RAC2[E62K] has altered TIAM1-mediated GDP exchange and p50RhoGAP-mediated GTP hydrolysis. (A) GDP exchange assay using RAC2[WT] (red) and RAC2[E62K] (blue) preloaded with fluorescent ^mantGDP and incubated with unlabeled GDP. Intrinsic nucleotide exchange (open circles) and TIAM1-mediated exchange (filled circles) are shown. Ratio TIAM1:RAC2 = 1:1. (B) ^mantGDP dissociation rate of RAC2[WT] and RAC2[E62K] with and without TIAM1 calculated from panel A. Number of replicates, N = 4 for intrinsic dissociation and N = 2 for TIAM1-mediated dissociation. ***P < .0005. (P value calculated using the 1-way ANOVA followed by Tukey multiple comparison test). (C) GTP hydrolysis of RAC2[WT] and RAC2[E62K] preloaded with GTP without and with p50RhoGAP (1:500, p50RhoGAP:RAC2). Colored as in panel A. (D) GTP hydrolysis rate of RAC2[WT] and RAC2[E62K] preloaded with GTP, calculated from panel C. Data acquired in triplicate. **P < .005 (P value calculated as in panel B).