Abstract:We analyze experimentally determined phase diagram of
γD–βB1 crystallin
mixture. Proteins are described as dumbbells decorated with attractive sites to
allow inter–particle interaction. We use thermodynamic perturbation
theory to calculate the free energy of such mixtures and, by applying
equilibrium conditions, also the compositions and concentrations of the
co–existing phases. Initially we fit the
Tcloud versus packing fraction
η measurements for pure
(x2 = 0) γD
solution in 0.1 M phosphate buffer at pH = 7.… Show more
“…1a ). These particles, with attractive patches decorated on a hard core, which in our case is spherical, have been used to model proteins in LLPS 30 – 33 . Figure 1 Illustration of the patchy particle model for protein-regulator mixtures and its two phases.…”
Recently many cellular functions have been associated with membraneless organelles, or protein droplets, formed by liquid-liquid phase separation (LLPS). Proteins in these droplets often contain RNA-binding domains, but the effects of RNA on LLPS have been controversial. To gain better understanding on the roles of RNA and other macromolecular regulators, here we used Gibbs-ensemble simulations to determine phase diagrams of two-component patchy particles, as models for mixtures of proteins with regulatory components. Protein-like particles have four patches, with attraction strength εPP; regulatory particles experience mutual steric repulsion but have two attractive patches toward proteins, with the strength εPR tunable. At low εPR, the regulator, due to steric repulsion, preferentially partitions in the dispersed phase, thereby displacing the protein into the droplet phase and promoting LLPS. At moderate εPR, the regulator starts to partition and displace the protein in the droplet phase, but only to weaken bonding networks and thereby suppress LLPS. At εPR > εPP, the enhanced bonding ability of the regulator initially promotes LLPS, but at higher amounts, the resulting displacement of the protein suppresses LLPS. These results illustrate how RNA can have disparate effects on LLPS, thus able to perform diverse functions in different organelles.
“…1a ). These particles, with attractive patches decorated on a hard core, which in our case is spherical, have been used to model proteins in LLPS 30 – 33 . Figure 1 Illustration of the patchy particle model for protein-regulator mixtures and its two phases.…”
Recently many cellular functions have been associated with membraneless organelles, or protein droplets, formed by liquid-liquid phase separation (LLPS). Proteins in these droplets often contain RNA-binding domains, but the effects of RNA on LLPS have been controversial. To gain better understanding on the roles of RNA and other macromolecular regulators, here we used Gibbs-ensemble simulations to determine phase diagrams of two-component patchy particles, as models for mixtures of proteins with regulatory components. Protein-like particles have four patches, with attraction strength εPP; regulatory particles experience mutual steric repulsion but have two attractive patches toward proteins, with the strength εPR tunable. At low εPR, the regulator, due to steric repulsion, preferentially partitions in the dispersed phase, thereby displacing the protein into the droplet phase and promoting LLPS. At moderate εPR, the regulator starts to partition and displace the protein in the droplet phase, but only to weaken bonding networks and thereby suppress LLPS. At εPR > εPP, the enhanced bonding ability of the regulator initially promotes LLPS, but at higher amounts, the resulting displacement of the protein suppresses LLPS. These results illustrate how RNA can have disparate effects on LLPS, thus able to perform diverse functions in different organelles.
“…These particles, with attractive patches decorated on a hard core, which in our case is spherical, have been used to model proteins in LLPS. [30][31][32][33] The details of the protein and regulator models were chosen based on a number of observations regarding LLPS of protein-regulator mixtures. First of all, in many experimental studies where LLPS has been demonstrated, there exists a single "driver" protein that can form droplets on its own, whereas RNA or other regulatory components can modulate the phase boundaries.…”
Recently many cellular functions have been associated with membraneless organelles, or protein droplets, formed by liquid-liquid phase separation (LLPS). Proteins in these droplets often contain RNA-binding domains, but the effects of RNA on LLPS have been controversial. To gain better understanding on the roles of RNA, here we used Gibbs-ensemble simulations to determine phase diagrams of two-component patchy particles, as models for mixtures of proteins with RNA or other regulatory components. Protein-like particles have four patches, with attraction strength ε PP ; regulatory particles experience mutual steric repulsion but have two attractive patches toward proteins, with the strength ε PR tunable. At low ε PR , the regulator, due to steric repulsion, preferentially partitions in the dispersed phase, thereby displacing the protein into the droplet phase and promoting LLPS. At moderate ε PR , the regulator starts to partition and displace the protein in the droplet phase, but only to weaken bonding networks and thereby suppress LLPS. At ε PR > ε PP , the enhanced bonding ability of the regulator initially promotes LLPS, but at higher amounts, the resulting displacement of the protein suppresses LLPS. These results illustrate how RNA can have disparate effects on LLPS, thus able to perform diverse functions in different organelles.
“…[138,139] These studies indicate that LLPS is sensitiven ot only to the net electric charge of an IDP buta lso the pattern of charged istribution along its sequence. [140][141][142] Mean-field and more coarse-grained approaches that do not address sequence dependence at the residue level have also provided useful insights into the LLPS of globular proteins, [143,144] IDPs, and IDRs [21,63,[145][146][147] such as the role of cation-p interactions [61,63] and RNA. [148] Possible physicalo rigins of LLPS with LCSTsw ere addressed in some of these studies, [33,138] but the theoretical basis of pressure dependenceo fb iomolecular LLPS [42,44,98] has not been quantitatively explored.…”
Section: At Entative Rationalization Of T- P- and Tmao-dependentpromentioning
confidence: 99%
“…[35] Folded proteins are here represented in an abstract fashion similar to that employed in patchy-sphere models of liquid mixtures. [144,148,191] LLPS of folded proteins is envisioned to be driven by transienti nteractions of all types between protein surfaces. The association of differentr esidue types with different LLPS scenarios in Figure 13 is only an indication of general behavioral tendency.F or real proteins, all interaction types contribute, although some may have am ore dominant impact than the others, dependingo nt he amino acid sequence.…”
Section: At Entative Rationalization Of T- P- and Tmao-dependentpromentioning
Liquid–liquid phase separation (LLPS) of proteins and other biomolecules play a critical role in the organization of extracellular materials and membrane‐less compartmentalization of intra‐organismal spaces through the formation of condensates. Structural properties of such mesoscopic droplet‐like states were studied by spectroscopy, microscopy, and other biophysical techniques. The temperature dependence of biomolecular LLPS has been studied extensively, indicating that phase‐separated condensed states of proteins can be stabilized or destabilized by increasing temperature. In contrast, the physical and biological significance of hydrostatic pressure on LLPS is less appreciated. Summarized here are recent investigations of protein LLPS under pressures up to the kbar‐regime. Strikingly, for the cases studied thus far, LLPSs of both globular proteins and intrinsically disordered proteins/regions are typically more sensitive to pressure than the folding of proteins, suggesting that organisms inhabiting the deep sea and sub‐seafloor sediments, under pressures up to 1 kbar and beyond, have to mitigate this pressure‐sensitivity to avoid unwanted destabilization of their functional biomolecular condensates. Interestingly, we found that trimethylamine‐N‐oxide (TMAO), an osmolyte upregulated in deep‐sea fish, can significantly stabilize protein droplets under pressure, pointing to another adaptive advantage for increased TMAO concentrations in deep‐sea organisms besides the osmolyte's stabilizing effect against protein unfolding. As life on Earth might have originated in the deep sea, pressure‐dependent LLPS is pertinent to questions regarding prebiotic proto‐cells. Herein, we offer a conceptual framework for rationalizing the recent experimental findings and present an outline of the basic thermodynamics of temperature‐, pressure‐, and osmolyte‐dependent LLPS as well as a molecular‐level statistical mechanics picture in terms of solvent‐mediated interactions and void volumes.
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