SNARE‐Reconstituted Liposomes as Controllable Zeptoliter Nanoreactors for Macromolecules

Abstract

store enzymes, proteins, DNA, and various drug molecules. [2,3] These vesicles contain an aqueous lumen confined by a lipid bilayer, which acts as a permeable barrier to generate confined volumes ranging from zeptoliters (10 −21 L) to attoliters (10 −18 L). [4] Liposomes provide feasible fluidic solutions especially in areas where biocompatibility and miniaturization are critical. The zeptoliter-scale cavity creates a high effective concentration for encapsulated molecules. As a biomedical tool, liposomes exhibit little to no toxicity to biological organization and can preserve drug molecules in a stable environment. For these reasons, catalyst-loaded liposomes for intracellular enzymatic therapy are expected to be the next generation drug delivery systems. [5] Here, we show that liposomes can provide a suitable environment for reactions involving molecules smaller than the membrane's threshold pore size. Moreover, we demonstrate that protein-reconstituted liposomes, called proteoliposomes, can function as two-compartment nanoreactors for large macromolecules through soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein-mediated membrane fusion in a fast and regulated manner.While liposomes are seen as ideal nanocontainers, [3,6] their lipid membranes are impermeable to macromolecules, ions, and small molecules, essentially making the inner lumen a closed space. To overcome this limitation, scientists have tested the use of porous liposomes generated by membrane phase changes at the lipid phase transition temperature [7] or by pore formation proteins. [8] Porous liposomes are promising technology for nanoreactors since molecules smaller than the threshold pore size can enter the lumen to trigger reactions on demand.We demonstrate the size-selective permeability of porous liposomes through the protection of encapsulated DNAzyme, a catalytic DNA system isolated by in vitro selection methods, from degradation. [9] The experimental scheme is shown in Figure 2a. Cy3/Cy5 dual-labeled DNAzyme complex ( Figure S2, Supporting Information), whose self-cleavage activity depends on the presence of UO 2 2+ , [10] is encapsulated in liposomes composed of dimyristoyl phospatidylcholine (DMPC) lipid molecules. Due to the close proximity of donor (Cy3) and acceptor (Cy5) molecules, the initial Förster (fluorescence) resonance energy transfer (FRET) efficiency, E, is high Liposomes are synthetic phospholipid vesicles containing an aqueous lumen confined by a lipid bilayer. These vesicles have been used as nanoreactors because they offer confined environments that can result in significant changes to chemistry. However, a major limitation of using liposomes as nanocontainers is the impermeability of their lipid membranes. To overcome this, scientists have tested the use of porous liposomes generated by membrane phase changes or by pore formation proteins. In this study, the selective permeability of porous liposomes to molecules smaller than the threshold pore size is demonstrated for triggering int...

A Proteoliposome Method for Assessing Nanotoxicity on Synaptic Fusion and Membrane Integrity

“…We have developed an in vitro system that closely simulates this environment; our system incorporates liposomes mimicking synaptic vesicles and the presynaptic membrane, which are reconstituted with the corresponding SNAREs so that fusogenic activity is triggered by Ca 2+ . This vesicle system has been used extensively in single vesicle fusion assays, ensemble fluorescence experiments, and transmission electron microscopy (TEM) imaging experiments among others . This in vitro vesicle approach has been instrumental in elucidating the roles of specific proteins during synaptic fusion.…”

“…Content‐mixing experiments monitor the effects of nanomaterials on fusogenic activity. Analogous to neuronal exocytosis, fusion between SNARE‐reconstituted SV and PM vesicles is triggered by Ca 2+ . The experimental scheme is depicted in Figure A.…”

Interface Colloidal Deposition of Nanoparticle Wire Structures

“…Gold nanoparticles have proved to be effective building blocks for electronic sensing purposes because of their biocompatibility and the strong dependence of their electronic properties on capping molecules . Chemical and biological molecules, such as proteins and viruses, can also affect the electronic behavior of nanowires . Naturally, because of the high surface‐to‐volume ratio resulting from their porous structures, conducting gold nanowires made through interface colloidal deposition are promising for the electric detection of biological and chemical molecules ( Figure A,B) .…”