Towards biomimics of cell membranes: Structural effect of phosphatidylinositol triphosphate (PIP₃) on a lipid bilayer

Phosphoinositide (PIP) lipids are anionic phospholipids playing a fundamental role for the activity of several transmembrane and soluble proteins. Among all, phosphoinositol-3′,4′,5′-trisphosphate (PIP₃) is a secondary signaling messenger that regulates the function of proteins involved in cell growth and gene transcription. The present study aims to reveal the structure of PIP-containing lipid membranes, which so far has been little explored. For this purpose, supported lipid bilayers (SLBs) containing 1,2-dioleoyl-sn-glycero-3-phospho-(1′-myo-inositol-3′,4′,5′-trisphosphate (DOPIP₃) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) were used as mimics of biomembranes. Surface sensitive techniques, i.e. Quartz Crystal Microbalance with Dissipation monitoring (QCM-D), Atomic Force Microscopy (AFM) and Neutron Reflectometry (NR), provided detailed information on the formation of the SLB and the location of DOPIP₃ in the lipid membrane. Specifically, QCM-D and AFM were used to identify the best condition for lipid deposition and to estimate the total bilayer thickness. On the other hand, NR was used to collect experimental structural data on the DOPIP₃ location and orientation within the lipid membrane. The two bilayer leaflets showed the same DOPIP₃ concentration, thus suggesting the formation of a symmetric bilayer. The headgroup layer thicknesses of the pure POPC and the mixed POPC/DOPIP₃ bilayer suggest that the DOPIP₃-headgroups have a preferred orientation, which is not perpendicular to the membrane surface, but instead it is close to the surrounding lipid headgroups. These results support the proposed PIP₃ tendency to interact with the other lipid headgroups as PC, so far exclusively suggested by MD simulations.