Arrest transitions in protein solutions – insight from combining scattering, microrheology, and computer simulations

The 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

Bio-, Magneto- and event-stratigraphy across the K-T boundary

Determining the time and the time structure of rare events in geology can be accomplished by applying three different and independent stratigraphic methods: Biostratigraphy, magneto-stratigraphy and event-stratigraphy. The optimal time resolution of the two former methods is about 1000 years, while by means of event-stratigraphy a resolution of approximately one year can be achieved. For biostratigraphy across the Cretaceous-Tertiary (K-T) boundary micro- and nannofossils have been found best suited. The qualitative and quantitative analyses of minerals and trace elements across the K-T boundary show anomalies on a millimeter scale and permit conclusions regarding the time structure of the K-T event itself. The results of the analyses find a most consistent explanation by the assumption of an extraterrestrial impact. The main portion of the material rain from the atmosphere evidently was deposited within a short time. The long-time components consist of the finest portion of the material rain from the atmosphere and the transported and redeposited fall-out

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 only depth and width of the attractive part of the potential are optimized, accounts in a remarkable way for the wavevector dependence of the X-ray scattering structure factor.Comment: 4 pages, 4 figure

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 proteindense 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

A colloid approach to self-assembling antibodies

Concentrated solutions of monoclonal antibodies have attracted considerable attention due to their importance in pharmaceutical formulations, yet their tendency to aggregate and the resulting high solution viscosity has posed considerable problems. It remains a very difficult task to understand and predict the phase behavior and stability of such solutions. Here we present a systematic study of the concentration dependence of the structural and dynamic properties of monoclonal antibodies using a combination of different scattering methods and microrheological experiments. To interpret these data, we use a colloid-inspired approach based on a simple patchy model, which explicitly takes into account the anisotropic shape and the charge distribution of the molecules. Combining theory, simulations and experiments, we are able to disentangle self-assembly and intermolecular interactions and to quantitatively describe the concentration dependence of structural and dynamic quantities such as the osmotic compressibility, the collective diffusion coefficient and the zero shear viscosity over the entire range of investigated concentrations. This simple patchy model not only allows us to consistently describe the thermodynamic and dynamic behavior of mAb solutions, but also provides a robust estimate of the attraction between their binding sites. It will thus be an ideal starting point for future work on antibody formulations, as it provides a quantitative assessment of the effects of additional excipients or chemical modifications on antibody interactions, and a prediction of their effect on solution viscosity

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

Density-dependent interactions and structure of charged colloidal dispersions in the weak screening regime

We determine the structure of charge-stabilized colloidal suspensions at low ionic strength over an extended range of particle volume fractions using a combination of light and small angle neutron scattering experiments. The variation of the structure factor with concentration is analyzed within a one-component model of a colloidal suspension. We show that the observed structural behavior corresponds to a non-monotonic density dependence of the colloid effective charge and the mean interparticle interaction energy. Our findings are corroborated by similar observations from primitive model computer simulations of salt-free colloidal suspensions.Comment: Revised version, accepted to Phys. Rev. Let

Accurate Correction of the "bulk Response" in Surface Plasmon Resonance Sensing Provides New Insights on Interactions Involving Lysozyme and Poly(ethylene glycol)

Surface plasmon resonance is a very well-established surface sensitive technique for label-free analysis of biomolecular interactions, generating thousands of publications each year. An inconvenient effect that complicates interpretation of SPR results is the "bulk response"from molecules in solution, which generate signals without really binding to the surface. Here we present a physical model for determining the bulk response contribution and verify its accuracy. Our method does not require a reference channel or a separate surface region. We show that proper subtraction of the bulk response reveals an interaction between poly(ethylene glycol) brushes and the protein lysozyme at physiological conditions. Importantly, we also show that the bulk response correction method implemented in commercial instruments is not generally accurate. Using our method, the equilibrium affinity between polymer and protein is determined to be KD = 200 μM. One reason for the weak affinity is that the interaction is relatively short-lived (1/koff < 30 s). Furthermore, we show that the bulk response correction also reveals the dynamics of self-interactions between lysozyme molecules on surfaces. Besides providing new insights on important biomolecular interactions, our method can be widely applied to improve the accuracy of SPR data generated by instruments worldwide

Punk::Schwab:Artaud oder Fick zurück was dich zufickt (die Sprache)

Wie steigt man aus im digitalen Zeitalter – aus der Sprache, aus der Gesellschaft, aus dem Ich? Wie lässt sich Widerstand überhaupt noch formulieren? Und wie kann Kunst opponieren angesichts eines gefräßigen, alles einverleibenden Markts? Werner Schwab, Kultfigur eines widerständigen Gestus in der kleinbürgerlich-bäuerlich-klerikal-faschistischen österreichischen Provinz in den späten 1980er, frühen 1990er Jahren, selbst noch analog arbeitend, aber mit neuesten Medientheorien vertraut, nimmt in seinem Schreiben – wie das System selbst – alles auf, amalgamiert, kompiliert, „verwurschtet“ es und spiegelt das Halbverdaute in seiner „nackten Obszönität“ der Gesellschaft wieder zurück, „das alles, um dem ‚System’ seine demaskierte Fratze entgegen zu halten“, zugleich „Zerrspiegel“ und in seiner die „Grenzen des semantischen Bereichs zur reinen Körperlichkeit hin überschreitenden“ Sprachbearbeitung „Subversionsübung“, wie es der frühverstorbene genialisch-grenzgängerische Grazer Journalist Richard Stradner in seinem Schwab-Beitrag „PUNK::SCHWAB:ARTAUD oder Fick zurück was dich zufickt (die Sprache)“ formuliert, der Schwab mit deutlich identifikatorischem Gestus in einen Gegenkanon zwischen Punk (im Gegensatz zum bereits systemkonformen Rock und Pop) und Artauds „Theater der Grausamkeit“ und am Kipppunkt zwischen dem „Authentizitätsanspruch“ einer „Überschreitungsästhetik“ (etwa durch die Fäkalsprache) und der Deutung als „intelligent angelegter Fake“ einreiht. Stradner entwirft auf wenigen Seiten eine Schwab-Poetik in nuce mit vielen Anschlussmöglichkeiten, die deutlich macht, dass Widerstand zu allen Zeiten sich nur über die Form artikulieren lässt und die in ihrer kursorisch-offenen anti-hierarchischen Form dem Schwabschen Schreiben kongenial ist