Formation of tethered bilayer lipid membranes on gold surface probed by in situ SEIRAS

Tethered bilayer lipid membranes (tBLMs) are versatile platforms for the analysis of biochemical and biophysical processes at biological membranes. To control the stability and functional properties of these artificial constructions, molecular-level knowledge on the organization and structure is required. We used surface-enhanced infrared absorption spectroscopy (SEIRAS) to elucidate the in situ formation of tBLMs on a gold substrate. The alkyl chains of long-chain anchoring thiol in a mixed self-assembled monolayer after 60 min of incubation in an adsorption methanol-d4 solution was found to be in a disordered state. Spectroscopic data revealed the complete formation of a bilayer after 60 min of incubation of a mixed anchoring monolayer in a phosphate buffer solution containing vesicles formed from partially deuterated lipid DPPC-d62 and cholesterol-d7. The temporal evolution of absorption bands from the lipid, anchoring mixed monolayer thiol, and water with increasing the bilayer formation time in the phosphate buffer solution containing vesicles revealed a two-stage process. Firstly, the adsorption of lipid molecules with a simultaneous withdrawal of water takes place at the interface. Secondly, the transformation of alkyl chains of the anchoring monolayer due to the insertion and interaction of lipids with the monolayer proceeds.