Self-Assembled Hybrid Nanomaterials: Interactions of Lipid Bilayers with Metal Oxide Surfaces of Nanoscale Curvature

Interfacing biological and man-made systems at the nanoscale level is essential for developing novel living-nonliving biotechnology platforms for applications in biology and medicine as well as for designing of biosensors. Despite of an impressive progress already achieved in designing new hybrid bio-nanomaterials including those based on self-assembling lipid bilayers, there is still a gap in understanding the influence of a nanostructured support and nanoconfinement on structure and properties of the membrane-protein interface. In this work we report on spin-labeling EPR studies to assess effects of solid inorganic interface, specifically, silica nanoparticle support and alumina nanotube confinement, on 1) the phospholipid membrane surface electrostatic potential and 2) effective pKa of the membrane-burred peptide ionisable sidechains. Novel EPR active pH-sensitive lipids IMTSL-PE and IKMTSL-PE were employed to measure the phospholipid membrane surface potential. The change in the protonation state of the label was directly observed by CW EPR allowing for determination of the effective pKa of the probe. Specific covalent attachment of EPR probes ensures that the spectroscopic signal originates from well-defined location at the interfaces. We have shown that by forming POPC or POPC/POPG mixed bilayers on the surfaces of silica nanoparticles the absolute value of the negative electric potential at the membrane surface could be increased significantly. The potential of mixed bilayer was observed to be more sensitive to the silica support, suggesting a different mechanism of the mixed bilayer response to the nanostructured surface. Only single protonation transition was observed for EPR pH-sensitive probe, thus, suggesting that both leaflets of the nanoparticle supported phospholipid bilayers have the same electrostatic surface potential. Addition of cholesterol to phospholipid bilayers did not diminish the bilayer response to silica. Effect the silica nanoparticle size on the lipid bilayer surface electrostatic potential was also observed for particles smaller than 100 nm. Effects of the silica support on the dynamics of lipids and transmembrane peptide have been also investigated. Specifically, a model transmembrane α-helical WALP peptide was covalently modified with cysteine-specific pH-sensitive nitroxides and incorporated into bilayers of various compositions. We have investigated the effect of placing a phospholipid bilayer with the integrated transmembrane α-helical WALP peptide on the surface of silica nanoparticles and inside nanochannels of anodic aluminum oxide on the peptide dynamics and the effective pKa of the probe. The silica support caused shift in the pKa of the probe consistent with the negative charge on the silica surface but induced a peptide transition upon the probe protonation not observed in liposomes.