Lipid bilayers suspended over microwells on Si substrates are promising platforms for nanobiodevices that mimic cell membranes. Using the biotin-avidin interaction, we have succeeded in selectively arranging vesicles on the freestanding region of a lipid bilayer. When ternary lipid mixtures of saturated lipid, unsaturated lipid, and cholesterol are used, they separate into liquid-order (Lo) and liquid-crystalline (Lα) domains. A freestanding lipid bilayer prefers the Lα-phase over the Lo-phase because of the difference in their flexibility. In addition, the type of biotinylated lipid determines whether it is localized in the Lα-phase domain or the Lo-phase domain. As a result, the biotinylated unsaturated lipids localized in the Lα-phase domain aggregate in the freestanding lipid bilayer, and vesicles labeled with biotin selectively bind to the freestanding lipid bilayer by the biotin-avidin interaction. This technique helps to introduce biomolecules into the freestanding lipid bilayer of nanobiodevices via vesicles.
Vesicle fusion plays an important role in molecular transport and signal transduction in vivo. Fusion occurs even in artificial vesicles without proteins, as long as cations are present similar to the physiological environment. The progress of fusion between vesicles encapsulating a fluorescent dye (calcein) and giant unilamellar vesicles (GUVs) was examined using a fluorescence microscope. In the case of vesicles formed with phosphatidylcholine (PC) alone, the fusion begins upon the suppression of electrostatic repulsion by cations such as Ca 2+ . However, almost all vesicles remain in the state of adsorption or hemifusion even after 1 h or more, and it is difficult to proceed to full fusion. In contrast, mixing phosphatidylethanolamine (PE) with vesicles facilitates the progression from hemifusion to full fusion, and the transfer of calcein to the GUV was observed immediately after beginning the vesicle fusion. This is probably because the small head group makes the fluid phase lipid bilayer unstable.
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