Abstract
In the field of bottom-up synthetic biology, lipid membranes are the scaffold to create minimal cells and mimic reactions and processes at or across the membrane. In this context, we employ here a versatile microfluidic platform that enables precise positioning of nanoliter droplets with user-specified lipid compositions and in a defined pattern. Adjacent droplets make contact and form a droplet interface bilayer to simulate cellular membranes. Translocation of molecules across membranes are tailored by the addition of alpha-hemolysin to selected droplets. Moreover, we developed a protocol to analyze the translocation of non-fluorescent molecules between droplets with mass spectrometry. Our method is capable of automated formation of one- and two-dimensional droplet networks, which we demonstrated by connecting droplets containing different compound and enzyme solutions to perform translocation experiments and a multistep enzymatic cascade reaction across the droplet network. Our platform opens doors for creating complex artificial systems for bottom-up synthetic biology.
Highlights
In the field of bottom-up synthetic biology, lipid membranes are the scaffold to create minimal cells and mimic reactions and processes at or across the membrane
One of the key challenges in the creation of artificial cells is the formation of a cell-like architecture, where different biochemical compounds are retained in smaller organelles within the cell and each organelle has a specific biological function
Aqueous droplets of ~25 nanoliters were generated in hexadecane/squalane with dissolved lipids
Summary
In the field of bottom-up synthetic biology, lipid membranes are the scaffold to create minimal cells and mimic reactions and processes at or across the membrane In this context, we employ here a versatile microfluidic platform that enables precise positioning of nanoliter droplets with user-specified lipid compositions and in a defined pattern. Many pathways require multiple steps in different organelles, translocation of intermediate products between organelles, and signaling cascades to enable the cell to sense the environment or relay information between different cellular regions In this regard, major achievements were presented recently, reporting the realization of multistep reactions[4,5], genetic circuits[8,9,10], controlled division[11] or communication pathways across membranes[12,13,14] in artificial cell models. The α-HL pores are used to connect selected compartments with each other
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