Peter Schurtenberger

Equilibrium cluster formation in concentrated protein solutions and colloids.

Controlling interparticle interactions, aggregation and cluster formation is of central importance in a number of areas, ranging from cluster formation in various disease processes to protein crystallography and the production of photonic crystals. Recent developments in the description of the interaction of colloidal particles with short-range attractive potentials have led to interesting findings including metastable liquid–liquid phase separation and the formation of dynamically arrested states (such as the existence of attractive and repulsive glasses, and transient gels). The emerging glass paradigm has been successfully applied to complex soft-matter systems, such as colloid–polymer systems and concentrated protein solutions. However, intriguing problems like the frequent occurrence of cluster phases remain. Here we report small-angle scattering and confocal microscopy investigations of two model systems: protein solutions and colloid–polymer mixtures. We demonstrate that in both systems, a combination of short-range attraction and long-range repulsion results in the formation of small equilibrium clusters. We discuss the relevance of this finding for nucleation processes during protein crystallization, protein or DNA self-assembly and the previously observed formation of cluster and gel phases in colloidal suspensions.

Internal structure and colloidal behaviour of covalent whey protein microgels obtained by heat treatment

Covalently cross-linked whey protein microgels (WPM) were produced without the use of a chemical cross-linking agent. The hierarchical structure of WPM is formed by a complex interplay of heat denaturation, aggregation, electrostatic repulsion, and formation of disulfide bonds. Therefore, well-defined spherical particles with a diameter of several hundreds of nanometers and with relatively low polydispersity are formed in a narrow pH regime (5.8–6.2) only. WPM production was carried out on large scale by heating a protein solution in a plate-plate heat exchanger. Thereafter, the microgels were concentrated by microfiltration and spray dried into a powder. The spherical structure of the WPM was conserved in the powder. After re-dispersion, the microgel dispersions fully recovered their initial structure and size distribution. Due to the formation of disulfide bonds the particles were internally covalently cross-linked and were remarkably stable in a large pH range. Because of the pH dependent charge of the constituents the particles underwent significant size changes upon shifting the pH. Small angle X-ray scattering experiments were used to reveal their internal structure, and we report on the pH-induced structural changes occurring on different length scale. Our experiments showed that close analogies could be drawn to internally cross-linked and pH-responsive microgels based on weak polyelectrolytes. WPM also exhibited a pronounced swelling at pH values below the isoelectric point (IEP), and a collapse at the IEP. However, in contrast to classical microgels, WPM are not build up by simple polymer chains but possess a complex hierarchical structure consisting of strands formed by clusters of aggregated denatured proteins that act as primary building blocks. They were flexible enough to respond to changes of the environment, and were stable enough to tolerate pH values where the proteins were highly charged and the strands were stretched.

Interplay between Spinodal Decomposition and Glass Formation in Proteins Exhibiting Short-Range Attractions

We investigate the competition between spinodal decomposition and dynamical arrest using aqueous solutions of the globular protein lysozyme as a model system for colloids with short-range attractions. We show that quenches below a temperature Ta lead to gel formation as a result of a local arrest of the protein-dense phase during spinodal decomposition. The rheological properties of these gels allow us to use centrifugation experiments to determine the local densities of both phases and to precisely locate the gel boundary and the attractive glass line close to and within the unstable region of the phase diagram.

Depolarization of backscattered linearly polarized light

We formulate a quantitative description of backscattered linearly polarized light with an extended photon diffusion formalism taking explicitly into account the scattering anisotropy parameter g of the medium. From diffusing wave spectroscopy measurements, the characteristic depolarization length for linearly polarized light, lp , is deduced. We investigate the dependence of this length on the scattering anisotropy parameter g spanning an extended range from -1 (backscattering) to 1 (forward scattering). Good agreement is found with Monte Carlo simulations of multiply scattered light.

The Largest Synthetic Structure with Molecular Precision: Towards a Molecular Object

Pushing the limits: A 200A - 10 Da structurally defined, linear macromolecule (PG5) has a molar mass, cross-section dimension, and cylindrical shape that are comparable to some naturally occurring objects, such as amyloid fibrils or certain plant viruses. The macromolecule is resistant against flattening out on a surface; the picture shows PG5 embracing the tobacco mosaic virus (TMV).

Photoinitiated Coupling of Unmodified Monosaccharides to Iron Oxide Nanoparticles for Sensing Proteins and Bacteria

We report a versatile approach for the immobilization of unmodified monosaccharides onto iron oxide nanoparticles. Covalent coupling of the carbohydrate onto iron oxide nanoparticle surfaces was accomplished by the CH insertion reaction of photochemically activated phosphate-functionalized perfluorophenylazides (PFPAs), and the resulting glyconanoparticles were characterized by IR, TGA, and TEM. The surface-bound d-mannose showed the recognition ability toward Concanavalin A and Escherichia coli strain ORN178 that possesses mannose-specific receptor sites. Owing to the simplicity and versatility of the technique, together with the magnetic property of iron oxide nanoparticles, the methodology developed in this study serves as a general approach for the preparation of magnetic glyconanoparticles to be used in clinical diagnosis, sensing, and decontamination.

Modeling equilibrium clusters in lysozyme solutions

We 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.

A simple patchy colloid model for the phase behavior of lysozyme dispersions

We propose a minimal model for spherical proteins with aeolotopic pair interactions to describe the equilibrium phase behavior of lysozyme. The repulsive screened Coulomb interactions between the particles are taken into account assuming that the net charges are smeared out homogeneously over the spherical protein surfaces. We incorporate attractive surface patches, with the interactions between patches on different spheres modeled by an attractive Yukawa potential. The parameters entering the attractive Yukawa potential part are determined using information on the experimentally accessed gas-liquid-like critical point. The Helmholtz free energy of the fluid and solid phases is calculated using second-order thermodynamic perturbation theory. Our predictions for the solubility curve are in fair agreement with experimental data. In addition, we present new experimental data for the gas-liquid coexistence curves at various salt concentrations and compare these with our model calculations. In agreement with earlier findings, we observe that the strength and the range of the attractive potential part only weakly depend on the salt content.

Cluster-Driven Dynamical Arrest in Concentrated Lysozyme Solutions

We 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 ≳ 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.

Mode-selective dynamic light scattering: theory versus experimental realization

We present a quantitative experimental comparison of fiber-based, single- and few-mode dynamic light scattering with the classical pinhole-detection optics. The recently presented theory of mode-selective dynamic light scattering [Appl. Opt. 32, 2860 (1993)] predicts a collection efficiency and a signal-tobaseline ratio superior to that of a classical pinhole setup. These predictions are confirmed by our experiments. Using single-mode optical fibers with different cutoff wavelengths and commercially available mechanical components, we have constructed a mode-selective detection optics in a simple and compact dynamic light-scattering spectrometer that permits an optimal compromise between signal intensity and dynamical resolution.