Characterization of PA-binding proteins. A, Quantitative analysis of the binding of PA-sensor proteins TAPAS-1 and Raf-1 to different phospholipids (phospholipid strip assay; 100 pmol for each spotted lipid). Quantification of the level of TAPAS-1-GST and Raf-1-GST binding to individual lipids was determined from the integrated densities of spots on immunoblots. Values are given as arbitrary units (mean ± SEM; n = 4 independent experiments with proteins from three distinct protein preparations). B, Example of lipid dot blot with the indicated amounts of PA spotted onto nitrocellulose filters (5–100 nmol). Binding of TAPAS-1 wild type (lanes 1–6), TAPAS-1(WW19,20DD) (lanes 7–12), or TAPAS-1(WW19,20AA) (lanes 13–18) was detected with anti-GST antibodies. TAPAS-1 wild-type GST significantly bound to PA but not TAPAS-1(WW19,20AA)-GST or TAPAS-1(WW19,20DD)-GST. PE, Phosphatidylethanolamine; PC, phosphatidylcholine; PtdIns, phosphatidylinositol.