Building block aspect ratio controls assembly, architecture, and mechanics of synthetic and natural protein networks.

Fibrous networks constructed from high aspect ratio protein building blocks are ubiquitous in nature. Despite this ubiquity, the functional advantage of such building blocks over globular proteins is not understood. To answer this question, we engineered hydrogel network building blocks with varying numbers of protein L domains to control the aspect ratio. The mechanical and structural properties of photochemically crosslinked protein L networks were then characterised using shear rheology and small angle neutron scattering. We show that aspect ratio is a crucial property that defines network architecture and mechanics, by shifting the formation from translationally diffusion dominated to rotationally diffusion dominated. Additionally, we demonstrate that a similar transition is observed in the model living system: fibrin blood clot networks. The functional advantages of this transition are increased mechanical strength and the rapid assembly of homogenous networks above a critical protein concentration, crucial for in vivo biological processes such as blood clotting. In addition, manipulating aspect ratio also provides a parameter in the design of future bio-mimetic and bio-inspired materials.

Reaction Rate Governs the Viscoelasticity and Nanostructure of Folded Protein Hydrogels.

Hydrogels constructed from folded protein domains are of increasing interest as resilient and responsive biomaterials, but their optimization for applications requires time-consuming and costly molecular design. Here, we explore a complementary approach to control their properties by examining the influence of crosslinking rate on the structure and viscoelastic response of a model hydrogel constructed from photochemically crosslinked bovine serum albumin (BSA). Gelation is observed to follow a heterogeneous nucleation pathway in which BSA monomers crosslink into compact nuclei that grow into fractal percolated networks. Both the viscoelastic response probed by shear rheology and the nanostructure probed by small-angle X-ray scattering (SAXS) are shown to depend on the photochemical crosslinking reaction rate, with increased reaction rates corresponding to higher viscoelastic moduli, lower fractal dimension, and higher fractal cluster size. Reaction rate-dependent changes are shown to be consistent with a transition between diffusion- and rate-limited assembly, and the corresponding changes to viscoelastic response are proposed to arise from the presence of nonfractal depletion regions, as confirmed by SAXS. This controllable nanostructure and viscoelasticity constitute a potential route for the precise control of hydrogel properties, without the need for molecular modification.

In Silico Modeling of the Rheological Properties of Covalently Cross-Linked Collagen Triple Helices.

Biomimetic hydrogels based on natural polymers are a promising class of biomaterial, mimicking the natural extra-cellular matrix of biological tissues and providing cues for cell attachment, proliferation, and differentiation. With a view to providing an upstream method to guide subsequent experimental design, the aim of this study was to introduce a mathematical model that described the rheological properties of a hydrogel system based on covalently cross-linked collagen triple helices. In light of their organization, such gels exhibit limited collagen bundling that cannot be described by existing fibril network models. The model presented here treats collagen triple helices as discrete semiflexible polymers, permits full access to metrics for network microstructure, and should provide a comprehensive understanding of the parameter space associated with the development of such multifunctional materials. Triple helical hydrogel networks were experimentally obtained via the reaction of type I collagen with both aromatic and aliphatic diacids. The complex modulus G* was found from rheological testing in linear shear and quantitatively compared to model predictions. In silico data from the computational model successfully described the experimental trends in hydrogel storage modulus with either (i) the concentration of collagen triple helices during the cross-linking reaction or (ii) the type of cross-linking segment introduced in resulting hydrogel networks. This approach may pave the way to a step change in the rational design of biomimetic triple helical collagen systems with controlled multifunctionality.

Ecological approaches to oral biofilms: control without killing.

Humans have co-evolved with micro-organisms and have a symbiotic or mutualistic relationship with their resident microbiome. As at other body surfaces, the mouth has a diverse microbiota that grows on oral surfaces as structurally and functionally organised biofilms. The oral microbiota is natural and provides important benefits to the host, including immunological priming, down-regulation of excessive pro-inflammatory responses, regulation of gastrointestinal and cardiovascular systems, and colonisation by exogenous microbes. On occasions, this symbiotic relationship breaks down, and previously minor components of the microbiota outcompete beneficial bacteria, thereby increasing the risk of disease. Antimicrobial agents have been formulated into many oral care products to augment mechanical plaque control. A delicate balance is needed, however, to control the oral microbiota at levels compatible with health, without killing beneficial bacteria and losing the key benefits delivered by these resident microbes. These antimicrobial agents may achieve this by virtue of their recommended twice daily topical use, which results in pharmacokinetic profiles indicating that they are retained in the mouth for relatively long periods at sublethal levels. At these concentrations they are still able to inhibit bacterial traits implicated in disease (e.g. sugar transport/acid production; protease activity) and retard growth without eliminating beneficial species. In silico modelling studies have been performed which support the concept that either reducing the frequency of acid challenge and/or the terminal pH, or by merely slowing bacterial growth, results in maintaining a community of beneficial bacteria under conditions that might otherwise lead to disease (control without killing).

Prospects of oral disease control in the future - an opinion.

The mouth supports a diverse microbiota which provides major benefits to the host. On occasions, this symbiotic relationship breaks down (dysbiosis), and disease can be a consequence. We argue that progress in the control of oral diseases will depend on a paradigm shift away from approaches that have proved successful in medicine for many diseases with a specific microbial aetiology. Factors that drive dysbiosis in the mouth should be identified and, where possible, negated, reduced or removed, while antimicrobial agents delivered by oral care products may function effectively, even at sub-lethal concentrations, by modulating the activity and growth of potentially pathogenic bacteria. In this way, the beneficial activities of the resident oral microbiota will be retained and the risk of dysbiosis occurring will be reduced.

Non-lethal control of the cariogenic potential of an agent-based model for dental plaque.

Dental caries or tooth decay is a prevalent global disease whose causative agent is the oral biofilm known as plaque. According to the ecological plaque hypothesis, this biofilm becomes pathogenic when external challenges drive it towards a state with a high proportion of acid-producing bacteria. Determining which factors control biofilm composition is therefore desirable when developing novel clinical treatments to combat caries, but is also challenging due to the system complexity and the existence of multiple bacterial species performing similar functions. Here we employ agent-based mathematical modelling to simulate a biofilm consisting of two competing, distinct types of bacterial populations, each parameterised by their nutrient uptake and aciduricity, periodically subjected to an acid challenge resulting from the metabolism of dietary carbohydrates. It was found that one population was progressively eliminated from the system to give either a benign or a pathogenic biofilm, with a tipping point between these two fates depending on a multiplicity of factors relating to microbial physiology and biofilm geometry. Parameter sensitivity was quantified by individually varying the model parameters against putative experimental measures, suggesting non-lethal interventions that can favourably modulate biofilm composition. We discuss how the same parameter sensitivity data can be used to guide the design of validation experiments, and argue for the benefits of in silico modelling in providing an additional predictive capability upstream from in vitro experiments.

High-frequency affine mechanics and nonaffine relaxation in a model cytoskeleton.

The cytoskeleton is a network of crosslinked, semiflexible filaments, and it has been suggested that it has properties of a glassy state. Here we employ optical-trap-based microrheology to apply forces to a model cytoskeleton and measure the high-bandwidth response at an anterior point. Simulating the highly nonlinear and anisotropic stress-strain propagation assuming affinity, we found that theoretical predictions for the quasistatic response of semiflexible polymers are only realized at high frequencies inaccessible to conventional rheometers. We give a theoretical basis for determining the frequency when both affinity and quasistaticity are valid, and we discuss with experimental evidence that the relaxations at lower frequencies can be characterized by the experimentally obtained nonaffinity parameter.

Local mechanical response in semiflexible polymer networks subjected to an axisymmetric prestress.

Analytical and numerical calculations are presented for the mechanical response of fiber networks in a state of axisymmetric prestress, in the limit where geometric nonlinearities such as fiber rotation are negligible. This allows us to focus on the anisotropy deriving purely from the nonlinear force-extension curves of individual fibers. The number of independent elastic coefficients for isotropic, axisymmetric, and fully anisotropic networks are enumerated before deriving expressions for the response to a locally applied force that can be tested against, e.g., microrheology experiments. Localized forces can generate anisotropy away from the point of application, so numerical integration of nonlinear continuum equations is employed to determine the stress field, and induced mechanical anisotropy, at points located directly behind and in front of a force monopole. Results are presented for the wormlike chain model in normalized forms, allowing them to be easily mapped to a range of systems. Finally, the relevance of these findings to naturally occurring systems and directions for future investigation are discussed.

Spindles and active vortices in a model of confined filament-motor mixtures.

BACKGROUND: Robust self-organization of subcellular structures is a key principle governing the dynamics and evolution of cellular life. In fission yeast cells undergoing division, the mitotic spindle spontaneously emerges from the interaction of microtubules, motor proteins and the confining cell walls, and asters and vortices have been observed to self-assemble in quasi-two dimensional microtubule-kinesin assays. There is no clear microscopic picture of the role of the active motors driving this pattern formation, and the relevance of continuum modeling to filament-scale structures remains uncertain. RESULTS: Here we present results of numerical simulations of a discrete filament-motor protein model confined to a pressurised cylindrical box. Stable spindles, nematic configurations, asters and high-density semi-asters spontaneously emerge, the latter pair having also been observed in cytosol confined within emulsion droplets. State diagrams are presented delineating each stationary state as the pressure, motor speed and motor density are varied. We further highlight a parameter regime where vortices form exhibiting collective rotation of all filaments, but have a finite life-time before contracting to a semi-aster. Quantifying the distribution of life-times suggests this contraction is a Poisson process. Equivalent systems with fixed volume exhibit persistent vortices with stochastic switching in the direction of rotation, with switching times obeying similar statistics to contraction times in pressurised systems. Furthermore, we show that increasing the detachment rate of motors from filament plus-ends can both destroy vortices and turn some asters into vortices. CONCLUSIONS: We have shown that discrete filament-motor protein models provide new insights into the stationary and dynamical behavior of active gels and subcellular structures, because many phenomena occur on the length-scale of single filaments. Based on our findings, we argue the need for a deeper understanding of the microscopic activities underpinning macroscopic self-organization in active gels and urge further experiments to help bridge these lengths.

Critical scaling and aging in cooling systems near the jamming transition.

We conduct athermal simulations of freely cooling, viscous soft spheres around the jamming transition density varphi(J) and find evidence for a growing length xi(t) that governs relaxation to mechanical equilibrium. xi(t) is manifest in both the velocity correlation function and the spatial correlations in a scalar measure of local force balance which we define. Data for different densities varphi can be collapsed onto two master curves by scaling xi(t) and t by powers of |varphi-varphi(J)|, indicative of critical scaling. Furthermore, particle transport for varphi>varphi(J) exhibits aging and superdiffusion similar to a range of soft matter experiments, suggesting a common origin. Finally, we explain how xi(t) at late times maps onto known behavior away from varphi(J).

Deformation of cross-linked semiflexible polymer networks.

Networks of filamentous proteins play a crucial role in cell mechanics. These cytoskeletal networks, together with various cross-linking and other associated proteins largely determine the (visco)elastic response of cells. In this Letter we study a model system of cross-linked, stiff filaments in order to explore the connection between the microstructure under strain and the macroscopic response of cytoskeletal networks. We find two distinct regimes as a function primarily of cross-link density and filament rigidity: one characterized by affine deformation and one by nonaffine deformation. We characterize the crossover between these two.