PROTEIN PARTITIONING IN 2-PHASE AQUEOUS POLYMER SYSTEMS .1. NOVEL PHYSICAL PICTURES AND A SCALING-THERMODYNAMIC FORMULATION

Novel physical pictures are proposed for the interactions between globular proteins and flexible nonionic polymers that are responsible for the observed partitioning of proteins in two-phase aqueous polymer systems. For the system poly(ethylene oxide) (PEO)-dextran-water these novel physical pictures are based on the observation that a transition occurs in the nature of the top PEO-rich phase, from the dilute to the semidilute polymer solution regimes, with increasing PEO molecular weight. In systems containing low molecular weight PEO (M < 10 000 Da), individual polymer coils, which may be larger or smaller than the proteins, interact with the proteins. Mathematically simple geometric and scaling arguments are used to probe the qualitative form of the free-energy change arising from polymer-protein and polymer-polymer interactions within each picture. Through a statistical thermodynamic framework, used to relate the change in free energy to the experimentally measurable protein partition coefficient, K(p), it is possible to discriminate between the novel physical pictures on the basis of the predicted qualitative trends. At THETA-solvent conditions for the polymer, a picture that incorporates solely the excluded-volume interactions between the proteins and the polymer coils is unable to account for the influence of polymer molecular weight on K(p); at athermal-solvent conditions, where repulsive polymer-polymer interactions also influence the protein chemical potential, the predicted protein partition coefficient is shifted in a direction qualitatively consistent with experimental trends. For both athermal- and THETA-solvent conditions, the observed change in K(p) is found to be qualitatively consistent with the presence of a weak attraction between the polymer coils and the proteins. The presence of a strong attraction between the polymer coils and the proteins, and associated formation of an adsorbed polymer layer at the surface of the proteins, can lead to a new partitioning behavior that has not yet been realized experimentally. In systems containing high molecular weight PEO (M >> 10 000 Da), the proteins interact with an entangled polymer network rather than with identifiable polymer coils, and the protein partition coefficient becomes independent of the PEO molecular weight.