The thermally reversible phase transitions in aqueous solutions of the triblock copolymers known as Pluronic and their related textures are well-researched. However, their corresponding rheological properties are less studied. In particular, their high-temperature behavior is difficult to access with classical rheology. Here we demonstrated that Diffusing Wave Spectroscopy (DWS)-based microrheology allows us to study the phase transition and the associated viscoelastic properties of Pluronic F127 solutions for temperatures from 5 C to 80 C. From the measured intensity-autocorrelation functions we can extract effective viscosities and determine the critical micellization temperature and concentration. Moreover,the high EO/PO (arm-to-core) ratio of F127 and its polydispersity play a critical role in the high-temperature re-entrant liquid phase, due to decreasing solubility of PEO along with the dehydration of the PPO core. The microscopic viscoelastic moduli G'(ω) and G”(ω) help to determine these phase transitions and provide mechanical properties in the solid phase that are not readily accessible with standard multi-particle tracking techniques due to limited Brownian motion.

During drying, particle-laden sessile droplets will leave so-called coffee-stain rings behind. This phenomenon is well known and well understood (Deegan et al., Nature 389, 827-829 (1997). Here, we show that when particle-laden droplets confined in a slit are allowed to evaporate very slowly, they do not deposit coffee rings, but form surprisingly intricate, circular maze-like patterns. We present experiments that illustrate this pattern formation and discuss the factors that determine when such patterns can form. We are not aware of reports of natural examples of the formation of such beautiful patterns under confinement, although it seems likely that they exist.

Clay-polymer mixtures, characterised by high mechanical stiffness, are widely utilised in strengthening soft and easy-to-break materials. Here, we present microrheological results on shake-gels made of natural (montmorillonite) and synthetic (Laponite) disk-shaped clay particles in combination with poly(ethyleneoxide). These clay-polymer suspensions represent shear-thickening fluids that display a large increase in viscosity upon large-amplitude shaking. By performing both bulk and microrheology experiments, we probe the phase behaviour and mechanical response of these nanocomposites. Slight tuning of either the particle or polymer concentration leads to dramatic changes in the macroscopic appearance of the mixture, transitioning from a low-viscosity fluid to a stiff gel, capable of sustaining its own weight. We relate these observations to microscopic measurements, providing insight into the local reorganisation of the constituting building blocks and the time-evolution of each phase.

Schizophyllan (SPG) is a semi-flexible, triple-helical polysaccharide with attractive properties as an efficient viscosifying compound and biological response modifier. We report microrheological characterization of schizophyllan as dispersed in solution and the changes associated when crosslinked with chitosan over an extended frequency range using diffusing wave spectroscopy (DWS). A SPG with high molecular weight (Mw = 1.1 × 106 Da) was selectively oxidized in the side chains (20 % or 40 %) to promote Schiff base formation with chitosan (CHI) amine groups, thus inducing crosslinking. The microrheological characterization of the dispersed SPG revealed characteristic features of the semiflexible structure, where also coupling between flexure and longitudinal modes was indicated based on scaling coefficient close to 7/8 of the loss modulus G"(ω) vs ω for ω in the range 3 × 103-105 rad/s. The in-situ characterization of the gelation process by DWS revealed changes in the scattered intensity-correlation function caused by the embedded colloidal probe-particles, from which the mean-square displacement of the probes and the shear moduli of the SPG-chitosan hydrogel samples were determined for various SPG concentrations and degrees of oxidation. It is found that SPG - chitosan hydrogels can be prepared with a polymer content in the range of 0.5-2.0 mg/mL and that tuning the molecular parameters allowed control of mechanical moduli in soft hydrogels in the range of 0.3 Pa up to 1000 Pa.

Focused ultrasound has experimentally been found to enhance the diffusion of nanoparticles; our aim with this work is to study this effect closer using both experiments and non-equilibrium molecular dynamics. Measurements from single particle tracking of 40 nm polystyrene nanoparticles in an agarose hydrogel with and without focused ultrasound are presented and compared with a previous experimental study using 100 nm polystyrene nanoparticles. In both cases, we observed an increase in the mean square displacement during focused ultrasound treatment. We developed a coarse-grained non-equilibrium molecular dynamics model with an implicit solvent to investigate the increase in the mean square displacement and its frequency and amplitude dependencies. This model consists of polymer fibers and two sizes of nanoparticles, and the effect of the focused ultrasound was modeled as an external oscillating force field. A comparison between the simulation and experimental results shows similar mean square displacement trends, suggesting that the particle velocity is a significant contributor to the observed ultrasound-enhanced mean square displacement. The resulting diffusion coefficients from the model are compared to the diffusion equation for a two-time continuous time random walk. The model is found to have the same frequency dependency. At lower particle velocity amplitude values, the model has a quadratic relation with the particle velocity amplitude as described by the two-time continuous time random walk derived diffusion equation, but at higher amplitudes, the model deviates, and its diffusion coefficient reaches the non-hindered diffusion coefficient. This observation suggests that at higher ultrasound intensities in hydrogels, the non-hindered diffusion coefficient can be used.

To minimize the incorrect use of antibiotics, there is a great need for rapid and inexpensive tests to identify the pathogens that cause an infection. The gold standard of pathogen identification is based on the recognition of DNA sequences that are unique for a given pathogen. Here, we propose and test a strategy to develop simple, fast, and highly sensitive biosensors that make use of multivalency. Our approach uses DNA-functionalized polystyrene colloids that distinguish pathogens on the basis of the frequency of selected short DNA sequences in their genome. Importantly, our method uses entire genomes and does not require nucleic acid amplification. Polystyrene colloids grafted with specially designed surface DNA probes can bind cooperatively to frequently repeated sequences along the entire genome of the target bacteria, resulting in the formation of large and easily detectable colloidal aggregates. Our detection strategy allows “mix and read” detection of the target analyte; it is robust and highly sensitive over a wide concentration range covering, in the case of our test target genome Escherichia coli bl21-de3, 10 orders of magnitude from 10 1 to 10 10 copies/mL. The sensitivity compares well with state-of-the-art sensing techniques and has excellent specificity against nontarget bacteria. When applied to real samples, the proposed technique shows an excellent recovery rate. Our detection strategy opens the way to developing a robust platform for pathogen detection in the fields of food safety, disease control, and environmental monitoring.

Short, partially complementary, single-stranded (ss)DNA strands can form nanostructures with a wide variety of shapes and mechanical properties. It is well known that semiflexible, linear dsDNA can undergo an isotropic to nematic (IN) phase transition and that sufficiently bent structures can form a biaxial nematic phase. Here we use numerical simulations to explore how the phase behaviour of linear DNA constructs changes as we tune the mechanical properties of the constituent DNA by changing the nucleotide sequence. The IN phase transition can be suppressed in so-called DNA 'nunchakus': structures consisting of two rigid dsDNA arms, separated by a sufficiently flexible spacer. In this paper, we use simulations to explore what phase behavior to expect for different linear DNA constructs. To this end, we first performed numerical simulations exploring the structural properties of a number of different DNA oligonucleotides, using the oxDNA package. We then used the structural information generated in the oxDNA simulations to construct more coarse-grained models of the rodlike, bent-core, and nunchaku DNA. These coarse-grained models were used to explore the phase behavior of suspensions of the various DNA constructs. The approach explored in this paper makes it possible to 'design' the phase behavior of DNA constructs by a suitable choice of the constituent nucleotide sequence.

Nano-sized particles functionalised with short single-stranded (ss)DNAs can act as detectors of complementary DNA strands. Here we consider tri-block-copolymer-based, self-assembling DNA-coated nanoparticles. The copolymers are chemically linked to the DNA strands via azide (N3) groups. The micelles aggregate when they are linked with complementary ssDNA. The advantage of such block-copolymer-based systems is that they are easy to make. Here we show that DNA functionalisation results in inter-micellar attraction, but that N3-groups that have not reacted with the DNA detector strands also change the phase behaviour of the tri-block polymer solution. We studied the triblock copolymer, Pluronic® F108, which forms spherical micelles in aqueous solutions upon heating. We find that the triblock chains ending with either an N3 or N3-DNA complex show a dramatic change in phase behaviour. In particular, the N3-functionalisation causes the chain ends to cluster below the critical micelle temperature (CMT) of pure F108, forming flower-micelles with the N3-groups at the core, while the PPO groups are exposed to the solvent. Above the CMT, we see an inversion with the PPO chains forming the micellar core, while the N3-groups are now aggregating on the periphery, inducing an attraction between the micelles. Our results demonstrate that, due to the two competing self-assembling mechanisms, the system can form transient hydrogels.

We characterize the self-assembly of colloidal particles with surface mobile DNA linkers under kinetically limited valence conditions. For this, we put forward a computer simulation model that captures the interplay between the main dynamic processes governing these systems and allows the simulation of the long time scales reached in experiments, predicting the final structures from a set of physical parameters mapped to experimental conditions. The model is validated by direct comparison with available experimental results, showing an overall good agreement that includes measurements of the average effective valence and its probability distribution as a function of the density of DNA linkers on the particles surface. We show that our simulation model is capable to not only reproduce the experimental structures but that it also provides dynamic information on the self-assembly process and shows how particle concentration and characteristic hybridization time influence the effective valence of the particles.

Abstract In this article, we show the DNA-functionalization of supraparticles, form their network, and manipulate the optical features of these networks by applying a magnetic field. We start with preparing the supraparticles (SPs) of semiconducting InP/ZnSeS/ZnS quantum dots (QDs), plasmonic silver nanoparticles, and superparamagnetic iron oxide nanoparticles. These SPs are prepared by employing azide-functionalized amphiphilic diblock or triblock copolymers as well as by using their combinations. Subsequently, we attached single-stranded DNAs to these SPs by employing copper-free click chemistry. Next, we hybridized DNA-coated QD SPs with the iron oxide SPs and formed a network. By applying a magnetic field, we restructured this network such that the iron oxide SPs are aligned. This led to an anisotropic emission from the QD SPs with a polarization ratio of 1.9. This study presents a proof-of-concept scheme to control the optical features of a self-assembled supraparticle system using an external interaction. We believe that our work will further contribute to the utilization of smart self-assembly techniques in optics and photonics. Impact statement The self-assembly of the nanoparticles, which lies at the heart of this article, enables achieving unconventional physical responses that cannot be obtained by the individual nanomaterials. Therefore, controlling the self-assembly process can lead to unprecedented control over the mechanical, optical, or electronic features of such novel architectures made of various types of nanoparticles. This smart self-assembly process will undoubtedly enable the realization of novel applications and open new avenues in materials science and engineering. In this article, we employ the self-assembly in two different aspects. First, we self-assembled magnetic, plasmonic, and semiconducting nanoparticles to obtain their supraparticles with the help of various amphiphilic polymers. Next, single-stranded DNA molecules were attached to them for the first time to achieve a precise control over their physical forming a “network of supraparticles of nanoparticles.” As a proof-of concept demonstration, we hybridized DNA-functionalized magnetic supraparticles made of iron oxide nanoparticles and DNA-functionalized light-emitting supraparticles containing InP/ZnSe/ZnS quantum dots. We explored the potential of tailoring the emission characteristics of the emitted light by utilizing an external magnetic field. We observed that under an external magnetic field the hybrid network reshapes; as a result, the polarization of the emitted light can be changed such that polarization anisotropy reaches ~1.9. We believe that our work presented here can initiate advanced-level active polarization control in displays and other light-emitting devices in the future. Considering that LCD-based displays require polarized light to function, the results of this work may serve for realizing efficient polarizer-free displays that rely on the manipulation of the structure under the magnetic field. Graphical abstract Single-stranded DNA molecules are attached to the supraparticles of semiconductor, metal, and magnetic nanoparticles to tailor optical features of the self-assembled network. An external magnetic field is employed to actively control the polarization of the light emitted by the network of semiconductor and magnetic supraparticles.

Prevention of protein aggregation and thus stabilization of proteins has large biological and biotechnological implications. Here we introduce Dynamic Light Scattering (DLS) and DLS-based microrheology to show how native bovine serum albumin (nBSA) forms amyloid fibrils in weakly denaturing conditions as function of time, and how stoichiometric conjugation of BSA with polymer-surfactants (PSpBSA) protects the protein form such aggregation. Employing a combination of Thioflavin-T fluorescence, Fourier transform infrared spectroscopy and other methods, we show that nBSA forms filamentous aggregates with amyloid-like structure, while PSpBSA proteins remain fully dispersed with only minor changes in their folding state, even when continuously heated for up to 5 days in denaturation conditions at 65 °C. Time-resolved DLS-based microrheology studies demonstrate that suspensions of the filamentous nBSA aggregates become viscoelastic for concentrations ≥200 μ M. Our results indicate that after 6 days in aggregation conditions, the elastic modulus G ′( ω ) of nBSA solutions went from zero initially to values of up to 3.6 Pa, indicating that the filaments become long enough to form an entangled, viscoelastic network. Interestingly, heating 200 μ M native BSA solutions at 65 °C for 2 days in Eppendorf tubes resulted in self-standing films rather than dispersed filaments. These films exhibited strong ThT-fluorescence intensities and a predominant β -sheet secondary structure in FTIR studies, suggesting that the self-standing microstructure of the film resulted from hierarchical self-assembly of the amyloid fibrils.

Spatially ordered arrangements of spherical colloids are known to exhibit structural colours. The intensity and brilliance of these structural colours typically improve with colloidal monodispersity, low concentrations of point and line defects and with increasing refractive index contrast between the colloids and the embedding medium. Here we show that suspensions of charge stabilised, fluorinated latex particles with low refractive-index contrast to their aqueous background form Wigner crystals with FCC symmetry for volume fractions between 13 and 40%. In reflection they exhibit both strong, almost angle-independent structural colours and sharp, more brilliant Bragg peaks despite the particle polydispersity and bimodal distribution. Simultaneously, these suspensions appear transparent in transmission. Furthermore, binary AB, A 2 B and A 13 B type mixtures of these fluorinated and similarly sized polystyrene particles appeared predominantly white but with clear Bragg peaks indicating a CsCl-like BCC structure and more complex crystals. We characterised the suspensions using a combination of reflectivity measurements and small-angle x-ray scattering, complemented by reflectivity modelling.

We study the elastic response of concentrated suspensions of rigid wire frame particles to a step strain. These particles are constructed from infinitely thin, rigid rods of length L. We specifically compare straight rod-like particles to bent and branched wire frames. In dense suspensions, the wire frames are frozen in a disordered state by the topological entanglements between their arms. We present a simple, geometric method to find the scaling of the elastic stress with concentration in these glassy systems. We apply this method to a simple 2D model system where a test particle is placed on a plane and constrained by a random distribution of points with number density ν. Two striking differences between wire frame and rod suspensions are found: (1) The linear elasticity per particle for wire frames is very large, scaling like ν2L4, whereas for rods, it is much smaller and independent of concentration. (2) Rods always shear thin but wire frames shear harden for concentrations less than ∼K/kBTL4, where K is the bending modulus of the particles. The deformation of wire frames is found to be important even for small strains, with the proportion of deformed particles at a particular strain, γ, being given by (νL2)2γ2. Our results agree well with simple numerical calculations for the 2D system.

Article Figures and data Abstract Introduction Results Discussion Materials and methods Data availability References Decision letter Author response Article and author information Metrics Abstract Measuring protein-protein interaction (PPI) affinities is fundamental to biochemistry. Yet, conventional methods rely upon the law of mass action and cannot measure many PPIs due to a scarcity of reagents and limitations in the measurable affinity ranges. Here, we present a novel technique that leverages the fundamental concept of friction to produce a mechanical signal that correlates to binding potential. The mechanically transduced immunosorbent (METRIS) assay utilizes rolling magnetic probes to measure PPI interaction affinities. METRIS measures the translational displacement of protein-coated particles on a protein-functionalized substrate. The translational displacement scales with the effective friction induced by a PPI, thus producing a mechanical signal when a binding event occurs. The METRIS assay uses as little as 20 pmols of reagents to measure a wide range of affinities while exhibiting a high resolution and sensitivity. We use METRIS to measure several PPIs that were previously inaccessible using traditional methods, providing new insights into epigenetic recognition. Introduction Protein-protein interactions (PPIs) are essential to cellular biology and both high- and low-affinity interactions are required to maintain robust and dynamic responses in biological circuits (Nooren and Thornton, 2003; Kastritis and Bonvin, 2013; Evans, 2001). Low-affinity interactions are commonly leveraged, as is seen for multivalent recognition (Markin et al., 2010), readers of highly abundant proteins, and in protein allostery (Daily and Gray, 2009). In particular, recognition of the epigenome is recognized to rely on the interplay between post-translational modifications (PTMs), like methylation, phosphorylation, and ubiquitination (Yau et al., 2017; McGinty and Tan, 2016; Patel, 2016). Furthermore, multidomain-containing proteins are often regulated by allostery through weak interdomain interactions (Gladkova et al., 2018; Peng, 2015). Increasingly, the importance of weak interactions or relatively small changes in PPI affinity has been realized. Despite the increasing sophistication of studying PPIs, biochemical characterization of these weaker and similar strength interactions remain a significant hurdle. Many techniques are useful for examining protein-binding strength, each with its own set of limitations (Rowe, 2011; Syafrizayanti et al., 2014; Pollard, 2010). However, virtually all the commonly used techniques to measure biological interactions, for example, like ELISA, FP, SPR, NMR, BLI, AUC, and ITC, rely on the law of mass action, and to measure protein binding affinities in the μM range and above, highly concentrated proteins or ligands are required (Xing et al., 2016). For many systems, obtaining such large quantities of materials can be unattainable. Furthermore, high protein concentrations leads to thermodynamic non-ideality and proteins can aggregate, self-associate, and non-specific interactions occur, thus obfuscating the binding signal (White et al., 2010; Saluja and Kalonia, 2008). NMR is the gold standard method to measure weak interactions; however, in addition to requiring copious amounts of materials, the proteins must also be isotopically labeled, a single affinity measurement requires substantial instrument time and complex data analysis, and of all the methods mentioned is the lowest throughput. BLI and SPR are methods that can measure interactions while using a small quantity of the immobilized partner, however, binding is still governed by mass action and the soluble analyte must be at concentrations above the Kd; for weak binders, this can still use large quantities of materials (Helmerhorst et al., 2012; Weeramange et al., 2020). Additionally, the signal is highly dependent on the mass change of the interaction, and for smaller ligands, binding to larger molecules this signal could be small. Moreover, discerning between background binding and specific binding can be difficult, especially for weak interactions which requires the analyte to be at a high concentration. Another difficulty in determining the binding affinity of PPIs arises when measuring similar strength interactions, for example, two- to fivefold differences. Several factors contribute to this limitation, but determining the active fraction of protein is significant because, for most fitting techniques, the calculated affinity is a dependent variable of the protein concentration (Jarmoskaite et al., 2020; Hulme and Trevethick, 2010). Moreover, to achieve accurate fitting of a protein binding isotherm requires accurate determination of the end point of the saturation curve, which for weak interactions necessitates high concentrations of ligand. Another factor in differentiating similar strength interactions is that most binding measurements have low statistical power due to the resource intensiveness of performing multiple replicates. A method where binding strength can be measured independent of protein concentration, that uses small amounts of reagents, and that has high statistical power would be valuable. Here, we present a novel approach to measuring the strength of biological interactions that is moderately high-throughput, requires a minimal amount of protein material, and can measure a wide range of Kd values from 10-2 to 10-15 M. This technique was initially inspired by the rolling of biological cells, like neutrophils exhibiting haptotaxis on endothelial cells. Neutrophil motion is driven by chemical or ligand gradients (Voisin and Nourshargh, 2013). The neutrophils roll on the endothelial cells due to PPIs between the cell surface receptors. The PPIs increase the effective friction between the two cells, allowing the rotational motion to be converted into translational displacement. We aimed to create a single particle biomimetic technique that leveraged this fundamental physical concept of friction to produce a mechanical signal to indicate binding events, the Mechanically Transduced Immunosorbent assay (METRIS). METRIS utilizes protein functionalized ferromagnetic particles to mimic the rolling cells. These ferromagnetic particles are made active via actuation of an externally applied rotating magnetic field and the particles proceed to roll, henceforth referred to as rollers, and translate across the surface using a similar mode of locomotion as the neutrophils. When the rollers are placed on a functionalized surface, the amount of rotational motion converted into translational motion depends on the effective friction between the rollers and the substrate. That effective friction scales with the strength of the binding interaction. Thus, a higher affinity PPI between the roller and the substrate will result in a larger translational displacement of the roller. Since both the roller and surface have immobilized proteins, the method is not dependent on mass action and requires approximately 20 pmols to measure PPIs regardless of their strength. Using the METRIS assay, we reproduced well-characterized binding preferences for two different methyllysine histone reader domains (Gatchalian et al., 2013; Kuo et al., 2012) and weak interactions between the E2 Ube2D (Buetow et al., 2015) and UBL-domains (DaRosa et al., 2018). These affinities range between 10-4 and 10-6 M. However, we were also able to measure several weaker interactions between unmodified histone peptides, which allowed us to measure the ΔΔGs for the phospho/methyl switch phenomenon in DIDO1-PHD (Andrews et al., 2016). Finally, we also show that this method can be used to measure a weak interdomain interaction between the isolated UHRF1-UBL domain and SRA domain, which is known to control the E3 ligase specificity and epigenetic DNA methylation inheritance (Foster et al., 2018; DaRosa et al., 2018). Collectively, our results show that the METRIS assay can be a very powerful technique which has the potential to provide additional insight into PPI interactions that were difficult to measure using other methods. Results Rolling parameter scales with interaction affinity of PPI In the METRIS assay, rollers are placed in a Helmholtz coil inspired apparatus (see Figure 1A and Figure 1—figure supplement 1) where an externally rotating magnetic field is applied at a constant frequency, ω. The permanent magnetic moment of the roller, couples with the applied magnetic field, producing a magnetic torque and subsequent rotation of the ferromagnetic bead (Steimel et al., 2014; Sing et al., 2015). In the absence of effective friction, the rollers would rotate mostly in place with the frequency of the applied magnetic field; however, effective friction induced by binding between the rollers and the substrate will convert some of that rotational motion into translational displacement, Δ⁢x, thus indirectly measuring the effective friction between the substrate and the rollers. Since the magnetic field is many orders of magnitude stronger than the strength of noncovalent interactions, in this system the higher the effective friction between the rollers and the surface corresponds to larger translational displacement. The effective friction is determined by the strength and density of PPIs between the roller and the coated substrate. Thus, the translational displacement will scale with the density and affinity of the PPIs being measured, such that a higher Δ⁢x corresponds to a higher affinity. However, the displacement is also a function of several other parameters, specifically the diameter of the roller, D, and the frequency of rotation of the applied magnetic field, ω. Here we define a dimensionless parameter, which is the ratio of the observed translational displacement of the roller to the maximum theoretical translational displacement of a sphere that we refer to as the rolling parameter, RP (Figure 1B) (1) R⁢P=Δ⁢xπ⁢D⁢τ⁢ω Figure 1 with 3 supplements see all Download asset Open asset Experimental schematic of the Mechanically Transduced Immunosorbent Assay (METRIS) used to measure protein-protein interactions. (A) General schematic of roller and surface functionalization. Both binder A and binder B are attached to the roller or the surface by biotin-streptavidin interactions. The direction of the rotating magnetic field is indicated by the curved arrow. (B) The rolling parameter (RP) is a dimensionless parameter that measures rolling. The RP is calculated by taking the ratio between the observed displacement of a roller, Δ⁢x, and the maximum theoretic rolling of a sphere, which is calculated from the circumference of each spherical particle ΠD, the frequency of the rotation of the magnetic field ω, and the actuation time τ. In the schematic three scenarios are depicted: (top) a RP of 1 where the roller moves the maximum theoretical displacement, (middle) a RP less than 1, and (bottom) a RP near 0 where the particle does not move. (C) Representative microscopy images of streptavidin rollers (black points) on an avidin (left) or a biotin surface (right). The scale bar in black is 100 μm and the images size are 1.28 mm × 0.96 mm. The position of the rollers prior to magnetic field actuation are indicated by the transparent spots and after actuation is opaque. The top panels are after a CW actuation and the bottom is after a CCW actuation. The magnification illustrates the difference between the null (streptavidin-avidin) and biotin-streptavidin interaction translational displacement. (D) Plot of a single roller from a streptavidin-biotin (black) and an avidin-streptavidin (red) experiment. The y-axis (X) represents the position of each roller in the field of view. The magnification above shows how Δx is calculated for each roller by subtracting the preactuation position from the postactuation position. CW and CCW actuations are repeated as described in the methods. Translational displacement is calculated as a vector. (E) The distribution of rolling parameters (RPs) from the streptavidin-avidin (N=8 rollers) and biotin-streptavidin (N=9 rollers) experiments. RP is calculated using the equation in (B) for each actuation period for each roller so the distributions contain N × 36 points. See Figure 1—source data 1 for the rolling parameter for each actuation. Figure 1—source data 1 Rolling parameter from all rolls for either biotin-streptavidin and avidin-streptavidin. https://cdn.elifesciences.org/articles/67525/elife-67525-fig1-data1-v1.xlsx Download elife-67525-fig1-data1-v1.xlsx where Δx is the translational displacement of the roller, D is the diameter of the roller, τ is the actuation period of the magnetic field, and ω is the rotational frequency of the magnetic field. The RP is a dimensionless parameter that varies from 0 to 1 where 0 is no translational displacement and one being a sphere perfectly rolling at a single hinge point and translating a distance equivalent to its circumference. Here, the sphere also undergoes a number of rotations given by product of the rotational frequency of the magnetic field and actuation time. The density of the interactions between the roller and the substrate are kept as constant as possible from experiment to experiment by fully saturating both the rollers and the substrate with proteins and peptides. As described in the Materials and methods, both the rollers and substrate are coated 50× the theoretical number of binding sites, so virtually all the sites should be occupied. Additionally, a series of washing steps are carried out to make sure no unbound protein or peptide remains on the surface. If the surface was not uniformly functionalized, the roller's displacement in these regions would be detected by correlations to either the individual roller or areas on the substrate. However, no such anomalies were observed in these experiments. To measure the translation displacement Δ⁢x and to calculate the RP of the rollers, a clockwise (CW) field was actuated at ω=1⁢H⁢z for τ=5 s. The field was then turned off for τ=5 s. A counter-clockwise (CCW) field was actuated at ω=1⁢H⁢z for τ=5 s and then the field was turned off for τ=5 s again. This process was repeated 18 times, and several example images of rollers and roller trajectories can be seen in Figure 1C and Figure 1D and in the supplemental videos. Appropriate parameters for the magnetic field strength and frequency were previously determined (Steimel, 2017). The rolling parameter is calculated from the observed roller translational displacement divided by the maximum theoretical translational displacement of a rolling sphere where all the rotational torque is converted into translation, so the rolling parameter varies from 0 to 1. A rolling parameter of 0 corresponds to a surface with no effective friction. Experimentally, a rolling parameter of 0 is never observed due to hydrodynamic friction between the roller and the substrate. A rolling parameter of 1 corresponds to the maximum theoretical rolling of a sphere. We first measured the rolling of streptavidin rollers on an avidin surface or a biotin surface. Still images of a CW (top) and CCW (bottom) actuation (Figure 1C) show that on the streptavidin surface the rollers hardly move, while on the biotin surface the rollers translate well over 100 μm. A full trajectory for a roller on the biotin and streptavidin surfaces (Figure 1D) show the Δx for each roller remains relatively constant through each actuation (see Figure 1—video 1 and Figure 1—video 2 for movies of the experiment). Δ⁢x is calculated for each actuation for each roller and then converted to a rolling parameter (RP) (Figure 1D). The RPs have a gaussian distribution (Figure 1E) and the average RP on the avidin surface is 0.081 ± 0.004 while on biotin we observed a RP of 0.918 ± 0.002. The interaction between biotin and streptavidin is reported to be Kd=10-15 M. While it is impossible to know the true Kd value for a null interaction, the weakest PPI measured are in the 10-2 M range (Yoo et al., 2016) and enzymes with Kd values in the 10° M range have been reported (Bar-Even et al., 2011), so we assume that null interaction must be between 10° M and the concentration of water 5.5 × 102M, and we settled on 10°M as an estimation of the null interaction, which based on our subsequent fitting seems like a suitable assumption. These two values provide an idea about the range of affinities that can be measured with METRIS. DIDO1-PHD phospho/methyl switch characterized by METRIS Next, we wanted to determine whether we can quantitatively correlate the measured RP to binding affinities for known PPIs and test the robustness of the METRIS assay as an experimental approach to measure PPIs. We focused our attention on weak interactions and interactions between several protein pairs that are similar in binding strength, given that these PPIs are typically difficult to accurately measure. We first examined the well-established interaction between DIDO1-PHD and H3K4 methylation. DIDO1 is responsible for interchanging between active and silent chromatin states in embryonic stem cells, and its chromatin localization is regulated through a phospho/methyl switch, where phosphorylation of H3T3 evicts DIDO1 from chromatin during mitosis (Fütterer et al., 2017; Di Lorenzo and Bedford, 2011; Liu et al., 2014). The affinities for mono-, di-, and trimethylated peptides are well described in the literature (Gatchalian et al., 2013) and interactions with the unmodified peptide and H3T3pK4me3 were too weak to be measured in the experiment setup. H3K4 peptides and DIDO1-PHD were both immobilized to the rollers and substrate through biotin-streptavidin interactions. The H3 N-terminus (a.a. 1–20) was biotinylated and immobilized on the roller, and biotinylated avi-tagged GST-DIDO1-PHD was attached to the substrate. DIDO1 has a preference for H3K4me3 > H3K4me2 > H3K4me1 (Gatchalian et al., 2013). The measured Δ⁢x and RP match this preference, with the largest rolling parameter for H3K4me3 (0.233 ± 0.012) > H3K4me2 (0.213 ± 0.010) > H3K4me1 (0.176 ± 0.005) and H3 and H3T3pK4me3 being the lowest, although still above the baseline rolling parameter value of 0.081 (Figure 2A and B). While the overall change to the RPs is small, these differences are all statistically significant because the data set has good statistical power and small percentage errors (<5%) (Figure 2—source data 2). Additionally, the distribution of rolling parameters can be found in Figure 2—figure supplement 1A, Figure 2—video 1 and 2, and Figure 2—animation 3, show the rolling for this family of interactions. Figure 2 with 4 supplements see all Download asset Open asset DIDO1-PHD interactions with H3 peptides characterized using METRIS. (A) Plot showing the average translational displacement per actuation for the rollers coated with the indicated H3K4 methylated peptide on a DIDO1-PHD surface. Streptavidin-biotin and streptavidin-avidin (PBS) are included for references. See Figure 2—source data 2 for results of statistical analysis; all comparisons are statistically significant (p<0.0001). (B) Plot showing the calculated average rolling parameter per interaction. (C) Log-Log plot of the rolling parameters (RP) from panel B with the reported Kds. Extrapolated points for the unknown interactions are represented by unfilled markers, and the 95% confident interval for the fitting is depicted. (D) Table of rolling parameters and associated Kd estimates for the DIDO1-PHD interactions. Fold change is calculated as the ratio between the Kd values for the indicated peptide and for H3K4me3. These ratios are used to calculate ΔΔG at T=298K. The published values are from Gatchalian et al., 2013 using NMR (me1) and tryptophan fluorescence (me2/3); *ND = Not determined. (E) Image of the DIDO1-PHD crystal structure with H3K4me3 peptide, with the PHD surface electrostatic potentials shown (red = negative, blue = positive), the ΔΔG for K4me3, and the estimated ΔΔG for the rest of the peptide. The PTM reader sites are shown with greater detail to the right. Here, ΔΔG is calculated between the sequential methyl states, and the ratio of H3T3pK4me3 and H3K4me3 give the ΔΔG for T3p. (F) Results of the DIDO1-PHD histone peptide microarray assay against the indicated peptides (see Figure 1—figure supplement 1B for results of all peptides). Only H3K4me3 is statistically significant (P<0.05). (see Figure 2—source data 2 for results of statistical analysis). While these results indicate general binding trends, they cannot provide Kd estimates and do not have high enough resolution to distinguish between weaker binding interactions. Figure 2—source data 1 Rolling parameter from all rolls for the indicated rollers on a DIDO1-PHD surface. Each row is a different roller and each column is an actuation. https://cdn.elifesciences.org/articles/67525/elife-67525-fig2-data1-v1.xlsx Download elife-67525-fig2-data1-v1.xlsx Figure 2—source data 2 Statistical analysis of results from METRIS measurements and histone peptide microarray results for DIDO1-PHD. Results from statistical tests comparing the indicated pairs of rolling parameters (METRIS) or microarray results (Array). https://cdn.elifesciences.org/articles/67525/elife-67525-fig2-data2-v1.xlsx Download elife-67525-fig2-data2-v1.xlsx In order to correlate binding affinity to RP, we developed an empirical fitting method based on available data. We noticed that the log-log plot of Kd vs. RP showed a linear relationship between the three known DIDO1-PHD binding interactions to the methylated peptides (R2=0.995). We also included a no-binding avidin-streptavidin interaction (RP=0.081) estimated to have a Kd = 1M and the streptavidin-biotin interaction where Kd = 10-15 M (DeChancie and Houk, 2007; Figure 2C). Overall, this experiment suggests that there is a linear dependence of the log of the RP to the log of Kd that spans roughly fifteen-orders of magnitude. There is a clear correlation between RP and the measured Kd, the equilibrium constant for interactions, despite METRIS being a non-equilibrium technique. Kd is a ratio between the first-order dissociation rate (Koff) and the second-order association rate (Kon) (Sanders, 2004). For most PPIs, the Kon rates are very similar, and thus the Kd constant is mostly dependent on Koff. However, kinetic constants for binding interactions are rarely reported since few techniques can access this information, so for many interactions, only Kd is known. Since we do not have a theoretical model that relates RP to Kd, we sought to use an empirical fitting method based on the excellent correlation we observed between RP and Kd (Figure 2C). Using this fitting method, we could reproduce the literature Kd values with high accuracy; all of the predicted Kd values were roughly twofold tighter than the published values (Gatchalian et al., 2013) and the fold difference between the different methylation states is similar (Figure 2D). Remarkably, we were also able to estimate METRIS-Kd values for the weak interaction between the H3T3pK4me3 peptide (340 μM ± 90) and the unmodified H3 tail (1200 μM ± 440). While these are empirically derived estimates for Kd, it is clear from the RP measurements that these interactions are statistically distinct, and they represent a missing piece of data that is fundamental to a quantitative understanding of epigenetic recognition. The utility of the METRIS data is exemplified when evaluating the ΔΔGo (ΔΔG) values, a common way to report the energetic contributions of individual amino acids for a set of related PPIs. ΔΔG is calculated by taking the natural log of the ratio of two Kd values (Kd1 and Kd2 in equation 2) in the Gibbs free energy equation, where R is the gas constant and T is the temperature in Kelvin (Sidhu and Koide, 2007). (2) Δ⁢Δ⁢Go⁢(Δ⁢Δ⁢G)=R⁢T⁢ln⁡Kd⁢1Kd⁢2 This analysis allows for calculating the energetic contributions of the individual PTMs for binding to the DIDO1-PHD domain. For example, K4me3 is worth −4.2 k⁢c⁢a⁢l⁢sm⁢o⁢l while T3p is worth +3.4 k⁢c⁢a⁢l⁢sm⁢o⁢l (Figure 2D). To our knowledge, this is the first energetic analysis of the DIDO1 phospho/methyl switch. These values have more context when viewed with the crystal structure of DIDO1-PHD (Figure 2E; Gatchalian et al., 2013). The hydrophobic trimethyl-lysine binding site accounts for a significant amount of the total binding to the peptide, however, there are clearly other residues on H3 that interact with DIDO1-PHD, such as the N-terminus, R2, and T3, and therefore, it is not surprising that unmodified H3 can still bind and account for roughly −4 k⁢c⁢a⁢lm⁢o⁢l when using 1 M Kd as the null reference. The deleterious effect of T3p is also resolved, since residue E308 of the PHD domain would clash and repel a T3p modified histone tail. Furthermore, this analysis also provides new insights into discrimination of methylation states by the DIDO1-PHD. For example, the greatest change in ΔΔG occurs between H3 from H3K4me1 (−1.9 k⁢c⁢a⁢lm⁢o⁢l), then H3K4me1 versus H3K4me2 (−1.6 k⁢c⁢a⁢lm⁢o⁢l), and H3K4me2 from H3K4me3 is the weakest (−0.7 k⁢c⁢a⁢lm⁢o⁢l). Thus, despite the DIDO1-PHD having the highest affinity for H3K4me3, it has the greatest discrimination between non-methylated H3K4 versus H3K4me1. The structure agrees with this observation, where two of the methyl binding sites are the most buried and the third is the most exposed one. One of the significant advantages of the METRIS assay is that only 10 μl of 2 μM (20 pmol) is required to load the substrate and less is needed for the rollers, which is significantly less than any conventional method to measure PPI affinities. We compared METRIS to histone peptide microarrays, which is another methodology that can produce binding data with a minimal amount of protein (e.g. 500 μl of 0.5 μM [250 pmol] protein). While microarrays offer high-throughput screening, they lack the sensitivity to determine weak binding and small affinity differences. For DIDO1-PHD, we could observe a statistically significant difference between H3K4me3 and the other methylation states, but there were no other statistically significant differences (Figure 2F, Figure 2—figure supplement 1B, and Figure 2—source data 2). Given this result, METRIS is significantly more sensitive and quantitative than other common methods to measure protein affinities that use comparable amounts of reagents at low concentrations (i.e. ELISA and microarrays). Determining ORC1-BAH methyl preferences using METRIS analysis We further validated the METRIS assay using another methyllysine reader, the BAH domain of ORC1. ORC1 functions in licensing origins of replication by discriminating H4K20me2 from H4K20me1, a PTM on active chromatin, and H4K20me3 a repressive PTM (Bicknell et al., 2011a; Bicknell et al., 2011b; Kuo et al., 2012). We selected ORC1 because the reported affinities are within an order of magnitude, with a twofold difference reported between H4K20me1 and H4K20me3. The Δx and RP values we obtained matched the published binding preferences (Kuo et al., 2012) H4K20me2 (0.263±0.011) > H4K20me1 (0.226 ± 0.008) >H4K20me3 (0.215 ± 0.005)> H4 (0.202 ± 0.005) (Figure 3A, Figure 3B, Figure 3—figure supplement 1A, Figure 3—video 1, Figure 3—video 2, and Figure 3—animation 1). Using the same fitting method, we observe a linear log-log dependence (R2 = 0.967) and the METRIS calculated Kd values were between four- and eightfold tighter than the published values, yet there was good agreement between the fold-change and accordingly the ΔΔGs. (Figure 3C and Figure 3D). Thus, the METRIS assay is sensitive enough to measure changes that are 0.4 k⁢c⁢a⁢lm⁢o⁢l. Figure 3 with 4 supplements see all Download asset Open asset ORC1-BAH domain interactions characterized using METRIS. (A) Plot showing the average translational displacement per actuation for the rollers immobilized with the H4K20 methylated peptide on a ORC1-BAH domain surface. Streptavidin-Biotin and Streptavidin-avidin (PBS) are included for references. See Figure 3—source data 2 for results of statistical analysis; all comparisons are significant (p<0.0001). (B) Plot showing the calculated RP for the indicated interactions (C) Log-Log plot of the rolling parameters, RP, from panel A. Extrapolated point markers are unfilled and the 95% Kd confident interval for the fitting is depicted. (D) Table of rolling parameters and associated Kd estimates for the ORC1-BAH. Fold change is calculated as the ratio between the Kd for the indicated peptide and the Kd for H4K20me2. These ratios are used to calculate ΔΔG at T=298K. The published values are from Kuo et al., 2012 using ITC; *ND = Not determined. (E) Image of the ORC1-BAH crystal structure with H4K20me2 peptide, with the BAH surface electrostatic potentials shown (red = negative, blue = positive) as well as the ΔΔG for K20me2 and the estimate for the rest of the peptide. The PTM reader site is shown with greater detail to the right. Here the ΔΔG is calculated between the sequential methyl states. (F) Results of the ORC1-BAH histone peptide microarray assay against the indicated peptides from panel A (see Figure 3—figure supplement 1B for complete peptide plot). Only H4K20me2 is statistically significantly different (p<0.05) from the other H4 peptides (Figure 3—source data 2). Again, we see that microarrays can indicate general binding trends but they cannot provide Kd estimates and do not have high enough resolution to distinguish between weaker binding interactions. Figure 3—source data 1 Rolling parameter from all rolls for the indicated rollers on a ORC1-BAH surface. Each row is a different roller and each column is an actuation. https://cdn.el

We study the elastic response of rigid wire frame particles in concentrated glassy suspensions to a step strain by applying the simple geometric methods developed in Paper I. The wire frame particles are comprised of thin rigid rods of length L, and their number density, ρ, is such that ρL3 ≫ 1. We specifically compare rigid rods to L-shapes made of two equal length rods joined at right angles. The behavior of wire frames is found to be strikingly different from that of rods. The linear elasticity scales like ρ3L6 for L-shaped particles, whereas it scales proportional to ρ for rods and the non-linear response shows a transition from shear hardening to shear softening at a critical density ρc∼K/kBTL6, where K is the bending modulus of the particles. For realistic particles made of double stranded DNA, this transition occurs at densities of about ρL3 ∼ 10. The reason for these differences is that wire frames can be forced to bend by the entanglements with their surroundings, whereas rods always remain straight. This is found to be very important even for small strains, with most particles being bent above a critical strain γc∼ρL3 -1.

We measure by experiment and particle-based simulation the rheology of concentrated, non-Brownian droplet emulsions functionalized with surface-bound single-stranded (ss), "sticky," DNA. In the absence of ssDNA, the emulsion viscosity increases with the dispersed phase volume fraction ϕ, before passing through a liquid-solid transition at a critical ϕ_{c} related to random close packing. Introducing ssDNA leads to a liquid-solid transition at ϕ<ϕ_{c}, the onset being set by the droplet valency N and the ssDNA concentration (or simulated binding strength ε). Using insight from simulation, we identify three key behaviors: (i) jammed suspensions (ϕ>ϕ_{c}≈0.64) show weak effects of functionalization, with elastic rheology instead governed by droplet stiffness; (ii) suspensions with ϕ<ϕ_{c} and N=1, 2 always exhibit viscous rheology, regardless of functionalization; and (iii) for ϕ<ϕ_{c} and N>3, functionalization leads to a controllable viscous-elastic transition. We present state diagrams showing the range of rheological tuning attainable by these means.

Prevention of protein aggregation and thus stabilization of proteins has large biological and biotechnological implications. Here, we show that inhibition of amyloid-like aggregates is possible in stoichiometric conjugates of polymer surfactant and bovine serum albumin (BSA) chosen as a model protein. We investigate using a combination of Thioflavin-T fluorescence spectroscopy, dynamic light scattering and FTIR spectroscopy the aggregation behavior in polymer surfactant modified and unmodified (native) BSA solutions. The BSA-polymer surfactant conjugates are stable up to 5 days under aggregation conditions, while native BSA forms amyloid fibrillar structures. Further, DLS-based micro-rheology studies performed with heat-treated 100 to 200 {\mu}M native BSA aggregates provided understanding of the equilibrium elastic and viscous moduli over a very large frequency range, reaching MHz, which are inaccessible using bulk rheology. Our results indicate that after 6 days of aggregation conditions, elastic moduli showed values between 1.2 to 3.6 Pa corresponding to an entanglement length ({\xi}) of 105 nm. Interestingly, heating 200 {\mu}M native BSA solution at 65 degree C for 2 days in a plastic Eppendorf resulted in self-standing films. These films exhibited strong ThT-fluorescence intensity and a predominant \b{eta}-sheet secondary structure from the FTIR studies, suggesting that self-standing microstructure resulted from hierarchical self-assembly of amyloid fibrils.

Inspired by the relationship between the well-ordered architecture of aragonite crystals and biopolymers found in natural nacre, we present a facile strategy to construct large-scale organic/inorganic nacre-mimetics with hierarchical structure via a water-evaporation driven self-assembly process. We connect LAPONITE®-nanoclay platelets with each other using carboxymethyl cellulose, a cellulose derivative, thus creating thin, flexible films with a local brick-and-mortar architecture. The dried films show a pronounced resistance against tensile forces allowing for stronger thin films than nacre. In terms of functionalities, we report excellent glass-like transparency along with exceptional shape-persistent flame shielding. We also demonstrate that through metal ion-coordination we can further strengthen the interactions between the polymers and the nanoclays, and thus enhanced mechanical, and thermal properties as well as resistance against swelling and dissolution in aqueous environments. We believe that our simple pathway to fabricate such versatile polymer/clay nanocomposites can open avenues for inexpensive production of environmentally friendly, biomimetic materials in aerospace, wearable electrical devices, and in the food packaging industry.

Laponite is a synthetic clay that is known to form gels in aqueous suspensions at low concentrations (0.01 g/cm3). Although it is expected to form lyotropic liquid crystals, such phases usually do not form, as a consequence of laponite's tendency to form gels at concentrations below the threshold for liquid crystal formation. Here we show that macroscopic, birefringent phases of laponite can be prepared through osmotic compression of a laponite solution by an aqueous solution of carboxy methyl cellulose (CMC). We present polarization imaging studies showing how the initially dilute, isotropic laponite phase shrinks while developing typical birefringence colors between crossed polarizers. Using the Michel-Lévy interference charts, we were able to extract the refractive index and orientation of the laponite nanodisks in the compressed region. Our observations allow us to propose a tentative state diagram, indicating the concentration regions for which we obtain optically anisotropic gels.

It is well known that suspensions of particles in a viscous fluid can affect the rheology significantly, producing a pronounced non-Newtonian response even in dilute suspension. However, it is unclear a priori which particle shapes lead to this behavior. We present two simple symmetry conditions on the shape which are sufficient for a dilute suspension to be Newtonian for all strain sizes and one sufficient for Newtonian behavior for small strains. We also construct a class of shapes out of thin, rigid rods not found by the symmetry argument which share this property for small strains.