ASYMMETRIC LARGE AREA MODEL BIOMEMBRANES
All biological cell membranes maintain an electric transmembrane potential of around 100 mV, due in part to an asymmetric distribution of charged phospholipids across the membrane. This asymmetry is crucial to cell health and physiological processes such as intracell signaling, receptor-mediated endocytosis, and membrane protein conformation and function as well as active processes involving flippase and floppase proteins. Despite the biological significance, there are limited studies linking the consequences of lipid asymmetry to critical membrane properties and processes involving ion channels. One reason for this is the scarcity of reliable methods to create artificial membrane systems that incorporate both transverse lipid asymmetry and ion channels. Experimental artificial membrane systems incorporate essential cell membrane structures, namely the phospholipid bilayer, in a controllable manner where specific properties and processes can be isolated and examined in an environment much simpler than living systems. It is of particular interest to study asymmetry in transverse lipid composition across the phospholipid bilayer on such a system to probe the effects of the lipid composition and asymmetric arrangement of these lipids on the physicochemical properties of the membrane. By doing so, an understanding of how membrane asymmetry dictates membrane properties and in turn impacts cellular processes will be achieved.
The primary goal of this thesis is to develop a platform for fabricating and characterizing compositionally controlled planar, free-standing, asymmetric membranes. This asymmetry was qualitatively demonstrated using a fluorescence quenching assay, and it has been quantified using a combination of anionic and zwitterionic lipids in concert with a patch-clamp amplifier system. Initial measurements of a transmembrane potential on a partially asymmetric bilayer were found to be between 10 and 25 mV. Increasing membrane charge asymmetry increases the offset voltage, as expected, and also modifies the stiffness of the membrane. These initial successes demonstrate a viable pathway to fabricate and quantitatively characterize asymmetric bilayers that can be extended to accommodate more complex membrane processes in the future.