Supplementary Figure 5: Further analyses of the phase diagrams of the membrane-reconstituted Lck-CD45-CD3ζ network.

(a-d) Phase diagrams of the membrane-reconstituted Lck-CD45-CD3ζ network measured at different time points. Liposome reconstitution of Lck (WT) with CD45 and CD3ζ and FRET assays were performed as described in Fig. 5a,b and the % donor quenching at 30 min, 60 min, 90 min and 120 min after ATP addition were used to construct heat maps. As in Fig. 5c, the black dashed lines indicate conditions with equal molar ratio of Lck and CD45. The red dashed boxes highlight the regime with physiological densities of Lck and CD45. Similar phase diagrams were acquired in another experiment using independent materials. (e-g) Dose response plots for Lck (Y394F). FRET data as shown in Fig. 5b,c for Lck (Y394F) were plotted in the same manner as for Lck (WT) in Fig. 5d-f. (h-j) Dose response plots for Lck (Y505F). FRET data as shown in Fig. 5b,c for Lck (Y505F) were plotted in the same manner as for Lck (WT) in Fig. 5d-f. All data shown in this figure were fit with sigmoidal dose response curves (variable slopes) using Graphpad Prism 5.0 and the Hill-coefficients (n_H) are indicated with s.e.m. in brackets. Error bars represent s.e.m. from three independent measurements.