Binding of chemical derivatives of Phaseolus vulgaris phytohemagglutinin to pig spleen lymphocytes

Abstract

The mitogenic activity of Phaseolus vulgaris phytohemagglutinin (PHA) and its acetyl, citraconyl, glycinyl, homoarginyl, N-bromosuccinimide, and nitro derivatives has been assayed using pig spleen lymphocytes. Nitrated and N-bromosuccinimide-treated PHA were not mitogenic. Whereas the mitogenicity of acetyl PHA was impaired, the other derivatives were as active as untreated lectin. Binding studies of [125I]PHA and derivatives to purified pig spleen lymphocytes were performed at 37 °C. Results show that the lymphocytes bound various amounts of lectin derivatives. Maximum amounts were observed in the case of mitogenic lectins. Data were analyzed by Scatchard plots and two kinds of binding sites were found for PHA and the mitogenic derivatives. High affinity and low affinity binding sites have been characterized in terms of (apparent) association constant and (apparent) number of sites. Binding of PHA or its chemical derivatives to a small fraction (4.6–9.0%) of all available lectin binding sites was sufficient for maximal lymphocyte stimulation. Nonmitogenic PHA derivatives (nitro or N-bromosuccinimide) bound, with low affinity, to one kind of binding site. The relevancy of these findings to lymphocyte activation is discussed.