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.