Experimental Method to Distinguish between a Solution and a Suspension

Dispersion of objects in a fluid phase can be classified as solutions (Gibbs free energy of mixing, Delta G(mix) < 0) or suspensions (Delta G(mix) > 0) depending on their thermodynamic stability. Small objects tend to form solutions, larger ones suspensions, e.g., molecules versus micrometer-sized colloids. Proteins and nanomaterials fall between these two size regimes. The long-standing issue of whether proteins and nanoparticles are dissolved or suspended remains an important research question. Here, a simple, versatile, and experimentally robust method, based on sedimentation equilibrium analytical ultracentrifugation (SE-AUC), which can determine whether proteins, nanoparticles, or polymers form solutions or suspensions, is presented. SE-AUC determines the osmotic pressure profile for a dispersion. Such a profile for solutions (equilibrium one-phase systems) is independent of the initial and the operating conditions. The opposite is true for suspensions that are nonequilibrium two-phase systems. This study proves that bovine serum albumin and lysozyme form solutions while ferritin and apoferritin form suspensions.

Polymer-theory insights into biomolecular systems

Salvatore Assenza

Boosted by the technological advances in experimental techniques, cellular biology is nowadays facing the need for quantitative approaches in order to rationalize the huge amount of collected data. A particularly succesfull theoretical framework is provided by Polymer Theory which, combined with molecular simulations, can capture the essential features of biomacromolecules and describe the cellular processes they participate to. This thesis provides a compendium of works showing the strength of this combination. In a first project, we model the twisting properties of amyloid fibrils by means of a simple coarse-grained approach, based on the competition between elasticity and electrostatic repulsion of nearby portions of the fibrils. The model quantitatively recapitulates the evolution of fibril periodicity as a function of the ionic strength of the solution and of the fibril width. A universal mesoscopic structural signature of the fibrils emerges from this picture, predicting a general, parameter-free law for the periodicity of the fibrils which is validated on several experimental results. A second work is focused on the role played by mitochondrial Hsp70 chaperone in the import of cytoplasmic proteins. Particularly, we computed by means of molecular simulations the effective free-energy profile for substrate translocation upon chaperone binding. We then used the resulting free energy to quantitatively characterize the kinetics of the import process and outline the essential role played by Hsp70 in this context. Finally, in a third project we studied the shape properties of a polymer under tension, a physical condition typically realized both in single-molecule experiments and in vivo. By means of analytical calculations and Monte Carlo simulations, we develop a theoretical framework which quantitatively describes these properties, highlighting the interplay between external force and chain size in determining the spatial distribution of a stretched chain.

EPFL2015

Polymer-theory insights into biomolecular systems

Salvatore Assenza

EPFL2015