Figure 2.
Direct SPA-binding data for the WASP GBD and mutant variants, with Cdc42. The indicated concentration of [3H]GTP-labeled Cdc42 was incubated with GST-tagged WASP GBD variants, as appropriate. The SPA signal was corrected by subtraction of the background signal from parallel measurements in which the effector protein was omitted. The effect of the concentration of Cdc42 on this corrected SPA signal was fitted to a binding isotherm to give an apparent Kd value and the signal at saturating Cdc42 concentrations. The data and curve fits are displayed as a percentage of this maximal signal. A, binding of representative mutants in the WASP GBD to Cdc42. B, binding of WASP BR hexa-mutant and C-terminal deletion mutant to Cdc42. 2–4 experimental replicates were performed for each WASP variant with 12 data points in each. A summary of all the binding data can be found in Table 2.