Mechanism of human lymphocyte stimulation by concanavalin A: role of valence and surface binding sites.

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Abstract
A monovalent form of concanavalin A (m-Con A) was prepared to determine the importance of valence for human lymphocyte surface binding and subsequent lymphocyte stimulation as measured by blast transformation and cytotoxicity. Concanavalin A (Con A) was fragmented by a proteolytic process, and the m-Con A derivative was isolated by elution with an ascending D-glucose gradient on a Sephadex G-200 column. The MW of m-Con A was 18,00 by sodium dodecyl sulfate-polyacrylamide electrophoresis. Equilibrium dialysis with .alpha.-methyl D-glucoside and subsequent Scatchard plot analysis revealed an association constant (Ka) of 1.2 .times. 103 liters/mol and a valence of 1.1. Incubation of lymphocytes with 125I-labeled m-Con A demonstrated surface binding at 1.21 .times. 106 molecules/cell, which was comparable to the binding of [3H]Con A (1.02 .times. 106 molecules/cell). In contrast to the intact lectin, m-Con A had a markedly reduced capacity to agglutinate rabbit erythrocytes and human lymphocytes, and did not stimulate lymphocyte blast transformation or cytotoxicity at 1 and 10 .mu.g/ml. Pretreatment of lymphocytes with m-Con A blocked blast transformation induced by Con A. Thus, m-Con A binds to lymphocyte surface receptors, but does not stimulate blast transformation or cytotoxicity; Con A probably bridges binding sites on the lymphocyte surface to induce lymphocyte activation.