Furuki, T. | bioRxiv Science

bioRxiv2023

Reentrant condensation of a multicomponent complex system of biomolecules induced by polyphosphate

Reentrant condensation (RC) is a phase behavior of protein solution comprising at least two components. In RC, a protein state varies from one phase to two phases and then back to one phase as the concentration of one component monotonically increases. To understand the phase behavior of multicomponent complex solutions of biomolecules, it is worth constructing an experimental multicomponent system that exhibits RC behavior. Here, we used a cola/milk mixture to investigate RC of a multicomponent complex system and explained the RC mechanism by reducing the system to two pure components, polyphosphate (polyP) and casein. In the multicomponent complex system, RC was observed with 20-60% cola and 1% milk. In the pure system, RC occurred with 0.01-2 mM tetraphosphate and 0.5 mg/ml casein. Moreover, the phase diagram showed that the condensation of casein depended on the chain length of the polyP. The present study succeeded in experimentally inducing RC in a multicomponent system and reproducing RC even when the system was reduced to its pure components. The fact that RC can be experimentally induced using common materials will provide important insights into the understanding of phase-separation behavior of biomolecules.

biochemistry↗

bioRxiv2022

Competitive membrane wetting of polymer blends in artificial cells initiates phase separation and promotes fractionation

Biomolecular condensates driven by liquid-liquid phase separation (LLPS) have received attention as novel activity regulators of living organisms. In intracellular LLPS, an important question is what type of biomolecules form condensates under what conditions. In this regard, possible interactions between biomolecules have been investigated. Recently, LLPS condensates have been reported to regulate the membrane structure upon wetting. However, the possibility of membrane wetting, in which the membrane conversely regulates the LLPS, remains unexplored. Using droplets of short polyethylene glycol and long dextran blends encapsulated with a lipid membrane, we demonstrate that membrane wetting regulates LLPS in cell-size spaces and alters the equilibrium state. In smaller droplets, the two-phase region expands beyond the bulk system, and the fractionation degree increases, particularly during the separation between short PEG and long dextran. We explain the space-size dependent LLPS based on the competitive membrane wetting between the polymers. Smaller droplets promote the membrane wetting of short PEG, which enhances the depletion force between long dextran molecules and finally induces LLPS. This shows that competition for membrane wettability among various molecules can regulate LLPS in cell-size spaces, rendering this LLPS principle feasible in living cells.

biophysics↗