Isolation and biochemical characterization of the H-2Kk and H-2Dk antigens from the RDM-4 lymphoma

Milligram amounts of the H-2Kk and H-2Dk antigens from the RDM-4 lymphoma have been isolated in a one-step procedure employing affinity chromatography on monoclonal antibodies conjugated to Sepharose. The purified antigens have been characterized biochemically, including amino-terminal amino acid sequence and tryptic peptide analyses of the 45,000 mol. wt chains of the antigens. The amino acid sequence results for the H-2Kk antigen, when compared with previously reported data for this molecule, reveal a discrepancy in the sequence of this 45,000 mol. wt chain. We postulate that this discrepancy reflects a previously undescribed polymorphism for the Kk-gene. We also report the first amino-terminal amino acid sequence for the 45,000 mol. wt chain from the H-2Dk antigen and demonstrate that the sequence of the β2-mieroglobulin obtained from the H-2Kk antigen preparation is identical to that published previously. Finally, the structural integrity of the H-2Kk antigen is assessed using binding of β2-microglobulin, incorporation into liposomes and reaction with antibodies as criteria.