Fig. 2. Heterodimer formation and antigen binding of mutated DRß chains. Point mutations were introduced into DR1ß chain at residues Y123, D152 or W153 at Y123 and W153 and at Y123, D152 and W153, which were substituted by A. Mutant DR1 and wild-type DR1ß chains were co-expressed with DR in COS-7 cells. Residues Y, D and W, mutated to A are indicated on top. Cells were metabolically labeled with [³⁵S]methionine for 30 minutes. DR heterodimers were immunoprecipitated from cell lysates with mAb I251. Immunoprecipitates separated by SDS-PAGE were exposed to films for 24 hours. The positions of the DR and DRß bands are indicated at the right. (B) Binding of an antigenic influenza-virus-derived sequence (MAT) to DR1 mutants. A recombinant Ii (Ii-MAT), where the MHCII-binding sequence was replaced by a DR1-binding sequence derived from the matrix protein of influenza virus was co-expressed with mutant (lanes 2 to 6) and wild-type DR1ß chains (lane 1), combined with DR chain. DRß chains with mutated Y, D and W residues are indicated on top of lanes 2 to 6. DR was immunoprecipitated with I251. Upon separation of the immunocomplexes by SDS-PAGE, Ii-MAT was immunoblotted with In-1 against Ii. Separation of Ii-MAT served as a reference for the recombinant Ii (lane 7). Heavy (H) and light (L) chains are indicated on the left and the position of Ii-MAT is shown at the right.