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Protein Precipitation and Crystalization

Protein-protein interactions are central to many biological and biotechnology processes. We are developing molecular-thermodynamic descriptions of protein phase equilibria in electrolyte solutions, providing a framework for the design and optimization of protein separation systems such as precipitation and crystallization. Salt-induced precipitation is extensively used in industry as a first purification step, and while crystallization provides the purest form of proteins, finding conditions for crystallization remains the limiting and least understood step in X-ray crystallographic determination of protein crystal structure. Protein aggregation results in the loss of biological activity, and misfolding and formation of insoluble deposits are observed in debilitating diseases (e.g., Alzheimer’s, Parkinson’s and Huntington’s diseases), in biotechnology (inclusion body formation) and in the pharmaceutical industry (protein formulation). We combine experimental methods and computer simulations to study protein interactions.

As a first step in determining protein phase behavior, we consider proteins as spheres interacting via a two-body potential that depends on the physicochemical properties of the protein, the electrolyte and properties of the solution such as temperature and pH. Intermolecular interactions are measured using low angle laser light scattering, dynamic light scattering, membrane osmometry, fluorescence anisotropy and cloud point measurements. Chromatography also provides a source of data on specific protein-protein interactions. Integral equation approximations provide the link between potentials of mean force (pmf) and thermodynamic properties to yield the equilibrium phase behavior of the systems.

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