Liquid-liquid phase separation and static light scattering of concentrated ternary mixtures of bovine α and γB crystallins -: art. no. 134909

We have used light scattering, turbidimetry, and thermodynamic analysis to study the phase diagram of concentrated aqueous mixtures of the bovine lens proteins, gamma B crystallin, and alpha crystallin. We find that dilute alpha crystallin raises the phase separation temperature of concentrated gamma B crystallin, while more concentrated alpha crystallin suppresses phase separation. Very concentrated alpha/gamma B mixtures can reversibly cloud above 37 degrees C, even though gamma B alone phase separates only below temperatures near 0 degrees C, and alpha does not phase separate. At the scattering vector magnitude used, high-concentration alpha/gamma B mixtures scatter less light than the weighted average of their component alpha and gamma B solutions, while low-concentration alpha/gamma B mixtures scatter more than such a weighted average. We use a mean-field thermodynamic analysis of such ternary mixtures to show that the observed light scattering and phase boundaries of alpha and gamma B crystallin mixtures give evidence for prominent local fluctuations of relative protein composition. In the single phase, these fluctuations scatter comparatively little light, but are associated with enhanced thermodynamic instability. By applying this analysis to the experimental tie lines we estimate the magnitude of the saddlelike component of the free energy near the aqueous-gamma B critical point. (c) 2006 American Institute of Physics.