| Abstract: | The addition of polyethylene glycol (PEG) of MW 6000 to solutions of oxy- and deoxyhemoglobins results in an increase in the thermodynamic activity of these proteins. This in turn results, when PEG concentration is high enough, in phase separation into two phases; a protein-rich, PEG-poor phase and a PEG-rich, protein-poor phase. With increasing PEG concentration, the protein-rich amorphous phase becomes metastable and is converted into a well-defined crystalline or polymer phase. The logarithm of protein solubility is a linear function of PEG content up to a protein concentration of 150 g/L because the expression for the activity coefficient can, up to this concentration range, be approximated by a logarithmic function. Curvature appears at higher protein concentrations. Activities obtained by extrapolation from linear portions of the function, showing an unchanged, well-defined crystalline state, yield an activity coefficient for the saturated PEG-free protein solution in agreement with the appropriate values obtained from hard-sphere calculations of excluded volume [Ross, P. D. & Minton, A. P. (1977) J. Mol. Biol. 112 , 437–452]. Solutions containing two hemoglobin species showed cocrystallization of the hemoglobins with a triple point where two crystal forms can be shown to coexist. |
