Acidic residues in the DRβ chain third hypervariable region are required for stimulation of a DR(α,β1*0402)-restricted T-cell clone

We investigated the minimal requirements for stimulation of an antigen-specific HLA-DR(α,β1*0402)-restricted T-cell clone (Een 217) by using transfectants expressing mutant DRβ chains as APCs. Antigen-specific stimulation of Een217 was induced with transfectants expressing DR(α,β1*0402) and DR(α,β1*0403) but not other DR4 subtypes that also bind the peptide recognized by this clone. Analysis of the effects of single amino acid substitutions in the β chains of each of the DR4 subtypes revealed a requirement for acidic residues in the third HVR, particularly amino acid 71, in stimulation of clone Een217. Functional class II Mutants were generated from nonstimulatory DR4 subtype β chains by acidic residue substitutions within the third HVR. These data define the requirement for negatively charged residues in this region for peptide-induced stimulation of this T-cell clone. The required acidic residues can be located at either position 70, 71, or 74 in the DRβ chain. The negative charge in this segment of the DRβ chain α-helix may be required for direct interactions with the T-cell receptor of Een217 or may affect peptide conformation.