Purification and structural characterization of LFA-1, a lymphocyte function-associated antigen, and Mac-1, a related macrophage differentiation antigen associated with the type three complement receptor.

Abstract

LFA-1, an antigen associated with antigen-specific T lymphocyte-mediated killing, and Mac-1, a macrophage differentiation antigen associated with type three complement receptor function, contain alpha chains of Mr = 180,000 and 170,000, respectively, and beta chains of Mr = 95,000. The monoclonal antibodies defining these antigens do not cross-react. The LFA-1 and Mac-1 beta chains are highly homologous or identical, whereas the alpha chains are highly different by tyrosyl tryptic peptide mapping (Kürzinger, K., Ho, M. K., and Springer, T. A. (1982) Nature (Lond.) 296, 668-670). T lymphoma cell lines express LFA-1 but not Mac-1 as shown by immunofluorescence and immunoprecipitation. Conversely, some macrophage-like lines express Mac-1 but not LFA-1. Other macrophage-like lines co-express Mac-1 and small amounts of LFA-1. Mac-1 and LFA-1 are present as separate molecules in these cells. [35S]Methionine and [[3H]glucosamine are incorporated into both alpha and beta chains of Mac-1 and LFA-1, showing both chains are endogenously synthesized and are glycoproteins. Cross-linking and two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis experiments show that in both Mac-1 and LFA-1 the alpha and beta chains are noncovalently associated in alpha 1 beta 1 quaternary structures. By quantitative immunofluorescence flow cytometry, the EL-4 T lymphoma and P388D1 macrophage-like lines were estimated to express 10(5) LFA-1 and 1.6 x 10(5) Mac-1 molecules/cell, respectively. From these sources the antigens have been purified to homogeneity in 200-400-micrograms quantities by monoclonal antibody affinity chromatography. The purified antigens contain only the alpha and beta subunits.