Osmolyte-induced conformational stabilization of a hydrophobic polymer.

Elucidating the mechanistic role of osmolytes on conformations of hydrophobic prototypical macromolecules in principle is the stepping stone towards understanding the effect of osmolytes on proteins. Motivated by this, we use equilibrium simulations and umbrella sampling techniques to dissect the underlying mechanism of osmolyte-induced conformational stability of a hydrophobic polymer. Our results unveil a remarkable osmolyte-dependent conformational stabilization of the polymer. In an aqueous solution of 4 M choline chloride (ChCl), the polymer has an even more compact structure than in water. On the other hand, an aqueous solution of 8 M urea stabilizes the extended state of the polymer. Interestingly, the polymer adopts an intermediate hairpin conformation in a mixed osmolyte solution of 4 M ChCl and 8 M urea in water due to the interplay of ChCl and urea. Our simulations identify the relative accumulation of water and the hydrophilic part of choline or preferential binding of urea near the collapsed and the extended states, respectively. Analyses split out the enthalpic and entropic contributions to the overall free energy. This decides the stabilization of the preferred conformation in the chosen osmolyte solution. Our simulations show that in an aqueous solution of ChCl, the hairpin state is stabilized by entropy gain. In contrast, the enthalpic contribution stabilizes the hairpin state in mixed environments. However, a collapsed state is energetically not favored in the presence of urea. In brief, via employing an in silico approach, the current findings indicate the importance of osmolytes in stabilizing the conformational states of hydrophobic polymers.

Compound giant unilamellar vesicles as a bio-mimetic model for electrohydrodynamics of a nucleate cell.

The understanding obtained by studies on the electrohydrodynamics (EHD) of single giant unilamellar vesicles (sGUVs) has contributed significantly towards a better comprehension of the response of biological cells to electric fields. This has subsequently helped in developing technologies such as cell dielectrophoresis and cell electroporation. For nucleate eukaryotic cells though, a vesicle-in-vesicle compound giant unilamellar vesicle (cGUV) is a more appropriate bio-mimic than a sGUV. In this work, we present an improvised method for the formation of cGUVs, wherein the electrical conductivities of the inner, annular and outer regions of the cGUVs can be modified. A comprehensive experimental study is presented on the EHD of these cGUVs under weak AC fields over a wide range of frequencies, and an encouraging agreement is observed between the experiments and earlier published theoretical studies on concentric cGUVs. The spherical, prolate or oblate spheroidal deformations of a cGUV under AC electric fields depend upon the membrane electromechanical properties as well as the magnitude and direction of the electric traction at the membrane produced by the Maxwell stress that varies with the relative timescales associated with the frequency of the applied AC electric field and that of the membrane charging time and the Maxwell-Wagner relaxation time. This work establishes cGUVs as appropriate bio-mimics for conducting EHD studies relevant to eukaryotic cells.

Correction: A passive star polymer in a dense active bath: insights from computer simulations.

Correction for 'A passive star polymer in a dense active bath: insights from computer simulations' by Ramanand Singh Yadav et al., Soft Matter, 2024, 20, 3910-3922, https://doi.org/10.1039/D4SM00144C.

A passive star polymer in a dense active bath: insights from computer simulations.

Using computer simulations in two dimensions (2D), we explore the structure and dynamics of a star polymer with three arms made of passive monomers immersed in a bath of active Brownian particles (ABPs). We analyze the conformational and dynamical changes of the polymer as a function of activity and packing fraction. We also study the process of motility induced phase separation (MIPS) in the presence of a star polymer, which acts as a mobile nucleation center. The presence of the polymer increases the growth rate of the clusters in comparison to a bath without the polymer. In particular, for low packing fraction, both nucleation and cluster growth are affected by the inclusion of the star polymer. Clusters grow in the vicinity of the star polymer, resulting in the star polymer experiencing a caged motion similar to a tagged ABP in the dense phase. Due to the topological constraints of the star polymers and clustering nearby, the conformational changes of the star polymer lead to interesting observations. Inter alia, we observe the shrinking of the arm with increasing activity along with a short-lived hairpin structure of one arm formed. We also see the transient pairing of two arms of the star polymer, while the third is largely separated at high activity. We hope our findings will help in understanding the behavior of active-passive mixtures, including biopolymers of complex topology in dense active suspensions.

Decoding the dynamics of BCL9 triazole stapled peptide.

BCL9 is a key protein in Wnt signaling pathway. It acts as a transcriptional co-activator to ß-catenin, and dysregulation in this pathway leads to tumor growth. Inhibiting such a protein-protein interaction is considered as a therapeutic challenge. The interaction between ß-catenin and BCL9 is facilitated by a 23-residue helical domain from BCL9 and a hydrophobic groove of ß-catenin. To prevent this interaction, a peptide that mimics the alpha-helical domain of BCL9 can be designed. Stapling is considered a successful strategy in the pursuit of designing such peptides in which amino acids side are stitched together using chemical moieties. Among the various types of cross-linkers, triazole is the most rapid and effective one synthesized via click reaction. However, the underlying interactions behind maintaining the secondary structure of stapled peptides remain less explored. In the current work, we employed the molecular dynamics simulation to study the conformational behavior of the experimentally synthesized single and double triazole stapled BCL9 peptide. Upon the addition of a triazole staple, there is a significant reduction in the conformational space of BCL9. The helical character of the stapled peptide increases with an increase in separation between the triazole cross-linkers. Also, we encompassed the Replica Exchange with Solute Tempering (REST2) simulation to validate the high-temperature response of the stapled peptide. From REST2, the PCA and t-SNE show the reduction in distinct cluster formation on the addition of triazole staple. Our study infers further development of these triazole-stapled BCL9 peptides into effective inhibitors to target the interaction between ß-catenin and BCL9.

Escape dynamics of a self-propelled nanorod from circular confinements with narrow openings.

We perform computer simulations to explore the escape dynamics of a self-propelled (active) nanorod from circular confinements with narrow opening(s). Our results clearly demonstrate how the persistent and directed motion of the nanorod helps it to escape. Such escape events are absent if the nanorod is passive. To quantify the escape dynamics, we compute the radial probability density function (RPDF) and mean first escape time (MFET) and show how the activity is responsible for the bimodality of RPDF, which is clearly absent if the nanorod is passive. Broadening of displacement distributions with activity has also been observed. The computed mean first escape time decreases with activity. In contrast, the fluctuations of the first escape times vary in a non-monotonic way. This results in high values of the coefficient of variation and indicates the presence of multiple timescales in first escape time distributions and multimodality in uniformity index distributions. We hope our study will help in differentiating activity-driven escape dynamics from purely thermal passive diffusion in confinement.

Active dynamics of linear chains and rings in porous media.

To understand the dynamical and conformational properties of deformable active agents in porous media, we computationally investigate the dynamics of linear chains and rings made of active Brownian monomers. In porous media, flexible linear chains and rings always migrate smoothly and undergo activity-induced swelling. However, semiflexible linear chains though navigate smoothly, shrink at lower activities, followed by swelling at higher activities, while semiflexible rings exhibit a contrasting behavior. Semiflexible rings shrink, get trapped at lower activities, and escape at higher activities. This demonstrates how activity and topology interplay and control the structure and dynamics of linear chains and rings in porous media. We envision that our study will shed light on understanding the mode of transport of shape-changing active agents in porous media.

Structure and dynamics of an active polymer chain inside a nanochannel grafted with polymers.

We use computer simulations to investigate the complex dynamics of a polymer, made of active Brownian particles, inside a channel grafted internally with passive polymer chains. Our simulations reveal that this probe-polymer, if passive, exhibits a compact structure when its interaction is repulsive with the grafted chains as it tends to stay within the hollow space created along the axis of the channel. On increasing the attractive interaction, the passive probe-polymer is pulled towards the grafted polymeric region and adopts an extended structure. By contrast, switching on the activity helps the probe-polymer to escape from the local traps caused by the sticky grafted chains. The interplay between the activity of the probe-polymer and its sticky interaction with the grafted chains results in shrinking, followed by swelling as the activity is increased. To elucidate the dynamics we compute the mean square displacement (MSD) of the center of mass of the probe-polymer, which increases monotonically with activity and displays superdiffusive behavior at an intermediate time and enhanced diffusion at a long time period. In addition, compared with the attractive interaction, the active probe-polymer shows faster dynamics when the interaction is repulsive to the grafted polymers. We believe that our current study will provide insights into the structural changes and dynamics of active polymers in heterogeneous media and will be useful in designing polymer-based drug delivery vehicles.

The multiple links between actin and mitochondria.

Actin plays many well-known roles in cells, and understanding any specific role is often confounded by the overlap of multiple actin-based structures in space and time. Here, we review our rapidly expanding understanding of actin in mitochondrial biology, where actin plays multiple distinct roles, exemplifying the versatility of actin and its functions in cell biology. One well-studied role of actin in mitochondrial biology is its role in mitochondrial fission, where actin polymerization from the endoplasmic reticulum through the formin INF2 has been shown to stimulate two distinct steps. However, roles for actin during other types of mitochondrial fission, dependent on the Arp2/3 complex, have also been described. In addition, actin performs functions independent of mitochondrial fission. During mitochondrial dysfunction, two distinct phases of Arp2/3 complex-mediated actin polymerization can be triggered. First, within 5 min of dysfunction, rapid actin assembly around mitochondria serves to suppress mitochondrial shape changes and to stimulate glycolysis. At a later time point, at more than 1 h post-dysfunction, a second round of actin polymerization prepares mitochondria for mitophagy. Finally, actin can both stimulate and inhibit mitochondrial motility depending on the context. These motility effects can either be through the polymerization of actin itself or through myosin-based processes, with myosin 19 being an important mitochondrially attached myosin. Overall, distinct actin structures assemble in response to diverse stimuli to affect specific changes to mitochondria.

Acute ACAT1/SOAT1 Blockade Increases MAM Cholesterol and Strengthens ER-Mitochondria Connectivity.

Cholesterol is a key component of all mammalian cell membranes. Disruptions in cholesterol metabolism have been observed in the context of various diseases, including neurodegenerative disorders such as Alzheimer's disease (AD). The genetic and pharmacological blockade of acyl-CoA:cholesterol acyltransferase 1/sterol O-acyltransferase 1 (ACAT1/SOAT1), a cholesterol storage enzyme found on the endoplasmic reticulum (ER) and enriched at the mitochondria-associated ER membrane (MAM), has been shown to reduce amyloid pathology and rescue cognitive deficits in mouse models of AD. Additionally, blocking ACAT1/SOAT1 activity stimulates autophagy and lysosomal biogenesis; however, the exact molecular connection between the ACAT1/SOAT1 blockade and these observed benefits remain unknown. Here, using biochemical fractionation techniques, we observe cholesterol accumulation at the MAM which leads to ACAT1/SOAT1 enrichment in this domain. MAM proteomics data suggests that ACAT1/SOAT1 inhibition strengthens the ER-mitochondria connection. Confocal and electron microscopy confirms that ACAT1/SOAT1 inhibition increases the number of ER-mitochondria contact sites and strengthens this connection by shortening the distance between these two organelles. This work demonstrates how directly manipulating local cholesterol levels at the MAM can alter inter-organellar contact sites and suggests that cholesterol buildup at the MAM is the impetus behind the therapeutic benefits of ACAT1/SOAT1 inhibition.

A polymer chain with dipolar active forces in connection to spatial organization of chromatin.

A living cell is an active environment where the organization and dynamics of chromatin are affected by different forms of activity. Optical experiments report that loci show subdiffusive dynamics and the chromatin fiber is seen to be coherent over micrometer-scale regions. Using a bead-spring polymer chain with dipolar active forces, we study how the subdiffusive motion of the loci generate large-scale coherent motion of the chromatin. We show that in the presence of extensile (contractile) activity, the dynamics of the loci grows faster (slower) and the spatial correlation length increases (decreases) compared to the case with no dipolar forces. Hence, both the dipolar active forces modify the elasticity of the chain. Interestingly in our model, the dynamics and organization of such dipolar active chains largely differ from the passive chain with renormalized elasticity.

Dynamics of a spherical self-propelled tracer in a polymeric medium: interplay of self-propulsion, stickiness, and crowding.

We employ computer simulations to study the dynamics of a self-propelled spherical tracer particle in a viscoelastic medium, made of a long polymer chain. Here, the interplay between viscoelasticity, stickiness, and activity (self-propulsion) brings additional complexity to the tracer dynamics. Our simulations show that on increasing the stickiness of the tracer particle to the polymer beads, the dynamics of the tracer particle slows down as it gets stuck to the polymer chain and moves along with it. But with increasing self-propulsion velocity, the dynamics gets enhanced. In the case of increasing stickiness as well as activity, the non-Gaussian parameter (NGP) exhibits non-monotonic behavior, which also shows up in the re-scaled self part of the van-Hove function. Non-Gaussianity results owing to the enhanced binding events and the sticky motion of the tracer along with the chain with increasing stickiness. On the other hand, with increasing activity, initially non-Gaussianity increases as the tracer moves through the heterogeneous polymeric environment but for higher activity, the tracer escapes resulting in a negative NGP. For higher values of stickiness, the trapping time distributions of the passive tracer particle broaden and have long tails. On the other hand, for a given stickiness with increasing self-propulsion force, the trapping time distributions become narrower and have short tails. We believe that our current simulation study will be helpful in elucidating the complex motion of activity-driven probes in viscoelastic media.

Dynamics of self-propelled tracer particles inside a polymer network.

The transport of tracer particles through mesh-like environments such as biological hydrogels and polymer matrices is ubiquitous in nature. These tracers can be passive, such as colloids, or active (self-propelled), for example, synthetic nanomotors or bacteria. Computer simulations in principle could be extremely useful in exploring the mechanism of the active transport of tracer particles through mesh-like environments. Therefore, we construct a polymer network on a diamond lattice and use computer simulations to investigate the dynamics of spherical self-propelled particles inside the network. Our main objective is to elucidate the effect of the self-propulsion on the tracer particle dynamics as a function of the tracer size and the stiffness of the polymer network. We compute the time-averaged mean-squared displacement (MSD) and the van-Hove correlations of the tracer. On the one hand, in the case of a bigger sticky particle, the caging caused by the network particles wins over the escape assisted by the self-propulsion. This results an intermediate-time subdiffusion. On the other hand, smaller tracers or tracers with high self-propulsion velocities can easily escape from the cages and show intermediate-time superdiffusion. The stiffer the network, the slower the dynamics of the tracer, and bigger tracers exhibit longer lived intermediate time superdiffusion, since the persistence time scales as ∼σ3, where σ is the diameter of the tracer. At the intermediate time, non-Gaussianity is more pronounced for active tracers. At the long time, the dynamics of the tracer, if passive or weakly active, becomes Gaussian and diffusive, but remains flat for tracers with high self-propulsion, accounting for their seemingly unrestricted motion inside the network.

In Silico Studies of Active Probe Dynamics in Crowded Media.

Active systems are made of agents, each of which takes energy from the environment and converts it to directed motion. Therefore, by construction, these systems function out of equilibrium and cannot be described using equilibrium statistical mechanics. Though the most studied aspect has been the collective motion of active particles, the motion at the individual particle level in crowded media is also of prime importance. Examples include the motion of bacteria in hydrogels, single cell migration as a way to search for food or escape from toxic agents, and synthetic active agents transporting through soft crowded media. This review presents an overview of our understanding of single active probe dynamics in crowded media from computer simulations. The active probe is a Janus or a dumbbell-shaped particle, and the medium is made of crowders that are either sticky or repulsive to the probe and could be frozen or mobile. The density and the topology of the crowders also play an important role. We hope our in silico studies will help to elucidate the mechanism of activity-driven transport in crowded media in general and design nanomachines for targeted delivery.

Mitochondrial dysfunction triggers actin polymerization necessary for rapid glycolytic activation.

Mitochondrial damage represents a dramatic change in cellular homeostasis. One rapid response is perimitochondrial actin polymerization, termed acute damage-induced actin (ADA). The consequences of ADA are not understood. In this study, we show evidence suggesting that ADA is linked to rapid glycolytic activation upon mitochondrial damage in multiple cells, including mouse embryonic fibroblasts and effector CD8+ T lymphocytes. ADA-inducing treatments include CCCP, antimycin, rotenone, oligomycin, and hypoxia. The Arp2/3 complex inhibitor CK666 or the mitochondrial sodium-calcium exchanger (NCLX) inhibitor CGP37157 inhibits both ADA and the glycolytic increase within 5 min, supporting ADA's role in glycolytic stimulation. Two situations causing chronic reductions in mitochondrial ATP production, mitochondrial DNA depletion and mutation to the NDUFS4 subunit of complex 1 of the electron transport chain, cause persistent perimitochondrial actin filaments similar to ADA. CK666 treatment causes rapid mitochondrial actin loss and a drop in ATP in NDUFS4 knock-out cells. We propose that ADA is necessary for rapid glycolytic activation upon mitochondrial impairment, to re-establish ATP production.

Distinct mode of membrane interaction and disintegration by diverse class of antimicrobial peptides.

The exploitation of conventional antibiotics in conjunction with the adeptness of microbes has led to the emergence of multi-drug-resistant pathogens. This has posed a severe threat to combating life-threatening infectious diseases. Antimicrobial peptides (AMP), which are considered to be the first line of defense in all living organisms, are being developed for therapeutic use. Herein, we determined the NMR solution structure of Rhesus macaque Myeloid Alpha Defensin-4 (RMAD4), a defensin AMP. Additionally, the distinct modes of membrane perturbation for two structurally dissimilar classes of AMPs was studied using biophysical methods namely, Solid-state 31P NMR, DSC and cryo-TEM. The cathelicidin - Bovine myeloid antimicrobial peptide (BMAP-28 (1-18)), which adopts a helical conformation, and the defensin RMAD4 peptide that natively folds to form ß-sheets appeared to engage differently with the bacterial membrane. The helical BMAP-28 (1-18) peptide initiates lipid segregation and membrane thinning followed by pore formation, while the ß-stranded RMAD4 peptide demonstrates fragmentation of the bilayer by the carpet or detergent-like mechanism of action. Molecular dynamics studies sufficiently corroborated these findings. The structure and mechanism of action of the AMPs studied using experimental and computational approaches are believed to help in providing a platform for the rational design of new competent and cost-effective antimicrobial peptides for therapeutic applications.

Migration of active rings in porous media.

Inspired by how the shape deformations in active organisms help them to migrate through disordered porous environments, we simulate active ring polymers in two-dimensional random porous media. Flexible and inextensible active ring polymers navigate smoothly through the disordered media. In contrast, semiflexible rings undergo transient trapping inside the pore space; the degree of trapping is inversely correlated with the increase in activity. We discover that flexible rings swell while inextensible and semiflexible rings monotonically shrink upon increasing the activity. Together, our findings identify the optimal migration of active ring polymers through porous media.

Structure and Orientation of Water and Choline Chloride Molecules around a Methane Hydrophobe: A Computer Simulation Study.

Recent studies have reported manifold industrial applications of aqueous choline chloride (ChCl) solution as an alternative to deep eutectic solvent. ChCl also serves as a protecting co-solvent for proteins by restricting urea to approach the protein surface and thereby maintaining the water structure around the protein. However, a detailed molecular-level picture of the ChCl and water, even in the absence of urea around a representative hydrophobe is largely lacking. This motivates us to probe the effect of varying wt % of ChCl on the occupancy and orientations of the constituents around a representative solute like methane using computer simulations. Accumulation of water molecules and preferential exclusion of ChCl from the surface of methane perturb the tetrahedral geometry of water around it. We find a tangential alignment of the polar part of the ChCl molecules that interact with water, whereas its hydrophobic part is preferentially facing the methane. With an increase in ChCl wt %, a disruption in the tetrahedrality is evident for water molecules accompanied by a reduction in hydrogen bonds between water pairs in the solution. In short, ChCl induces crowding and modifies the microscopic arrangement and hydrogen bonding structure of the water around the methane and beyond.

Computational design of stapled peptide inhibitor against SARS-CoV-2 receptor binding domain.

Since its first detection in 2019, the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has been the cause of millions of deaths worldwide. Despite the development and administration of different vaccines, the situation is still worrisome as the virus is constantly mutating to produce newer variants some of which are highly infectious. This raises an urgent requirement to understand the infection mechanism and thereby design therapeutic-based treatment for COVID-19. The gateway of the virus to the host cell is mediated by the binding of the receptor binding domain (RBD) of the virus spike protein to the angiotensin-converting enzyme 2 (ACE2) of the human cell. Therefore, the RBD of SARS-CoV-2 can be used as a target to design therapeutics. The α1 helix of ACE2, which forms direct contact with the RBD surface, has been used as a template in the current study to design stapled peptide therapeutics. Using computer simulation, the mechanism and thermodynamics of the binding of six stapled peptides with RBD have been estimated. Among these, the one with two lactam stapling agents has shown binding affinity, sufficient to overcome RBD-ACE2 binding. Analyses of the mechanistic detail reveal that a reorganization of amino acids at the RBD-ACE2 interface produces favorable enthalpy of binding whereas conformational restriction of the free peptide reduces the loss in entropy to result higher binding affinity. The understanding of the relation of the nature of the stapling agent with their binding affinity opens up the avenue to explore stapled peptides as therapeutic against SARS-CoV-2.