FIGURE 5.

Role of membrane deformation in the effect of the BAR domain on GAP activity. A, effect of BAR-PZA on the geometry of LUVs lacking cholesterol and containing saturated lipids. Negative stained electron microscopy was used to analyze the shape of LUVs containing 55% DSPC, 20% PE, 15% PS, 7.5% PI, and 2.5% PIP₂ formed by extrusion through 0.1-μm pores. The LUVs were incubated with either 0.5 μm BAR-PZA or 0.5 μm PZA. B, determination of size of vesicles used in experiments. The autocorrelation function of dynamic light scattering of LUVs is shown. LUVs containing either egg PC or DSPC and prepared by extrusion through a membranes of either 0.1- or 1.0-μm pores were analyzed by dynamic light scattering as described under “Experimental Procedures.” C, effect of BAR-PH on the activity of PZA. BAR-PH was titrated into a reaction containing 0.1 nm PZA, LUVs with 0.5% PIP₂, and myrArf1·GTP as a substrate. The relative GAP activity was estimated as the ln(GTP present at time 0/GTP) as described previously (29). BAR-PH would not be predicted to bind to PZA (33).