FIGURE 5.

Membrane-delimited active RhoA facilitates activation of membrane localized RhoA by soluble RhoGEF. A, increasing amounts of RhoA-His₆-GTPγS were reconstituted on the surface of phospholipid vesicles along with RhoA-His₆-mant-GDP (1 μm, substrate) as described under “Experimental Procedures.” Facilitated dissociation of mant-GDP from the preloaded RhoA was initiated by addition of the PRG-DHPH domains (10 nm) and monitored by the decrease in fluorescence of dissociated mant-GDP. Concentrations of activated RhoA for representative curves are noted in the legend. B, initial rates for dissociation of mant-GDP were determined as described under “Experimental Procedures” and plotted against the concentration of RhoA-His₆-GTPγS bound to vesicles.