Looping-in complexation and ion partitioning in nonstoichiometric polyelectrolyte mixtures

Abstract

A wide variety of intracellular membraneless compartments are formed via liquid-liquid phase separation of charged proteins and nucleic acids. Understanding the stability of these compartments, while accounting for the compositional heterogeneity intrinsic to cellular environments, poses a daunting challenge. We combined experimental and theoretical efforts to study the effects of nonstoichiometric mixing on coacervation behavior and accurately measured the concentrations of polyelectrolytes and small ions in the coacervate and supernatant phases. For synthetic polyacrylamides and polypeptides/DNA, with unequal mixing stoichiometry, we report a general “looping-in” phenomenon found around physiological salt concentrations, where the polymer concentrations in the coacervate initially increase with salt addition before subsequently decreasing. This looping-in behavior is captured by a molecular model that considers reversible ion binding and electrostatic interactions. Further analysis in the low-salt regime shows that the looping-in phenomenon originates from the translational entropy of counterions that are needed to neutralize nonstoichiometric coacervates.

Closed-Loop Phase Behavior of Nonstoichiometric Coacervates in Coarse-Grained Simulations

“…With addition of salt to the nonstoichiometric mixture, the coacervate phase first shrinks in the low-salt regime and then swells with further addition of salt (see Figure (right)). This behavior is consistent with experiments of Friedowitz et al using coacervates of synthetic polyacrylamides. Shrinking with low added salt is not observed for symmetrically charged coacervates, which only swell as salt is added.…”

“…We first vary the “mixing ratio”, i.e., the ratio of polycation and polyanion chains of equal length and charge densities, to investigate the impact of asymmetry on phase behavior. We then add salt to such nonstoichiometric systems and find a closed-loop phase behavior, which we compare to recent experimental results on synthetic polyacrylamide coacervates . Although there are different ways of breaking symmetry as mentioned above, in this work, we introduce asymmetry only by varying stoichiometry.…”

Influence of “Hydrophobicity” on the Composition and Dynamics of Polyelectrolyte Complex Coacervates

“…The high density of charges within PECs has stimulated much recent theory − and experiment − aimed at understanding both the driving forces for the formation of PECs from solution and their physical properties. While these driving forces are often assumed to be electrostatic in nature, entropy actually dominates PEC association: , the release of polyelectrolyte counterions offers much more entropy change than does mixing two polymers …”

Interfacial Structure and Tension of Polyelectrolyte Complex Coacervates

“…Polyelectrolyte complex coacervation refers to an associative liquid–liquid phase separation in solutions of oppositely charged polyelectrolytes into a polymer-rich liquid coacervate phase and an equilibrium polymer-deficient supernatant phase. − Due to its fundamental role in a variety of fields such as hydrogel fabrication, − drug delivery, , and formation of membrane-less organelles in cells, − polyelectrolyte complex coacervation has received a great deal of attention in recent years from both the polymer physics and biophysics communities, and significant progresses have been made in understanding the bulk phase behaviors. ,− For example, it is now well accepted that the addition of monovalent salt can weaken the degree of phase separation, and the miscibility gap vanishes above a critical salt concentration. , Likewise, the effects of other parameters such as temperature, charge fraction, chain length, polymer concentration, sequence, and stoichiometry on the bulk properties of complex coacervation have been extensively examined in the literature; we refer to several recent excellent reviews ,− for these progresses.…”