Membrane Fluctuation Model for Understanding the Effect of Receptor Nanoclustering on the Activation of Natural Killer Cells through Biomechanical Feedback.

We investigated the role of ligand clustering and density in the activation of natural killer (NK) cells. To that end, we designed reductionist arrays of nanopatterned ligands arranged with different cluster geometries and densities and probed their effects on NK cell activation. We used these arrays as an artificial microenvironment for the stimulation of NK cells and studied the effect of the array geometry on the NK cell immune response. We found that ligand density significantly regulated NK cell activation while ligand clustering had an impact only at a specific density threshold. We also rationalized these findings by introducing a theoretical membrane fluctuation model that considers biomechanical feedback between ligand-receptor bonds and the cell membrane. These findings provide important insight into NK cell mechanobiology, which is fundamentally important and essential for designing immunotherapeutic strategies targeting cancer.

The Stokes-Einstein-Sutherland Equation at the Nanoscale Revisited.

The Stokes-Einstein-Sutherland (SES) equation is at the foundation of statistical physics, relating a particle's diffusion coefficient and size with the fluid viscosity, temperature, and the boundary condition for the particle-solvent interface. It is assumed that it relies on the separation of scales between the particle and the solvent, hence it is expected to break down for diffusive transport on the molecular scale. This assumption is however challenged by a number of experimental studies showing a remarkably small, if any, violation, while simulations systematically report the opposite. To understand these discrepancies, analytical ultracentrifugation experiments are combined with molecular simulations, both performed at unprecedented accuracies, to study the transport of buckminsterfullerene C60 in toluene at infinite dilution. This system is demonstrated to clearly violate the conditions of slow momentum relaxation. Yet, through a linear response to a constant force, the SES equation can be recovered in the long time limit with no more than 4% uncertainty both in experiments and in simulations. This nonetheless requires partial slip on the particle interface, extracted consistently from all the data. These results, thus, resolve a long-standing discussion on the validity and limits of the SES equation at the molecular scale.

Assessment of Hydrophilicity/Hydrophobicity in Mesoporous Silica by Combining Adsorption, Liquid Intrusion, and Solid-State NMR Spectroscopy.

We have developed a comprehensive strategy for quantitatively assessing the hydrophilicity/hydrophobicity of nanoporous materials by combining advanced adsorption studies, novel liquid intrusion techniques, and solid-state NMR spectroscopy. For this, we have chosen a well-defined system of model materials, i.e., the highly ordered mesoporous silica molecular sieve SBA-15 in its pristine state and functionalized with different amounts of trimethylsilyl (TMS) groups, allowing one to accurately tailor the surface chemistry while maintaining the well-defined pore structure. For an absolute quantification of the trimethylsilyl group density, quantitative 1H solid-state NMR spectroscopy under magic angle spinning was employed. A full textural characterization of the materials was obtained by high-resolution argon 87 K adsorption, coupled with the application of dedicated methods based on nonlocal-density functional theory (NLDFT). Based on the known texture of the model materials, we developed a novel methodology allowing one to determine the effective contact angle of water adsorbed on the pore surfaces from complete wetting to nonwetting, constituting a powerful parameter for the characterization of the surface chemistry inside porous materials. The surface chemistry was found to vary from hydrophilic to hydrophobic as the TMS functionalization content was increased. For wetting and partially wetting surfaces, pore condensation of water is observed at pressures P smaller than the bulk saturation pressure p0 (i.e., at p/p0 < 1) and the effective contact angle of water on the pore walls could be derived from the water sorption isotherms. However, for nonwetting surfaces, pore condensation occurs at pressures above the saturation pressure (i.e., at p/p0 > 1). In this case, we investigated the pore filling of water (i.e., the vapor-liquid phase transition) by the application of a novel, liquid water intrusion/extrusion methodology, allowing one to derive the effective contact angle of water on the pore walls even in the case of nonwetting. Complementary molecular simulations provide density profiles of water on pristine and TMS-grafted silica surfaces (mimicking the tailored, functionalized experimental silica surfaces), which allow for a molecular view on the water adsorbate structure. Summarizing, we present a comprehensive and reliable methodology for quantitatively assessing the hydrophilicity/hydrophobicity of siliceous nanoporous materials, which has the potential to optimize applications in heterogeneous catalysis and separation (e.g., chromatography).

From Water Solutions to Ionic Liquids with Solid State Nanopores as a Perspective to Study Transport and Translocation Phenomena.

Solid state nanopores are single-molecular devices governed by nanoscale physics with a broad potential for technological applications. However, the control of translocation speed in these systems is still limited. Ionic liquids are molten salts which are commonly used as alternate solvents enabling the regulation of the chemical and physical interactions on solid-liquid interfaces. While their combination can be challenging to the understanding of nanoscopic processes, there has been limited attempts on bringing these two together. While summarizing the state of the art and open questions in these fields, several major advances are presented with a perspective on the next steps in the investigations of ionic-liquid filled nanopores, both from a theoretical and experimental standpoint. By analogy to aqueous solutions, it is argued that ionic liquids and nanopores can be combined to provide new nanofluidic functionalities, as well as to help resolve some of the pertinent problems in understanding transport phenomena in confined ionic liquids and providing better control of the speed of translocating analytes.

Quantitative Structural Organization of Bulk Apical Membrane Traffic in Pollen Tubes.

Pollen tube tip growth depends on balancing secretion of cell wall material with endocytic recycling of excess material incorporated into the plasma membrane (PM). The classical model of tip growth, which predicts bulk secretion, occurs apically, and is compensated by subapical endocytosis, has been challenged in recent years. Many signaling proteins and lipids with important functions in the regulation of membrane traffic underlying tip growth associate with distinct regions of the pollen tube PM, and understanding the mechanisms responsible for the targeting of these regulatory factors to specific PM domains requires quantitative information concerning the sites of bulk secretion and endocytosis. Here, we quantitatively characterized the spatial organization of membrane traffic during tip growth by analyzing steady-state distributions and dynamics of FM4-64-labeled lipids and YFP-tagged transmembrane (TM) proteins in tobacco (Nicotiana tabacum) pollen tubes growing normally or treated with Brefeldin A to block secretion. We established that (1) secretion delivers TM proteins and recycled membrane lipids to the same apical PM domain, and (2) FM4-64-labeled lipids, but not the analyzed TM proteins, undergo endocytic recycling within a clearly defined subapical region. We mathematically modeled the steady-state PM distributions of all analyzed markers to better understand differences between them and to support the experimental data. Finally, we mapped subapical F-actin fringe and trans-Golgi network positioning relative to sites of bulk secretion and endocytosis to further characterize functions of these structures in apical membrane traffic. Our results support and further define the classical model of apical membrane traffic at the tip of elongating pollen tubes.

Biomechanics as driver of aggregation of tethers in adherent membranes.

Cell adhesion is an important cellular process and is mediated by adhesion proteins residing on the cell membrane. Sometimes, two types of linker proteins are involved in adhesion, and they can segregate to form domains through a poorly understood size-exclusion process. We present an experimental and theoretical study of adhesion via linkers of two different sizes, realised in a mimetic model-system, based on giant unilamellar vesicles interacting with supported lipid bilayers. Here, adhesion is mediated by DNA linkers with two different lengths, but with the same binding enthalpy. We study the organisation of these linkers into domains as a function of relative fraction of long and short DNA constructs. Experimentally, we find that, irrespective of the composition, the adhesion domains are uniform with coexisting DNA bridge types, despite their relative difference in length of 9 nm. However, simulations suggest formation of nanodomains of the minority fraction at short length scales, which is below the optical resolution of the microscope. The nano-aggregation is more significant for long bridges, which are also more stable.

Regimes of motion of magnetocapillary swimmers.

The dynamics of a triangular magnetocapillary swimmer is studied using the lattice Boltzmann method. We extend on our previous work, which deals with the self-assembly and a specific type of the swimmer motion characterized by the swimmer's maximum velocity centred around the particle's inverse viscous time. Here, we identify additional regimes of motion. First, modifying the ratio of surface tension and magnetic forces allows to study the swimmer propagation in the regime of significantly lower frequencies mainly defined by the strength of the magnetocapillary potential. Second, introducing a constant magnetic contribution in each of the particles in addition to their magnetic moment induced by external fields leads to another regime characterized by strong in-plane swimmer reorientations that resemble experimental observations.

Molecular Biomechanics Controls Protein Mixing and Segregation in Adherent Membranes.

Cells interact with their environment by forming complex structures involving a multitude of proteins within assemblies in the plasma membrane. Despite the omnipresence of these assemblies, a number of questions about the correlations between the organisation of domains and the biomechanical properties of the involved proteins, namely their length, flexibility and affinity, as well as about the coupling to the elastic, fluctuating membrane, remain open. Here we address these issues by developing an effective Kinetic Monte Carlo simulation to model membrane adhesion. We apply this model to a typical experiment in which a cell binds to a functionalized solid supported bilayer and use two ligand-receptor pairs to study these couplings. We find that differences in affinity and length of proteins forming adhesive contacts result in several characteristic features in the calculated phase diagrams. One such feature is mixed states occurring even with proteins with length differences of 10 nm. Another feature are stable nanodomains with segregated proteins appearing on time scales of cell experiments, and for biologically relevant parameters. Furthermore, we show that macroscopic ring-like patterns can spontaneously form as a consequence of emergent protein fluxes. The capacity to form domains is captured by an order parameter that is founded on the virial coefficients for the membrane mediated interactions between bonds, which allow us to collapse all the data. These findings show that taking into account the role of the membrane allows us to recover a number of experimentally observed patterns. This is an important perspective in the context of explicit biological systems, which can now be studied in significant detail.

Statistical Mechanics of an Elastically Pinned Membrane: Equilibrium Dynamics and Power Spectrum.

In biological settings, membranes typically interact locally with other membranes: the extracellular matrix in the exterior or internal cellular structures such as the cytoskeleton, locally pinning the membrane. Characterizing the dynamical properties of such interactions presents a difficult task. Significant progress has been achieved through simulations and experiments, yet analytical progress in modeling pinned membranes has been impeded by the complexity of governing equations. Here, we circumvent these difficulties by calculating analytically the time-dependent Green's function of the operator governing the dynamics of an elastically pinned membrane in a hydrodynamic surrounding and subject to external forces. This enables us to calculate the equilibrium power spectral density for an overdamped membrane pinned by an elastic, permanently attached spring subject to thermal excitations. By considering the effects of the finite experimental resolution on the measured spectra, we show that the elasticity of the pinning can be extracted from the experimentally measured spectrum. Membrane fluctuations can thus be used as a tool to probe mechanical properties of the underlying structures. Such a tool may be particularly relevant in the context of cell mechanics, in which the elasticity of the membrane's attachment to the cytoskeleton could be measured.

Statistical Mechanics of an Elastically Pinned Membrane: Static Profile and Correlations.

The relation between thermal fluctuations and the mechanical response of a free membrane has been explored in great detail, both theoretically and experimentally. However, understanding this relationship for membranes locally pinned by proteins is significantly more challenging. Given that the coupling of the membrane to the cell cytoskeleton, to the extracellular matrix, and to other internal structures is crucial for the regulation of a number of cellular processes, understanding the role of the pinning is of great interest. In this manuscript, we consider a single protein (elastic spring of a finite rest length) pinning a membrane modeled in the Monge gauge. First, we determine the Green's function for the system and complement this approach by the calculation of the mode-coupling coefficients for the plane wave expansion and the orthonormal fluctuation modes, in turn building a set of tools for numerical and analytic studies of a pinned membrane. Furthermore, we explore static correlations of the free and the pinned membrane, as well as the membrane shape, showing that all three are mutually interdependent and have an identical long-range behavior characterized by the correlation length. Interestingly, the latter displays a nonmonotonic behavior as a function of membrane tension. Importantly, exploiting these relations allows for the experimental determination of the elastic parameters of the pinning. Last but not least, we calculate the interaction potential between two pinning sites and show that even in the absence of the membrane deformation, the pinnings will be subject to an attractive force because of changes in membrane fluctuations.

The Influence of Chemical Change on Protein Dynamics: A Case Study with Pyruvate Formate-Lyase.

Pyruvate formate-lyase (PFL) catalyzes the reversible conversion of pyruvate and coenzyme A (CoA) into formate and acetyl-CoA in two half-reactions. For the second half-reaction to take place, the S-H group of CoA must enter the active site of the enzyme to retrieve a protein-bound acetyl group. However, CoA is bound at the protein surface, whereas the active site is buried in the protein interior, some 20-30 Šaway. The PFL system was therefore subjected to a series of extensive molecular dynamics simulations (in the µs range) and a host of advanced analysis procedures. Models representing PFL before and after the first half-reaction were used to examine the possible effect of enzyme acetylation. All simulated structures were found to be relatively stable compared to the initial crystal structure. Although the adenine portion of CoA remained predominantly bound at the protein surface, the binding of the S-H group was significantly more labile. A potential entry channel for CoA, which would allow the S-H group to reach the active site, was identified and characterized. The channel was found to be associated with accentuated fluctuations and a higher probability of being in an open state in acetylated systems. This result suggests that the acetylation of the enzyme assumes a prominent functional role, whereby the formation of the acyl intermediate serves to initiate a subtle signaling cascade that influences the protein dynamics and facilitates the entry of the second substrate.

Correction: Capillary assemblies in a rotating magnetic field.

Correction for 'Capillary assemblies in a rotating magnetic field' by Galien Grosjean et al., Soft Matter, 2019, DOI: .

Geometric effects in random assemblies of ellipses.

Assemblies of anisotropic particles commonly appear in studies of active many-body systems. However, in two dimensions, the geometric ramifications of the finite density of such objects are not entirely understood. To fully characterize these effects, we perform an in-depth study of random assemblies generated by a slow compression of frictionless elliptical particles. The obtained configurations are then analysed using the Set Voronoi tessellation, which takes the particle shape into account. Not only do we analyse most scalar and vectorial morphological measures, which are commonly discussed in the literature or which have recently been addressed in experiments, but we also systematically explore the correlations between them. While in a limited range of parameters similarities with findings in 3D assemblies could be identified, important differences are found when a broad range of aspect ratios and packing fractions are considered. The data discussed in this study should thus provide a unique reference set such that geometric effects and differences from random assemblies could be clearly identified in more complex systems, including ones with soft and active particles that are typically found in biological systems.

Capillary assemblies in a rotating magnetic field.

Small objects floating on a fluid have a tendency to aggregate due to capillary forces. This effect has been used, with the help of a magnetic induction field, to assemble submillimeter metallic spheres into a variety of structures, whose shape and size can be tuned. Under time-varying fields, these assemblies can propel themselves due to a breaking of time reversal symmetry in their adopted shapes. In this article, we study the influence of an in-plane rotation of the magnetic field on these structures. Various rotational modes have been observed with different underlying mechanisms. The magnetic properties of the particles cause them to rotate individually. Dipole-dipole interactions in the assembly can cause the whole structure to align with the field. Finally, non-reciprocal deformations can power the rotation of the assembly. Symmetry plays an important role in the dynamics, as well as the frequency and amplitude of the applied field. Understanding the interplay of these effects is essential, both to explain previous observations and to develop new functions for these assemblies.

Exploring Reactive Conformations of Coenzyme A during Binding and Unbinding to Pyruvate Formate-Lyase.

Pyruvate formate-lyase (PFL) is a glycyl radical enzyme that converts pyruvate and coenzyme A (CoA) into formate and acetyl-CoA in two half-reactions. Recently, we showed that the acetylation of the PFL active site in the first half-reaction induces subtle conformational changes, leading to the opening of a potential channel for CoA entry. Entry of CoA into the active site is crucial for the second half-reaction, involving the acetyl transfer to CoA, and the completion of the catalytic cycle. Using steered molecular dynamics (SMD) simulations, performed on acetylated and nonacetylated monomeric PFL model systems, we first of all investigate the possible entry/exit pathways of CoA with respect to the active site through the previously identified channel. We then perform umbrella sampling simulations on multiple snapshots from SMD trajectories as well as unrestrained molecular dynamics simulations starting from the final structures obtained from entry SMD, with a view to identifying possible bound states of CoA in the near vicinity of the active site. Detailed study of the unrestrained dissociation processes reveals the presence of stable and reactive bound states of CoA close to the active site, one of which is in an ideal position for triggering the second half-reaction. Examination of the spatial distributions associated with the reactive bound states allows us to discuss the free energy barriers. Umbrella sampling, performed on snapshots from unrestrained dynamics confirms the above findings. The significance of the results for the catalysis are discussed for both acetylated and nonacetylated systems.

Optimal motion of triangular magnetocapillary swimmers.

A system of ferromagnetic particles trapped at a liquid-liquid interface and subjected to a set of magnetic fields (magnetocapillary swimmers) is studied numerically using a hybrid method combining the pseudopotential lattice Boltzmann method and the discrete element method. After investigating the equilibrium properties of a single, two, and three particles at the interface, we demonstrate a controlled motion of the swimmer formed by three particles. It shows a sharp dependence of the average center-of-mass speed on the frequency of the time-dependent external magnetic field. Inspired by experiments on magnetocapillary microswimmers, we interpret the obtained maxima of the swimmer speed by the optimal frequency centered around the characteristic relaxation time of a spherical particle. It is also shown that the frequency corresponding to the maximum speed grows and the maximum average speed decreases with increasing interparticle distances at moderate swimmer sizes. The findings of our lattice Boltzmann simulations are supported by bead-spring model calculations.

Mechanism of the Water-Gas Shift Reaction Catalyzed by Efficient Ruthenium-Based Catalysts: A Computational and Experimental Study.

Supported ionic liquid phase (SILP) catalysis enables a highly efficient, Ru-based, homogeneously catalyzed water-gas shift reaction (WGSR) between 100 °C and 150 °C. The active Ru-complexes have been found to exist in imidazolium chloride melts under operating conditions in a dynamic equilibrium, which is dominated by the [Ru(CO)3 Cl3 ]- complex. Herein we present state-of-the-art theoretical calculations to elucidate the reaction mechanism in more detail. We show that the mechanism includes the intermediate formation and degradation of hydrogen chloride, which effectively reduces the high barrier for the formation of the requisite dihydrogen complex. The hypothesis that the rate-limiting step involves water is supported by using D2 O in continuous catalytic WGSR experiments. The resulting mechanism constitutes a highly competitive alternative to earlier reported generic routes involving nucleophilic addition of hydroxide in the gas phase and in solution.

Effect of body deformability on microswimming.

In this work we consider the following question: given a mechanical microswimming mechanism, does increased deformability of the swimmer body hinder or promote the motility of the swimmer? To answer this we run immersed-boundary-lattice-Boltzmann simulations of a microswimmer composed of deformable beads connected with springs. We find that the same deformations in the beads can result in different effects on the swimming velocity, namely an enhancement or a reduction, depending on the other parameters. To understand this we determine analytically the velocity of the swimmer, starting from the forces driving the motion and assuming that the deformations in the beads are known as functions of time and are much smaller than the beads themselves. We find that to the lowest order, only the driving frequency mode of the surface deformations contributes to the swimming velocity, and comparison to the simulations shows that both the velocity-promoting and velocity-hindering effects of bead deformability are reproduced correctly by the theory in the limit of small bead deformations. For the case of active deformations we show that there are critical values of the spring constant - which for a general swimmer corresponds to its main elastic degree of freedom - which decide whether the body deformability is beneficial for motion or not.

Crowding of receptors induces ring-like adhesions in model membranes.

The dynamics of formation of macromolecular structures in adherent membranes is a key to a number of cellular processes. However, the interplay between protein reaction kinetics, diffusion and the morphology of the growing domains, governed by membrane mediated interactions, is still poorly understood. Here we show, experimentally and in simulations, that a rich phase diagram emerges from the competition between binding, cooperativity, molecular crowding and membrane spreading. In the cellular context, the spontaneously-occurring organization of adhesion domains in ring-like morphologies is particularly interesting. These are stabilized by the crowding of bulky proteins, and the membrane-transmitted correlations between bonds. Depending on the density of the receptors, this phase may be circumvented, and instead, the adhesions may grow homogeneously in the contact zone between two membranes. If the development of adhesion occurs simultaneously with membrane spreading, much higher accumulation of binders can be achieved depending on the velocity of spreading. The mechanisms identified here, in the context of our mimetic model, may shed light on the structuring of adhesions in the contact zones between two living cells. This article is part of a Special Issue entitled: Mechanobiology.

Nanometric thermal fluctuations of weakly confined biomembranes measured with microsecond time-resolution.

We probe the bending fluctuations of bio-membranes using highly deflated giant unilamellar vesicles (GUVs) bound to a substrate by a weak potential arising from generic interactions. The substrate is either homogeneous, with GUVs bound only by the weak potential, or is chemically functionalized with a micro-pattern of very strong specific binders. In both cases, the weakly adhered membrane is seen to be confined at a well-defined distance above the surface while it continues to fluctuate strongly. We quantify the fluctuations of the weakly confined membrane at the substrate proximal surface as well as of the free membrane at the distal surface of the same GUV. This strategy enables us to probe in detail the damping of fluctuations in the presence of the substrate, and to independently measure the membrane tension and the strength of the generic interaction potential. Measurements were done using two complementary techniques - dynamic optical displacement spectroscopy (DODS, resolution: 20 nm, 10 µs), and dual wavelength reflection interference contrast microscopy (DW-RICM, resolution: 4 nm, 50 ms). After accounting for the spatio-temporal resolution of the techniques, an excellent agreement between the two measurements was obtained. For both weakly confined systems we explore in detail the link between fluctuations on the one hand and membrane tension and the interaction potential on the other hand.