Biological condensates form percolated networks with molecular motion properties distinctly different from dilute solutions.

Formation of membraneless organelles or biological condensates via phase separation and related processes hugely expands the cellular organelle repertoire. Biological condensates are dense and viscoelastic soft matters instead of canonical dilute solutions. To date, numerous different biological condensates have been discovered, but mechanistic understanding of biological condensates remains scarce. In this study, we developed an adaptive single-molecule imaging method that allows simultaneous tracking of individual molecules and their motion trajectories in both condensed and dilute phases of various biological condensates. The method enables quantitative measurements of concentrations, phase boundary, motion behavior, and speed of molecules in both condensed and dilute phases, as well as the scale and speed of molecular exchanges between the two phases. Notably, molecules in the condensed phase do not undergo uniform Brownian motion, but instead constantly switch between a (class of) confined state(s) and a random diffusion-like motion state. Transient confinement is consistent with strong interactions associated with large molecular networks (i.e., percolation) in the condensed phase. In this way, molecules in biological condensates behave distinctly different from those in dilute solutions. The methods and findings described herein should be generally applicable for deciphering the molecular mechanisms underlying the assembly, dynamics, and consequently functional implications of biological condensates.

Numerical Techniques for Applications of Analytical Theories to Sequence-Dependent Phase Separations of Intrinsically Disordered Proteins.

Biomolecular condensates, physically underpinned to a significant extent by liquid-liquid phase separation (LLPS), are now widely recognized by numerous experimental studies to be of fundamental biological, biomedical, and biophysical importance. In the face of experimental discoveries, analytical formulations emerged as a powerful yet tractable tool in recent theoretical investigations of the role of LLPS in the assembly and dissociation of these condensates. The pertinent LLPS often involves, though not exclusively, intrinsically disordered proteins engaging in multivalent interactions that are governed by their amino acid sequences. For researchers interested in applying these theoretical methods, here we provide a practical guide to a set of computational techniques devised for extracting sequence-dependent LLPS properties from analytical formulations. The numerical procedures covered include those for the determination of spinodal and binodal phase boundaries from a general free energy function with examples based on the random phase approximation in polymer theory, construction of tie lines for multiple-component LLPS, and field-theoretic simulation of multiple-chain heteropolymeric systems using complex Langevin dynamics. Since a more accurate physical picture often requires comparing analytical theory against explicit-chain model predictions, a commonly utilized methodology for coarse-grained molecular dynamics simulations of sequence-specific LLPS is also briefly outlined.

Rearrangement of dicarboranyl methyl cation to icosahedral C3 B9 H12 + : An ab initio dynamics view.

Closo-carborane anions are prominent, whereas the cations of the same are less abundant in the literature. As these ions have similar size and are weakly coordinating, the ionic liquids of these two ions could have important applications in many areas of chemistry. In view of limited number of polyhedral carborane cations available, we revisited the rearrangement of dicarboranyl methyl cation (7-CH2 7,9-nido-C2 B9 H10 + ) using ab initio molecular dynamics calculations with metadynamics. Our simulations confirmed the concerted mechanism of the rearrangement. We believe this work will resume the interest in its synthesis and carborane cations in general.

Analytical Formulation and Field-Theoretic Simulation of Sequence-Specific Phase Separation of Protein-Like Heteropolymers with Short- and Long-Spatial-Range Interactions.

A theory for sequence-dependent liquid-liquid phase separation (LLPS) of intrinsically disordered proteins (IDPs) in the study of biomolecular condensates is formulated by extending the random phase approximation (RPA) and field-theoretic simulation (FTS) of heteropolymers with spatially long-range Coulomb interactions to include the fundamental effects of short-range, hydrophobic-like interactions between amino acid residues. To this end, short-range effects are modeled by Yukawa interactions between multiple nonelectrostatic charges derived from an eigenvalue decomposition of pairwise residue-residue contact energies. Chain excluded volume is afforded by incompressibility constraints. A mean-field approximation leads to an effective Flory-Huggins χ parameter, which, in conjunction with RPA, accounts for the contact-interaction effects of amino acid composition and the sequence-pattern effects of long-range electrostatics in IDP LLPS, whereas FTS based on the formulation provides full sequence dependence for both short- and long-range interactions. This general approach is illustrated here by applications to variants of a natural IDP in the context of several different amino-acid interaction schemes as well as a set of different model hydrophobic-polar sequences sharing the same composition. Effectiveness of the methodology is verified by coarse-grained explicit-chain molecular dynamics simulations.

Role of Hoogsteen interaction in the stability of different phases of triplex DNA.

A simple coarse-grained model of DNA which includes both Watson-Crick and Hoogsteen base pairing has been used to study the melting and unzipping of triplex DNA. Using Langevin dynamics simulations, we reproduce the qualitative features of one-step and two-step thermal melting of triplex as seen in experiments. The thermal melting phase diagram shows the existence of a stable interchain three-strand complex (bubble-bound state). Our studies based on the mechanical unzipping of a triplex revealed that it is mechanically more stable compared to an isolated duplex-DNA.

Field theory description of ion association in re-entrant phase separation of polyampholytes.

Phase separation of several different overall neutral polyampholyte species (with zero net charge) is studied in solution with two oppositely charged ion species that can form ion pairs through an association reaction. Hereby, a field theory description of the system, which treats polyampholyte charge sequence dependent electrostatic interactions as well as excluded volume effects, is given. Interestingly, analysis of the model using random phase approximation and field theoretic simulation consistently shows evidence of a re-entrant polyampholyte phase separation at high ion concentrations when there is an overall decrease of volume upon ion association. As an illustration of the ramifications of our theoretical framework, several polyampholyte concentration vs ion concentration phase diagrams under constant temperature conditions are presented to elucidate the dependence of phase separation behavior on the polyampholyte sequence charge pattern as well as ion pair dissociation constant, volumetric effects on ion association, solvent quality, and temperature.

Subcompartmentalization of polyampholyte species in organelle-like condensates is promoted by charge-pattern mismatch and strong excluded-volume interaction.

Polyampholyte field theory and explicit-chain molecular dynamics models of sequence-specific phase separation of a system with two intrinsically disordered protein (IDP) species indicate consistently that a substantial polymer excluded volume and a significant mismatch of the IDP sequence charge patterns can act in concert, but not in isolation, to demix the two IDP species upon condensation. This finding reveals an energetic-geometric interplay in a stochastic, "fuzzy" molecular recognition mechanism that may facilitate subcompartmentalization of membraneless organelles.

A Simple Explicit-Solvent Model of Polyampholyte Phase Behaviors and Its Ramifications for Dielectric Effects in Biomolecular Condensates.

Biomolecular condensates such as membraneless organelles, underpinned by liquid-liquid phase separation (LLPS), are important for physiological function, with electrostatics, among other interaction types, being a prominent force in their assembly. Charge interactions of intrinsically disordered proteins (IDPs) and other biomolecules are sensitive to the aqueous dielectric environment. Because the relative permittivity of protein is significantly lower than that of water, the interior of an IDP condensate is expected to be a relatively low-dielectric regime, which aside from its possible functional effects on client molecules should facilitate stronger electrostatic interactions among the scaffold IDPs. To gain insight into this LLPS-induced dielectric heterogeneity, addressing in particular whether a low-dielectric condensed phase entails more favorable LLPS than that posited by assuming IDP electrostatic interactions are uniformly modulated by the higher dielectric constant of the pure solvent, we consider a simplified multiple-chain model of polyampholytes immersed in explicit solvents that are either polarizable or possess a permanent dipole. Notably, simulated phase behaviors of these systems exhibit only minor to moderate differences from those obtained using implicit-solvent models with a uniform relative permittivity equals to that of pure solvent. Buttressed by theoretical treatments developed here using random phase approximation and polymer field-theoretic simulations, these observations indicate a partial compensation of effects between favorable solvent-mediated interactions among the polyampholytes in the condensed phase and favorable polyampholyte-solvent interactions in the dilute phase, often netting only a minor enhancement of overall LLPS propensity from the very dielectric heterogeneity that arises from the LLPS itself. Further ramifications of this principle are discussed.

Antithrombotic Management in Ischemic Stroke with Essential Thrombocythemia: Current Evidence and Dilemmas.

Thrombotic diseases like ischemic stroke are common complications of essential thrombocythemia (ET) due to abnormal megakaryopoiesis and platelet dysfunction. Ischemic stroke in ET can occur as a result of both cerebral arterial and venous thrombosis. Management of ET is aimed at preventing vascular complications including thrombosis. Acute management of ischemic stroke in ET is the same as that in the general population without myeloproliferative disorders. However, an ET patient with ischemic stroke is at high risk for rethrombosis and is therefore additionally managed with cytoreductive therapy and antithrombotic agents. Given abnormal platelet production in ET, there is suboptimal suppression of platelets with the standard recommended dose of aspirin for cardiovascular (CV) prevention. Hence, for optimal CV protection in ET, low-dose aspirin is recommended twice daily in an arterial thrombotic disease like atherothrombotic ischemic stroke in presence of the following risk factors: age >60 years, Janus kinase2 V617F gene mutation, and presence of CV risk factors. In the presence of the same risk factors, concurrent antiplatelet and anticoagulant therapy is suggested for venous thrombosis. However, increased risk of bleeding with dual antithrombotic agents poses a significant challenge in their use in cerebral venous thromboembolism or atrial fibrillation in presence of the above-mentioned risk factors. We discuss these dilemmas regarding antithrombotic management in ischemic stroke in ET in this case-based review of literature in the light of current evidence.

Interfacial Polarization and Ionic Structure at the Ionic Liquid-Metal Interface Studied by Vibrational Spectroscopy and Molecular Dynamics Simulations.

Ionic liquids (ILs) have both fundamental and practical value in interfacial science and electrochemistry. However, understanding their behavior near a surface is challenging because of strong Coulomb interactions and large and irregular ionic sizes, which affect both their structure and energetics. To understand this problem, we present a combined experimental and computational study using a vibrational probe molecule, 4-mercaptobenzonitrile, inserted at the junction between a metal and a variety of ILs. The vibrational frequency of the nitrile in the probe molecule reports on the local solvation environment and the electrostatic field at this junction. Within the ethylmethyl imidazolium (EMIM+) cation family of ILs, we varied the anions over a range of sizes and types. Complementing our surface spectroscopy, we also ran molecular dynamics simulations of these interfaces to better understand the ionic structures that produced the measured fields. The magnitude of the frequency shifts, and thereby fields, shows a general correlation with the size of anions, with larger anions corresponding to smaller fields. We find that the source of this correlation is partial intercalation of smaller anions into the probe monolayer, resulting in tighter packing of ionic layers near the surface. Larger anions reduce the overall lateral ion packing density near the surface, which reduces the net charge per unit area and explains the smaller observed fields. The insight from this work is important for developing a fundamental picture of concentrated electrolytes near interfaces and can help with designing ILs to create tailored electric fields near an electrode.

Biomolecular QM/MM Simulations: What Are Some of the "Burning Issues"?

QM/MM simulations have become an indispensable tool in many chemical and biochemical investigations. Considering the tremendous degree of success, including recognition by a 2013 Nobel Prize in Chemistry, are there still "burning challenges" in QM/MM methods, especially for biomolecular systems? In this short Perspective, we discuss several issues that we believe greatly impact the robustness and quantitative applicability of QM/MM simulations to many, if not all, biomolecules. We highlight these issues with observations and relevant advances from recent studies in our group and others in the field. Despite such limited scope, we hope the discussions are of general interest and will stimulate additional developments that help push the field forward in meaningful directions.

Rigidity of melting DNA.

The temperature dependence of DNA flexibility is studied in the presence of stretching and unzipping forces. Two classes of models are considered. In one case the origin of elasticity is entropic due to the polymeric correlations, and in the other the double-stranded DNA is taken to have an intrinsic rigidity for bending. In both cases single strands are completely flexible. The change in the elastic constant for the flexible case due to thermally generated bubbles is obtained exactly. For the case of intrinsic rigidity, the elastic constant is found to be proportional to the square root of the bubble number fluctuation.

Efimov-like phase of a three-stranded DNA and the renormalization-group limit cycle.

A three-stranded DNA with short range base pairings only is known to exhibit a classical analog of the quantum Efimov effect, viz., a three-chain bound state at the two-chain melting point where no two are bound. By using a nonperturbative renormalization-group method for a rigid duplex DNA and a flexible third strand, with base pairings and strand exchange, we show that the Efimov-DNA is associated with a limit cycle type behavior of the flow of an effective three-chain interaction. The analysis also shows that thermally generated bubbles play an essential role in producing the effect. A toy model for the flow equations shows the limit cycle in an extended three-dimensional parameter space of the two-chain coupling and a complex three-chain interaction.

A study of the prevalence of osteoporosis and hypovitaminosis D in patients with primary knee osteoarthritis.

INTRODUCTION: Osteoarthritis and Osteoporosis are highly prevalent disease, so is hypovitaminosis D. We tried to find out prevalence of osteoporosis and hypovitaminosis D in patients suffering from primary knee Osteoarthritis. We also compared the prevalence of oseotoporosis between general population and patients of primary osteoarthritis. METHODS: Patients suffering from primary knee OA were taken from Rheumatology OPD of Medical College Hospital and SSKM Hospital Kolkata, India. For each patient age and sex matched friend or relative of same locality was taken in the study as controls. Hospital staffs that come from different part of state was taken in the study as controls. The control population was the representative of general population. RESULTS: Total number of participants in this study was 206. Out of which there were 98 cases and 108 controls. BMD status correlates significantly with Primary OA. Serum Vitamin D3 status correlates significantly with Osteoarthritis. Age of the patients correlated significantly with both BMD Status and Knee OA but not with the vVitamin D level. There were significant correlation between the Serum Vitamin D3 status and BMD of the subjects. CONCLUSION: Osteoporosis is prevalent both in general population and patients suffering from Knee Osteoarthritis and may increase the disability. The matter is complicated by the fact hypovitaminosis D is also prevalent in the population and positively correlated with both Osteoporosis and osteoarthrosis, though we cannot comment on further pathogenesis because of cross sectional design of the study.

Renormalization group limit cycle for three-stranded DNA.

We show that there exists an Efimov-like three strand DNA bound state at the duplex melting point and it is described by a renormalization group limit cycle. A nonperturbative renormalization group is used to obtain this result in a model involving short range pairing only. Our results suggest that Efimov physics can be tested in polymeric systems.