Supported Lipid Bilayers on Skeletonized Zirconium Phosphonate Surfaces For the Study of Transmembrane Proteins

Supported lipid bilayers that can fully represent biological cell membranes are attractive biomimetic models for biophysical and biomedical applications. In previous work, we developed a new approach to engineer stable supported lipid membranes. This system uses the zirconium phosphonate substrate as a reactive surface that tethers the lipid membrane via a highly covalent bond between surface zirconium ions and divalent phosphate groups in the lipid assembly. An advantage of this approach is that the zirconium phosphonate can be applied to any surfaces (gold, glass, silicon) allowing different analytical techniques to be used on the same system. However, the covalent interaction between the zirconium phosphonate film and the inner lipid monolayer that is responsible for the bilayer stability restricts membrane components such as transmembrane proteins from penetrating into the membrane and retaining their mobility. In contrast to other approaches that incorporate pillars to support the bilayers, our strategy involves the creation of reservoirs beneath the supported lipid bilayers to accomodate transmembrane proteins. Skeletonized zirconium phosphonate surfaces have been designed using the Langmuir-Blodgett (LB) technique. Sizes of the nanometer-holes were varied by controlling the amount of octadecylphosphonic acid in mixed LB layers and characterized by atomic force microscopy. The skeletonization of the film was optimized so the hollow spaces were large enough to incorporate the proteins but small enough for the membrane to bridge the spaces. The efficiency of this system as a support for transmembrane proteins has been characterized by surface plasmon resonance enhanced ellipsometry (SPREE) using the proteins Annexin V and the maxi-K ion channel to demonstrate the utility of the system as a functional cell membrane. This concept is innovative in the area of bilayer platforms and can be used with membrane and transmembrane proteins.