Multivalent ions for tuning the phase behaviour of protein solutions

Orateur : Olga MATSARKAIA (Institut fuer Angewandte Physik, Universitaet Tuebingen, Auf der Morgenstelle 10, 72076 Tuebingen, Germany)

Protein phase behaviour is of importance in various areas of research such as structural biology, rational drug design and delivery, medicine (in particular protein condensation diseases), biotechnology, food science and cell biology. A particularly intriguing variety of phase behaviours can be induced in negatively charged, globular proteins in the presence of multivalent salts such as lanthanide (Ln) chlorides. These behaviours include reentrant condensation, crystallisation and cluster formation as well as liquid-liquid phase separation (LLPS) with an upper or a lower critical solution temperature (UCST-LLPS and LCST-LLPS, respectively). Here, we focus on a characterisation of LCST-LLPS in systems consisting of the protein bovine serum albumin (BSA) and the trivalent salts YCl3, HoCl3 and LaCl3. LCST-LLPS in BSA-YCl3 systems is investigated and shown to be an endothermic, hydration entropy-driven condensation. Subsequently, the influence of different cations and on LCST-LLPS is studied systematically employing small-angle scattering (SAS). Both cation-protein affinity and the strength of cation-induced interprotein attraction can be classified as Ho3+ > Y3+ > La3+. Cations and their properties are thus demonstrated to be a sensitive tool to fine-tune BSA-BSA interactions in solution. This knowledge is then exploited to investigate the kinetics of LLPS of BSA in the presence of different HoCl3/LaCl3 mixtures which induce different effective BSA-BSA attraction strengths. To this end, the characteristic length scales of the respective systems are monitored as functions of time and temperature using ultra-SAS. It is found that a stronger interprotein attraction brought across by higher HoCl3 concentrations leads to an increasing deviation from the t1/3 growth law. Finally, the local environment of Y3+ cation coordination by BSA is investigated as a probe of the molecular mechanism behind LCST-LLPS. The coordination number (CN) of Y3+ is found to be higher at low protein concentration, indicating a pronounced degree of cation hydration. The results obtained indicate that a careful choice of the multivalent cation used can fine-tune protein interactions and their phase behaviour in solution. These findings are of strong interest for a fundamental.

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