Core-shell coacervates formed from DNA nanostars

Phase-separating DNA coacervates have important potential as model protocells, uniquely combining tuneable material properties with programmable biomolecular interactions. However, the membrane-free nature of DNA coacervates leads to instability and heterogeneity, limiting their ability to mimic cell behaviour. Here, we develop multi-layered DNA coacervate ‘droplets’ composed of DNA nanostars, where nanostars with distinct DNA sequences form both the cytosolic core and the membrane-like shell. Nanostar properties were first explored to understand how structural changes in geometry, valency, and interaction strength affect phase-separation temperature, size, stability, and permeability. We show that when pairs of nanostars self-assemble in the same solution, the order of phase-separation determines core or shell destination, while the proportion of surfactant nanostars that link the two populations determines shell morphology. For 3 different core nanostars, we demonstrate a range of shell nanostars with different material properties and morphologies. Membrane-like systems, where the shell fully encloses the core, form if the difference in phase-separation temperatures of the two nanostars is greater than 7C with 16-25% surfactant nanostars. Core-shell droplets were more cell-like, with well-defined size, stability over time, and core permeability controlled by shell properties. Furthermore, droplet size and membrane thickness were controlled by adjusting the thermal annealing rate during assembly. These techniques provide a diverse library of droplets suitable to be used as protocells with predictable size, mono-dispersity, membrane thickness, and permeability. Core-shell DNA nanostar droplets will open new avenues for programming dynamic cell-like behaviour in simplified systems that combine DNA protocells with DNA molecular circuits.