Electric field-induced circulation and vacuolization regulate enzyme reactions in coacervate-based protocells

Abstract

Artificial protocells operating under non-equilibrium conditions offer a new approach to achieve dynamic features with life-like properties. Using coacervate micro-droplets comprising polylysine (PLL) and a short single-stranded oligonucleotide (ss-oligo) as a membrane-free protocell model, we demonstrate that circulation and vacuolization can occur simultaneously inside the droplet in the presence of an electric field. The circulation is driven by electrohydrodynamics and applies specifically to the major components of the protocell (PLL and ss-oligo). Significantly, under low electric fields (E = 10 V cm) the circulation regulates the movement of the vacuoles, while high levels of vacuolization produced at higher electric fields can deform or reshape the circulation. By taking advantage of the interplay between vacuolization and circulation, we achieve dynamic localization of an enzyme cascade reaction at specific droplet locations. In addition, the spatial distribution of the enzyme reaction is globalized throughout the droplet by tuning the coupling of the circulation and vacuolization processes. Overall, our work provides a new strategy to create non-equilibrium dynamic behaviors in molecularly crowded membrane-free synthetic protocells.

Dynamic Synthetic Cells Based on Liquid–Liquid Phase Separation

“…Higher-Order Structures and Behaviour 6.1. [161] Dynamic vacuolisation was also observed in RNA/peptide coacervate droplets produced through DNA transcription, and attributed to ar eentrantp hase-transition process. This chemical diversity can produce vacuolisationa nd multiphase structurationw ithin condensates in living cells, in which multiple aqueous phases coexist withoutm ixing.…”

“…Dynamic substructures (vacuoles) have been observed within PLys/oligonucleotide coacervate microdroplets subjected to ac ontinuous electric field. [161] Dynamic vacuolisation was also observed in RNA/peptide coacervate droplets produced through DNA transcription, and attributed to ar eentrantp hase-transition process. [126,127] Studies on ternary (or more) mixtures of phase-separating components are still relatively scarce.…”

Interface Engineering in Multiphase Systems toward Synthetic Cells and Organelles: From Soft Matter Fundamentals to Biomedical Applications

“…[245] External stimuli, such as UV and electric field, are engineered to investigate the nonequilibrium dynamics of coacervate droplets. [235,[246][247][248][249] Since the pioneer works of assembling aqueous two-phase droplets into lipid vesicles, [250][251][252] there also have been many attempts to encapsulate coacervate droplets in vesicles to further enhance their stability via electrostatic interactions between the membrane lipid and macromolecules in the coacervate (Figure 7c). [233,[253][254][255][256] These lipid membrane-coated coacervate droplets serve as advanced models for both cellular membranes and membraneless organelles.…”

“…[309] Additionally, the electric field also has potentially important impact on the behavior of synthetic membraneless coacervates. [246][247][248][249] Coacervates are resultant from electrostatic interactions of polyanions and polycations, such as nuclei acids. Applying electric field can induce complex and rich out-of-equilibrium phenomena, such as vacuolization, spontaneous splitting and circulations, which could be manipulated to regulate enzymatic reactions.…”

“…[ 245 ] External stimuli, such as UV and electric field, are engineered to investigate the nonequilibrium dynamics of coacervate droplets. [ 235,246–249 ]…”