SOLID LIQUID-PHASE BOUNDARIES OF LENS PROTEIN SOLUTIONS

We report measurement of the solid-liquid phase boundary, or liquidus line, for aqueous solutions of three pure calf gamma-crystallin proteins: gamma-II, gamma-IIIa, and gamma-IIIb. We also studied the liquidus line for solutions of native gamma-IV-crystallin calf lens protein, which consists of 85% gamma-IVa/15% gamma-IVb. In all four proteins the liquidus phase boundaries lie higher in temperature than the previously determined liquid-liquid coexistence curves. Thus, over the range of concentration and temperature for which liquid-liquid phase separation occurs, the coexistence of a protein crystal phase with a protein liquid solution phase is thermodynamically stable relative to the metastable separated liquid phases. The location of the liquidus lines clearly divides these four crystallin proteins into two groups: those in which liquidus lines flatten at temperatures > 70-degrees-C: gamma-IIIa and gamma-IV, and those in which liquidus lines flatten at temperatures < 50-degrees-C: gamma-II and gamma-IIIb. We have analyzed the form of the liquidus lines by using specific choices for the structures of the Gibbs free energy in solution and solid phases. By applying the thermodynamic conditions for equilibrium between the two phases to the resulting chemical potentials, we can estimate the temperature-dependent free energy change upon binding of protein and water into the solid phase.