102 research outputs found
Arrest transitions in protein solutions – insight from combining scattering, microrheology, and computer simulations
No full textThe static and dynamic properties of concentrated protein solutions are essential ingredients for our understanding of the cellular machinery or formulating biopharmaceuticals. Here a combination of advanced characterization techniques such as light and x-ray scattering, neutron spin echo measurements [1] and microrheology experiments [2], combined with the theoretical toolbox from colloid physics and state-of-the-art computer simulations [3], considerably enhances our understanding of the link between protein interactions and the stability, dynamics and flow properties of these solutions up to high concentrations. We will address the enormous influence of weak attractive interactions known to exist between many globular proteins, and demonstrate the dramatic effect of an interaction potential anisotropy [1] such as attractive patches and shape anisotropy [3] on the dynamic properties. We will also discuss how we can combine interparticle interaction effects and the formation of (transient) equilibrium clusters in an attempt to understand and predict properties such as the concentration dependence of the zero shear viscosity of dense protein solutions [4].
(1) Bucciarelli, S.; Myung, J. S.; Farago, B.; Das, S., Vliegenthart, G.; Holderer, O.; Winkler, R. G.; Schurtenberger, P.; Gompper, G.; Stradner, A. “Dramatic influence of patchy attractions on short-time protein diffusion under crowded conditions” Sci. Adv. 2016, 2:e1601432.
(2) Garting, T. and Stradner, A. “Optical Microrheology of Protein Solutions using Tailored Nanoparticles” Small 2018, 1801548.
(3) Myung, J. S.; Roosen-Runge, F.; Winkler, R. G.; Gompper, G.; Schurtenberger, P.; Stradner, A. “Weak shape anisotropy leads to non-monotonic crowding effects impacting protein dynamics under physiologically relevant conditions” J. Phys. Chem. B 2018, 122, 12396-12402.
(4) Bergman, M.; Garting, T.; Schurtenberger, P.; Stradner, A. “Experimental Evidence for a Cluster Glass Transition in Concentrated Lysozyme Solutions” submitted to J. Phys. Chem. B, 2019
Modeling equilibrium clusters in lysozyme solutions
No full textWe present a combined experimental and numerical study of the equilibrium cluster formation in globular-protein solutions under no-added salt conditions. We show that a cluster phase emerges as a result of a competition between a long-range screened Coulomb repulsion and a short-range attraction. A simple effective potential, in which electrostatic repulsion is fixed by experimental conditions and attraction is modeled with a generalized Lennard-Jones potential, accounts in a remarkable way for the wavevector dependence of the X-ray scattering structure factor
Self-assembly in patchy proteins: From transient networks to attractive glasses
No full textDynamic properties of crowded protein solutions are difficult to predict and control. This for example considerably limits our ability to create stable and injectable formulations of proteins or peptides at high concentrations. Another physiologically relevant case is presbyopia, or age-related farsightedness, where the pathological stiffening of the eye lens can be related to a liquid-solid transition of the protein mixtures inside the eye lens cells1. It is thus essential to achieve a quantitative understanding of the link between the molecular structure of the proteins and the interactions between them, and how these interactions influence the stability, dynamics and flow properties of the solutions as a function of their concentration. Here we show how we can use a combination of advanced characterization techniques1-4 such as neutron spin echo, small-angle scattering, 3D cross correlation light scattering and microrheology, combined with state-of-the-art computer simulations to assess and predict interparticle interactions and their impact on the dynamics and flow behavior of crowded protein solutions. We particularly point out the enormous influence of weak attractive interactions known to exist between many globular proteins, and demonstrate the dramatic effect of an interaction potential anisotropy such as attractive patches4 and shape anisotropy on the dynamic properties.
[1] G. Foffi, G. Savin, S. Bucciarelli, N. Dorsaz, G. Thurston, A. Stradner, P. Schurtenberger; A Hard Sphere-Like Glass Transition in Eye Lens Alpha Crystallin Solutions ; Proc. Natl. Acad. Sci. U. S. A., 111, 16748-16753 (2014).
[2] F. Cardinaux, E. Zaccarelli, A. Stradner, S. Bucciarelli, B. Farago, S. Egelhaaf, F. Sciortino, P. Schurtenberger; Cluster-driven dynamical arrest in concentrated lysozyme solutions J. Phys. Chem. B, 115, 7227 (2011).
[3] S. Bucciarelli, L. Casal-Dujat, C. De Michele, F. Sciortino, J. Dhont, J. Bergenholtz, B. Farago, P. Schurtenberger, and A. Stradner; Unusual Dynamics of Concentration Fluctuations in Solutions of Weakly Attractive Globular Proteins ; The Journal of Physical Chemistry Letters, 6, 4470-4474 (2015).
[4] S. Bucciarelli, J. S. Myung, B. Farago, S. Das, G. A. Viegenthart, O. Holderer, R. G. Winkler, P. Schurtenberger, G. Gompper, and A. Stradner; Dramatic Influence of Attractions on Short-Time Protein Diffusion under Crowded Conditions ; Science Advances, 2, e1601432 (2016)
Cluster-Driven dynamical arrest in concentrated lysozyme solutions
No full textWe present a detailed experimental and numerical study of the structural and dynamical properties of salt-free lysozyme solutions. In particular, by combining small-angle X-ray scattering (SAXS) data with neutron spin echo (NSE) and rheology experiments, we are able to identify that an arrest transition takes place at intermediate densities, driven by the slowing down of the cluster motion. Using an effective pair potential among proteins, based on the combination of short-range attraction and long-range repulsion, we account remarkably well for the peculiar volume fraction dependence of the effective structure factor measured by SAXS. We show that a transition from a monomer to a cluster-dominated fluid happens at volume fractions larger than ϕ ***Missing image substitution*** 0.05 where the close agreement between NSE measurements and Brownian dynamics simulations confirms the transient nature of the clusters. Clusters even stay transient above the geometric percolation found in simulation at ϕ > 0.15, though NSE reveals a cluster lifetime that becomes increasingly large and indicates a divergence of the diffusivity at ϕ ≃ 0.26. Macroscopic measurements of the viscosity confirm this transition where the long-lived-nature of the clusters is at the origin of the simultaneous dynamical arrest at all length scales
Unusual Dynamics of Concentration Fluctuations in Solutions of Weakly Attractive Globular Proteins
No full textThe globular protein γB-crystallin exhibits a complex phase behavior, where liquid–liquid phase separation characterized by a critical volume fraction ϕc = 0.154 and a critical temperature Tc = 291.8 K coexists with dynamical arrest on all length scales at volume fractions around ϕ ≈ 0.3–0.35, and an arrest line that extends well into the unstable region below the spinodal. However, although the static properties such as the osmotic compressibility and the static correlation length are in quantitative agreement with predictions for binary liquid mixtures, this is not the case for the dynamics of concentration fluctuations described by the dynamic structure factor S(q,t). Using a combination of dynamic light scattering and neutron spin echo measurements, we demonstrate that the competition between critical slowing down and dynamical arrest results in a much more complex wave vector dependence of S(q,t) than previously anticipated
Making Food Protein Gels via an Arrested Spinodal Decomposition
No full textWe report an investigation of the structural and dynamic properties of mixtures of food colloid casein micelles and low molecular weight poly(ethylene oxide). A combination of visual observations, confocal laser scanning microscopy, diffusing wave spectroscopy, and oscillatory shear rheometry is used to characterize the state diagram of the mixtures and describe the structural and dynamic properties of the resulting fluid and solid-like structures. We demonstrate the formation of gel-like structures through an arrested spinodal decomposition mechanism. We discuss our observations in view of previous experimental and theoretical studies with synthetic and food colloids, and comment on the potential of such a route toward gels for food processing
Optical Microrheology of Protein Solutions Using Tailored Nanoparticles
No full textThis work represents a critical re-examination of the application of dynamic light scattering (DLS)–based tracer particle microrheology to measure the zero shear viscosity of aqueous solutions of different proteins up to very high concentrations. It is demonstrated that a combination of surface-functionalized tracer particles, the use of the so-called 3D-DLS technique, and carefully chosen parameters for the scattering experiments is essential for a reliable and artifact-free determination of the viscosity of highly diverse protein solutions, while keeping the amount of protein to a minimum. The major challenges that arise in such microrheology experiments with protein solutions are discussed and used as guiding principles for the synthesis of all-purpose tracer particles with optimal size and an efficient surface functionalization, and the choice of the appropriate amount of tracers in the sample. Potential problems arising from depletion attractions between the tracer particles induced by the proteins are addressed, and compelling evidences for the absence of such effects are presented. The validity of the approach is corroborated by the perfect agreement between the zero shear viscosity obtained from 3D-DLS-based microrheology and literature data from classical rheological measurements for two vastly different protein–solvent systems up to concentrations close to the arrest transition
Synthesis and application of PEGylated tracer particles for measuring protein solution viscosities using Dynamic Light Scattering-based microrheology
No full textThe measurement of flow properties, such as the zero shear viscosity, of protein solutions is of paramount importance for many applications such as pharmaceutical formulations, where the syringeability of physiologically effective doses is a key property. However, the determination of these properties with classical rheological methods is often challenging due to e.g. detrimental surface effects or simply the lack of sufficient material. A possible alternative is Dynamic Light Scattering-based microrheology, where the Brownian motion of tracer particles embedded in the protein solution is monitored to access the zero shear viscosity of the sample. The prime advantages of this method compared to classical rheology are the absence of disturbing surface effects and the up to two orders of magnitude smaller protein quantities needed for an entire concentration series. This Protocol provides a detailed description of the synthesis of sterically stabilized tracer particles with surface and overall particle properties specifically designed to investigate the viscosity of protein solutions up to concentrations close to the arrest transition. These particles are tailored to avoid protein-particle as well as particle-particle aggregation at various sample conditions and thus allow for an artifact-free application of Dynamic Light Scattering-based tracer microrheology to determine the flow behaviour of biological samples. The Protocol concludes with step by step instructions for the characterization of protein solutions using a combination of the tracer particles and an advanced dynamic light scattering technique yielding the concentration-dependent zero shear viscosity
Potential and limits of a colloid approach to protein solutions
No full textLooking at globular proteins with the eyes of a colloid scientist has a long tradition, in fact a significant part of the early colloid literature was focused on protein solutions. However, it has also been recognized that proteins are much more complex than the typical hard sphere-like synthetic model colloids. Proteins are not perfect spheres, their interaction potentials are in general not isotropic, and using theories developed for such particles are thus clearly inadequate in many cases. In this perspective article, we now take a closer look at the field. In particular, we reflect on the fact that modern colloid science has been undergoing a tremendous development, where a multitude of novel systems have been developed in the lab and in silico. During the last decade we have seen a rapidly increasing number of reports on the synthesis of anisotropic, patchy and/or responsive synthetic colloids, that start to resemble their complex biological counterparts. This experimental development is also reflected in a corresponding theoretical and simulation effort. The experimental and theoretical toolbox of colloid science has thus rapidly expanded, and there is obviously an enormous potential for an application of these new concepts to protein solutions, which has already been realized and harvested in recent years. In this perspective article we make an attempt to critically discuss the exploitation of colloid science concepts to better understand protein solutions. We not only consider classical applications such as the attempt to understand and predict solution stability and phase behaviour, but also discuss new challenges related to the dynamics, flow behaviour and liquid-solid transitions found in concentrated or crowded protein solutions. It not only aims to provide an overview on the progress in experimental and theoretical (bio)colloid science, but also discusses current shortcomings in our ability to correctly reproduce and predict the structural and dynamic properties of protein solutions based on such a colloid approach
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