Table 2.

Comparison of properties of the wild-type and designed Cdc42 proteins

	Cdc42-wt	Cdc42-des1	Cdc42-des2
MWtheor, kDa	22.1	22.1	22.1
MWAUC, kDa	21.7 ± 1.1	21.1 ± 0.7	21.3 ± 0.9
Stability
T1/2,CD, ° C
60 μg/ml	59 ± 1	68 ± 1	69 ± 1
310 μg/ml	56 ± 1	63 ± 1	66 ± 1
310 μg/ml in 2M urea	51 ± 1	59 ± 1	60 ± 1
T1/2,activity,° C	60 ± 2	70 ± 2	70 ± 2
T1/2,SEC,° C	57 ± 2	66 ± 2	66 ± 2
Activity, 25 ° C
Km, M	(8.2 ± 0.9)·10−3	(8.2 ± 0.9)·10−3	(8.2 ± 0.9)·10−3
kcat, hr−1	2.8 ± 0.1	2.6 ± 0.1	3.1 ± 0.2
Kd,app,GAP/GTP, μM	0.3 ± 0.1	0.4 ± 0.1	0.3 ± 0.1
Kd,app,GAP/GDP, μM*	5 ± 2	12 ± 4	8 ± 3

MW_theor molecular mass calculated from the amino acid sequence. MW_AUC molecular mass in solution was obtained from the analytical ultracentrifugation experiments as described (experimental data is given in Fig. S2). T_1/2,CD values were obtained from the CD experiments, one of which is shown in Figure 3C; T_1/2,activity values were obtained from the experiments shown in Figure 3D; T_1/2,SEC values were obtained from experiments shown in Figure 3E; K_m and k_cat were obtained from the fit of the initial velocity versus concentration of GTP plot by using Michaelis-Menten relationship (see Fig S5); K_{d,app,GAP/GTP} was obtained from the analysis of data given in Fig. 4; and K_{d,app,GAP/GDP} values were obtained from ITC experiments performed at 5° C (shown in Fig. S6).