Spectroelectrochemical Properties of Floating Lipid Membranes in Tris Organic Buffer

In biologically and biotechnologically relevant applications, the use of organic biological buffers, e.g. Tris or HEPES, is essential to maintain the activity and stability of biomacromolecules. Knowledge of how organic biological buffers affect the properties of free-standing floating bilayers is very limited. In this work, the properties of floating lipid bilayers were investigated in Tris buffer solution. The model lipid bilayer contains zwitterionic, neutral, and negatively charged lipids, representing the composition of the outer segment of photoreceptor cells. The electrochemical characterization of the floating bilayer (membrane resistance, membrane conductance, and capacitance) differs significantly between Tris and inorganic electrolyte solutions. To discover the differences in the supramolecular structure of the floating model membrane which are introduced by a selection of an electrolyte solution, in situ infrared spectroscopies: polarization modulation infrared reflection absorption spectroscopy and surface-enhanced infrared absorption spectroscopy techniques, were used. The weakly hydrated [TrisH]⁺ is conducted through the floating lipid bilayer independently of the applied potential. Accumulation of [TrisH]⁺ in the spacer layer leads to the outflux of hydrated water molecules once the pores in the membrane at negative potentials are formed. The results described in this manuscript provide crucial information on the tuning of the properties of the spacer layer allowing for incorporation of transmembrane proteins of different sizes in the extra-membranous space aiming at construction of model outer segment membranes.