Crowded Environment Regulates the Coacervation of Biopolymers via Nonspecific Interactions.

The membrane-less organelles (MLOs) with subcompartments are formed via liquid-liquid phase separation (LLPS) in the crowded cell interior whose background molecules are up to 400 mg/mL. It is still a puzzle how the background molecules regulate the formation, dynamics, and functions of MLOs. Using biphasic coacervate droplets formed by poly(l-lysine) (PLL), quaternized dextran (Q-dextran), and single-stranded oligonucleotides (ss-oligo) as a model of MLO, we online monitored the LLPS process in Bovine Serine Albumin (BSA) solution up to 200.0 mg/mL. Negatively charged BSA is able to form complex or coacervate with positively charged PLL and Q-dextran and thus participates in the LLPS via nonspecific interactions. Results show that BSA effectively regulates the LLPS by controlling the phase distribution, morphologies, and kinetics. With increasing BSA concentration, the spherical biphasic droplets evolve in sequence into phase-inverted flower-like structure, worm-like chains, network structures, and confined coacervates. Each kind of morphology is formed via its own specific growth and fusion pathway. Our work suggests that MLOs could be controlled solely by the crowded environment and provides a further step toward understanding the life process in cell.

Coacervate Formed by an ATP-Binding Aptamer and Its Dynamic Behavior under Nonequilibrium Conditions.

Although numerous protocell models have been developed to explore the possible pathway of the origin of life on the early earth, few truly fulfill the roles of the DNA/RNA sequence and ATP molecules, which are keys to cell replication and evolution. The ATP-binding aptamer offers an opportunity to combine sequence and energy molecules. In this work, we choose the coacervate droplet as the protocell model and investigate the interaction of the DNA aptamer, poly(l-lysine)(PLL), and ATP under varying conditions. PLL and aptamers form solid precipitates, which spontaneously transform to coacervate droplets as ATP is introduced. The selective uptake and sequestration of exogenous molecules is achieved by the ATP-containing coacervates. As an electric field is applied to expel ATP, the portion of the droplet deficient in ATP becomes solid. The solid/liquid phase ratio is tunable by varying the electric field strength and excitation time. Together with the vacuolization process, a solid head with a soft mouth periodically opening and closing is created. Moreover, the composite coacervate droplet gradually grows as it is treated with an electric field and cannot recover to the original liquid phase after the power is turned off and replenished with ATP. Our work highlights that the proper integration of the DNA sequence, ATP, and energy input could be a powerful strategy for creating a protocell with certain living features.

Mass Transport in Coacervate-Based Protocell Coated with Fatty Acid under Nonequilibrium Conditions.

Construction of protocell models from prebiotically plausible components to mimic the basic features or functions of living cells is still a challenge. In this work, we prepare a hybrid protocell model by coating sodium oleate on the coacervate droplet constituted by poly(l-lysine) and oligonucleotide and investigate the transport of different molecules under electric field. Results show that sodium oleate forms a layered viscoelastic membrane on the droplet surface, which is selectively permeable to small, polar molecules, such as oligolysine. As the droplet is stimulated at 10 V cm-1, the oleate membrane slips along the direction of electric field while maintaining its integrity. Most of the molecules are still excluded under such conditions. As repetitive cycles of vacuolization occur at 20 V cm-1, all molecules are internalized and sequestrated in the droplet through their specific pathways except enzyme, which anchors in the oleate membrane and is immune to electric field. Cascade enzymatic reactions are then carried out, and the product generated from the membrane exhibits a time-dependent concentration gradient across the droplet. Our work makes a step toward the nonequilibrium functionalization of synthetic protocells capable of biomimetic operations.

Hydrogen-Bonded Polymer-Small Molecule Complexes with Tunable Mechanical Properties.

A novel type of polymeric material with tunable mechanical properties is fabricated from polymers and small molecules that can form hydrogen-bonded intermolecular complexes (IMCs). In this work, poly(vinyl alcohol) (PVA)-glycerol hydrogels are first prepared, and then they are dried to form IMCs. The tensile strengths and moduli of IMCs decrease dramatically with increasing glycerol content, while the elongations increase gradually. The mechanical properties are comparable with or even superior to those of common engineering plastics and rubbers. The IMCs with high glycerol content also show excellent flexibility and cold-resistance at subzero temperatures. Cyclic tensile and stress relaxation tests prove that there is an effective energy dissipation mechanism in IMCs and dynamic mechanical analysis confirms their physical crosslinking nature. FTIR and NMR characterizations prove the existence of hydrogen bonding between glycerol and PVA chains, which suppresses the crystallization of PVA from X-ray diffraction measurement. These PVA-glycerol IMCs may find potential applications in barrier films, biomedical packaging, etc., in the future.