7 gene. The peptide products of the two V beta 6.7 alleles differ because of nonconservative amino acid substitutions at positions 38 and 72. V beta 6.7a encodes ser38 and gly72, whereas V beta 6.7b encodes arg38 and glu72. We show here that the binding of mAb OT145 ot the beta-chain of TCR is lost when residue 72 of V beta 6.7a is mutated from gly to glu. The binding of OT145 is not affected by mutation of residue 38 from ser to arg. Thus, OT145 recognizes an epitope related to position 72. Residue 72 of the beta-chain of the TCR is located at a putative superantigen-binding site.