Hydrogel-supported bilayers for studying membrane protein functions

摘要

In vitro reconstitution of membrane proteins in lipid bilayers is thought to be an effective approach for understanding their function. Although a number of methods have been developed to create planer lipid bilayers and liposomes, each method has specific disadvantages. In this paper, we utilized hydrogels and reduced their disadvantages while retaining their original advantages. First, a lipid bilayer was formed between the surface of a hydrogel and a buffer solution with the contact method. Since the hydrogel supported the bilayer, stability is expected to be improved. Capacitance measurements evaluated the bilayer formation with an electrophysiological setup. The amplitude of the current responses reached up to 160 pA, suggesting that the surface area of the bilayer was approximately 8.3 × 10³ μm² (diameter of 50 μm). Second, the lipid bilayer was formed on patterned agarose using the self-spreading method. In this situation, the agarose works as a spacer between the membrane and substrate. Membrane formation was evaluated with fluorescence recovery after the photo bleaching method. Third, for future studies with membrane proteins, liposome formation was evaluated in various conditions, and we confirmed that liposomes could be formed in an in vitro protein synthesis solution.