Protein—Polyelectrolyte Phase Boundaries

Abstract

The separation of proteins by polyelectrolyte coacervation or precipitation is based on electrostatically-driven complex formation. We have investigated complexation between the globular protein BSA and the polyelectrolyte poly(dimethyldially1ammonium chloride) (PDMDAAC) using light-scattering techniques to monitor solution turbidity. Turbidimetric pH titrations were used to determine the specific pH values where soluble complex formation is initiated (pH,) and where phase separation occurs (pH& These values, collected a t different ionic strengths, can be presented as phase boundaries. The effects of macromolecular concentration, protein:polymer concentration ratio (r), and polymer molecular weight upon the phase boundary are examined. The macromolecular concentration and polymer molecular weight have little or no effect on the phase boundary. While pH, is independent of r, pH$ varies inversely with r. The use of phase boundaries in the selection of optimal pH and ionic strength for maximum yield and purity in protein separations is discussed.

Binding of Polycarboxylic Acids to Cationic Mixed Micelles: Effects of Polymer Counterion Binding and Polyion Charge Distribution

“…3. In the presence of TMAC, we find the same linear dependence of Y c on I 1/ 2 as revealed in previous studies on a number of polyelectrolyte-micelle systems (9,11,53). This linearity suggests that the phenomenon is governed by Eq.…”

“…"Type I" turbidimetric titrations (9,11), corresponding to the addition of ionic surfactant to polymer ϩ nonionic surfactant micelle solutions, were performed at 22 Ϯ 2°C by adding 40 mM cationic surfactant, either CTAB or CTAC, to a mixture of 0.50 g/L polymer and 20 mM C 12 E 8 at a constant ionic strength and pH.…”

“…In numerous studies with polyelectrolytes and oppositely charged micelles, we (7)(8)(9)(10) have obtained a comparable empirical result, namely: c ⅐ ϳ , [1] where c is the colloid critical surface charge density at the point of incipient complex formation, is the structural polymer linear charge density, and is the Debye-Hückel parameter, proportional to I 1/ 2 . This relationship also applies, at least at constant , to the binding of proteins to polyelectrolytes (11) despite the fact that such binding occurs even when the protein net charge is of the same sign as the polymer's.…”

Entering and Exiting the Protein−Polyelectrolyte Coacervate Phase via Nonmonotonic Salt Dependence of Critical Conditions

“…Athough the MW distribution of the chitosan sample has not been determined, the tendency of chitosan to aggregate is known to be highly sensitive to high MW species 50 but polyelectrolyte-protein binding affinity is little affected by MW. 11 The magnitude of coacervate turbidity decreases monotonically with increasing salt and disappears at 200 mM. It is likely that charge neutralization may be reached at some pH for all ionic strengths, but the level and strength of protein-polyelectrolyte interactions may be too weak to lead to counterion release, a likely driving force for coacervation.…”

“…11 Donati et al 14 found destabilization of a chitosan-alginate coacervate at ionic strengths outside of the range 0.02-0.10 M. Most recently Sperber et al 15 observed a maximum in pH φ at I ) 30 mM for mixtures of BLG and high charge density pectin. In general, the universality of such nonmonotonic behavior and its possible correlation with similar effects on pH c is not clear.…”