Specificity and competitive cation association to Phosphatidylinositol‐4,5‐bisphosphate model membranes

Abstract

Phosphatidylinositol‐4,5‐bisphosphate (PIP2) is an active signaling lipid implicated, among other functions, in the regulation of cell growth by activating the tumor suppressor PTEN. Using synchrotron surface‐sensitive x‐ray diffraction and fluorescence techniques we determined the preferential cation binding to PIP2 monolayers. The natural, highly unsaturated PIP2 was spread as a Langmuir monolayer on a physiological buffer containing 100 mM KCl at pH 7.2 to which divalent cations (Ca2+ and Mg2+) were added. X‐ray fluorescence of the PIP2 monolayer on the buffer shows an eight fold surface enhancement of monovalent K+ compared to its bulk concentration. When physiological levels of calcium are added (1‐100 µM), the Ca2+ gradually replaces bound K+ ions, leading to a significant change in the organization of the PIP2 model membrane. At higher concentrations (100‐1000 µM), which might be achieved during calcium signaling, we observe a 1000 fold surface enhancement of Ca2+. Similar experiments with Mg2+ ions also show strong ion binding to PIP2 at physiological levels (1 mM) with a lesser structural effect on the monolayer compared to that induced by Ca2+. For mixed solutions of Mg2+ and Ca2+ we find that Ca2+ occupies the majority of binding sites, and at mM concentrations completely removes the Mg2+ ions from the interface. Surprisingly, with both 1 mM Mg2+ and 1 mM Ca2+ in the subphase there is still a fourfold surface enrichment of K+ ions at the headgroup region.