Spatiotemporal control of signal-driven enzymatic reaction in artificial cell-like polymersomes

Abstract

Living cells can spatiotemporally control biochemical reactions to dynamically assemble membraneless organelles and remodel cytoskeleton. Herein, we present a microfluidic approach to prepare semi-permeable polymersomes comprising of amphiphilic triblock copolymer to achieve external signal-driven complex coacervation as well as biophysical reconstitution of cytoskeleton within the polymersomes. We also show that the microfluidic synthesis of polymersomes enables precise control over size, efficient encapsulation of enzymes as well as regulation of substrates without the use of biopores. Moreover, we demonstrate that the resulting triblock copolymer-based membrane in polymersomes is size-selective, allowing phosphoenol pyruvate to readily diffuse through the membrane and induce enzymatic reaction and successive coacervation or actin polymerization in the presence of pyruvate kinase and adenosine diphosphate inside the polymersomes. We envision that the Pluronic-based polymersomes presented in this work will shed light in the design of in vitro enzymatic reactions in artificial cell-like vesicles.

pH-Responsive Elastin-Like Polypeptide Designer Condensates

“…Such a protein sequence design approach, combined with microfluidic technology, can prove useful to understand the way natural cells form multiple, coexisting membraneless organelles in a dynamic fashion and to produce complex synthetic cells in the future. Given the recent surge of interest in forming coacervate-based organelles in biomimetic vesicles, – the presented pH-responsive ELP condensates will likely prove handy to the synthetic cell community. Ultimately, the pH-responsive nature of these condensates can be utilized to spatially and temporally regulate the function of enzymes, receptors, and signaling molecules, allowing for a highly precise control over biological processes in synthetic cells.…”

Coacervate Droplets for Synthetic Cells

“…Based on the capacity of some coacervates to reversible form and dissolve in response to a stimulus, various studies have demonstrated dynamic organelle assembly and biomolecular localization within membrane-bounded compartments, including waterin-oil emulsion droplets, GUVs (Figure 7a), and proteinosomes, e.g., in response to temperature, [194,215,[221][222][223] pH, [178,217,218] chemical, [212] and enzyme [170,178,181,219] reactions or light. [170,173] For instance, temperature-responsive polyU/spermidine coacervates were used to demonstrate storage and release of fluorescently labeled dsDNA strands within liposomes.…”

A Dissipative Supramolecular Glue for Temporal Control of Amplified Enzyme Activity and Biocatalytic Cascades

“…Temporal control of biomolecular components is key to several intracellular reaction networks, 1 robust signal transductions, 2 and efficient cell-cell communications. 3 Likewise, temporal regulation of enzyme activities allows dynamic control of biological transduction 4 and metabolism amongst other processes, helping elucidate biological events at the molecular level 5 and develop therapeutics with low side-effects. 6 While compartmentalized vesicles 7 or scaffold protein-mediated assembly of enzymes 8 allow organization of functional components inside the cell, dynamic regulation of activities requires transient assemblies in response to fuel consumption.…”