Properties and specificity of binding sites for the immunomodulator bestatin on the surface of mammalian cells.

The binding of the immunomodulator bestatin, an inhibitor of cell surface bound leucine aminopeptidase and aminopeptidase B, to mammalian cells of varying origin has been studied. The specific binding of [3H] bestatin was a rapid and saturable process exhibiting one affinity, characterized by an association constant of 0.8 x 10(5) M-1, as determined in the L5178y mouse lymphoma system. Optimal binding was observed at 37 degrees C. L-leucine and L-leucine-beta-naphthylamide prevented the binding, suggesting that the complex was formed between leucine aminopeptidase and bestatin. The protein nature of the bestatin-"receptor" was suggested by its susceptibility to trypsin. Under the conditions used here intracellular translocation of bestatin appeared to be negligible. A maximum of about 2.2 x 10(6) bestatin molecules could bind to L5178y mouse lymphoma cells. Under identical conditions by far the highest amount of bestatin was bound to macrophages from mice. Lower levels were measured with T-lymphocytes; very low binding capacity was observed with B-lymphocytes. Experiments with synchronized L5178y cells revealed a cell cycle dependent change of binding capacity for bestatin; the highest level was observed during the transition from S-to G2 phase and the lowest during G1- and early S phase. These data lend further support to the assumption that the immuno-potentiating activity of bestatin is due to a stimulation of T-lymphocyte proliferation probably mediated through the activation of macrophages.