Demonstration of two distinct light chains in HLA-DR-associated antigens by two-dimensional gel electrophoresis.

The polypeptide composition of HLA-DR-associated antigens was analyzed by two-dimensional nonequilibrium pH gradient electrophoresis (NEPHGE)/sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis (PAGE). Glycoprotein fractions from B lymphoblastoid cell lines homozygous for the DR antigen were used as antigen source. The HLA-DR-associated antigens were isolated by immunoprecipitation using a monoclonal anti-DR antibody, TDR31.1. Two polypeptides corresponding in molecular weight to the HLA-DR-associated antigen beta chain were detected. These polypeptides were shown to be different molecules on the basis of their separation on NEPHGE/SDS-PAGE analysis, their rates of turnover relative to the other polypeptides of the HLA-DR-associated antigens, and their noncoordinate alteration in position on NEPHGE/SDS-PAGE two-dimensional analysis of different cell lines. Of the two beta polypeptides, only one was invariant in position on NEPHGE/SDS-PAGE analyses of cell lines homozygous for the same DR specificity. It appears therefore that the position of one of these beta polypeptides correlates with DR specificity, while the other beta polypeptide exhibits positional variation on NEPHGE/SDS-PAGE analysis, indicative of polymorphism at a second HLA-DR-associated locus.