Direct measurement of surface charge distribution in phase separating supported lipid bilayers

Abstract

The local surface charge density of the cell membrane influences regulation and localization of membrane proteins. The local surface charge density could, until recently, not be measured directly under physiological conditions, and it was largely a hypothetical yet very important parameter. Here we use unsaturated lipids of a distinct charge (DOTAP, DOPC, and DOPG) and a neutral fully saturated lipid (DPPC) to create model membranes with phase separating domains of a defined charge. We then apply quantitative surface charge microscopy (QSCM) to investigate the local surface charge density; this is a technique based on a scanning ion conductance microscope (SICM) capable of measuring surface charge density with nanoscale lateral resolution. We are able to clearly distinguish lipid domains from charge and topography in all three model membranes. The measured surface charge densities furthermore reveal that disordered domains formed by charged lipids are in fact not only impure, but also incorporate uncharged saturated lipids. We estimate that at least 30% of disordered domains in DOPG : DPPC and DOTAP : DPPC will be DPPC. These ratios could present a limit for the formation of charged domains in lipid membranes.