KINETIC OF LIGAND-BINDING TO THE TYPE-1 FC-EPSILON RECEPTOR ON MAST-CELLS

Rates of association and dissociation of several specific monovalent ligands to and from the type I Fc-epsilon-receptor (Fc-epsilon-RI) were measured on live mucosal type mast cells of the rat line RBL-2H3. The ligands employed were a monoclonal murine IgE and Fab fragments prepared from three different, Fc-epsilon-RI-specific monoclonal IgG class antibodies. These monoclonals (designated H10, J17, and F4) were shown previously to trigger mediator secretion by RBL-2H3 mast cells upon binding to and dimerization of the Fc-epsilon-RI. Analysis of the kinetics shows that the minimal mechanism to which all data can be fitted involves two consecutive steps: namely, ligand binding to a low-affinity state of the receptor, followed by a conformational transition into a second, higher affinity state h of the receptor-ligand complex. These results resolve the recently noted discrepancy between the affinity of IgE binding to the Fc-epsilon-RI as determined by means of binding equilibrium measurements [Ortega et al. (1988) EMBO J. 7, 4101] and the respective parameter derived from the ratio of the rate constant of rat IgE dissociation and the initial rate of rat IgE association [Wank et al. (1983) Biochemistry 22, 954]. The probability of undergoing the conformational transition differs for the four different Fc-epsilon-RI-ligand complexes: while binding of Fab-H10 and IgE favors the h state, binding of Fab-J17 and Fab-F4 preferentially maintains the low-affinity 1 state (at 25-degrees-C). The temperature dependence of the ligand interaction kinetics with the Fc-epsilon-RI shows that the activation barrier for ligand association is determined by positive enthalpic and entropic contributions. The activation barrier of the 1 --> h transition, however, has negative enthalpic contributions counteracted by a decrease in activation entropy. The h --> 1 transition encounters a barrier that is predominantly entropic and similar for all ligands employed, thus suggesting that the Fc-epsilon-RI undergoes a similar conformational transition upon binding any of the ligands.