The tether compound/backfiller ratio impacts protein pore formation in the artificial lipid membrane

The tether that anchors the lipid bilayer to the solid surface is a key component of tethered lipid membranes, which mimic biological membranes. The obtained lipid bilayers with high electrical sealing properties facilitate the application of electrochemical impedance spectroscopy (EIS) to assess conductivity and defectiveness resulting, for example, from pore-forming toxins. However, it remains uncertain whether the tether compound in the lipid membrane impacts only EIS sensitivity or also influences analyte (protein) function. Thus, we investigated how the lipid membrane properties change by varying the amount and the length of the tether compound. Following this, the membranes were exposed to the pore-forming toxin pneumolysin produced by Streptococcus pneumoniae. Several methods were used, including surface plasmon resonance for binding analysis, surface-enhanced infrared absorption spectroscopy for structural insights, EIS for quantifying the pores, and atomic force microscopy for morphological characterization. The study highlights two key observations. First, the composition of mixed self-assembled monolayers deviates from that of the preparative solution. Second, the quantity of tether compounds present in the lipid membrane influences protein oligomerization due to the minor steric hindrance introduced by these compounds. These insights should be helpful in utilizing tethered lipid membranes as a tool to mimic cell membrane viscosity