Figure 4.

Dependence of PKCα and C2 domain membrane binding on PS mole density. Membrane binding density was quantified by single-molecule TIRF as described in the legend of Figure 3. A fixed concentration of full-length PKCα or C2 domain (5 pM) was added to supported PC bilayers containing increasing mole fractions of PS, and the density of fluorescent, membrane-bound proteins was measured for 20 temporally isolated frames from three separate movie streams in three separate titration experiments (n = 9). The best fit of the Hill equation yielded the indicated values of K_1/2 and the Hill coefficient. In both experiments, the free Ca²⁺ concentration was 6 μM in a physiological buffer (Materials and Methods) at 22 ± 0.5 °C.