Bioreactor droplets from liposome-stabilized all-aqueous emulsions

Abstract

Artificial bioreactors are desirable for in vitro biochemical studies and as protocells. A key challenge is maintaining a favourable internal environment while allowing substrate entry and product departure. We show that semipermeable, size-controlled bioreactors with aqueous, macromolecularly crowded interiors can be assembled by liposome stabilization of an all-aqueous emulsion. Dextran-rich aqueous droplets are dispersed in a continuous polyethylene glycol (PEG)-rich aqueous phase, with coalescence inhibited by adsorbed B130-nm diameter liposomes. Fluorescence recovery after photobleaching and dynamic light scattering data indicate that the liposomes, which are PEGylated and negatively charged, remain intact at the interface for extended time. Inter-droplet repulsion provides electrostatic stabilization of the emulsion, with droplet coalescence prevented even for submonolayer interfacial coatings. RNA and DNA can enter and exit aqueous droplets by diffusion, with final concentrations dictated by partitioning. The capacity to serve as microscale bioreactors is established by demonstrating a ribozyme cleavage reaction within the liposome-coated droplets.

Microscopic agents programmed by DNA circuits

“…Compartments efficiently restrict the diffusion of reactive species 35 , but conversely require specific strategies in order to open communication channels between the units [36][37][38][39] . Living organisms elegantly manage transfer across their boundaries.…”

Aqueous Emulsion Droplets Stabilized by Lipid Vesicles as Microcompartments for Biomimetic Mineralization

“…These structures are formed when ~100 nm diameter lipid vesicles (large unilamellar vesicles, or LUVs) self-assemble at 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 -4 -the aqueous/aqueous interface of a PEG/dextran biphasic system [29][30][31][32] , stabilizing enzyme-loaded droplets of dextran-rich phase in a continuous medium of PEG-rich phase. 33 and does not require membrane transporters or pore-forming molecules. 33 The combination of lipidstabilized, enzyme-loaded all-aqueous emulsion droplets with intermediate chelation provides excellent control over the mineralization process, enabling high-yield production of relatively monodisperse AMV populations.…”

“…33 and does not require membrane transporters or pore-forming molecules. 33 The combination of lipidstabilized, enzyme-loaded all-aqueous emulsion droplets with intermediate chelation provides excellent control over the mineralization process, enabling high-yield production of relatively monodisperse AMV populations. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 Scheme 1.…”

Combining Catalytic Microparticles with Droplets Formed by Phase Coexistence: Adsorption and Activity of Natural Clays at the Aqueous/Aqueous Interface

“…Candidates for early-Earth compartments include crevices or pores in rocks 17 , self-assemblies of lipids or simpler amphiphiles to form vesicles 18–23 , and aqueous droplets formed by liquid-liquid phase separation 15, 21, 24 . Combining catalytic mineral surfaces with compartments such as amphiphile vesicles or droplets formed by aqueous/aqueous phase coexistence is therefore appealing as primitive protocell models.…”

“…The immiscible aqueous phases that result upon demixing in aqueous polymer solutions provide compartments with different chemical and physical properties between which solutes such as biomolecules (or their prebiotic precursors) can partition. Consequently, aqueous two-phase systems (ATPS) have been used for biomolecule separations 25, 26 , food science 27, 28 , and biomimetic compartmentalization 16, 18, 29 , and have been proposed as simple protocell models 15, 24, 30 .…”