Casein Micelle Substructure and Calcium Phosphate Interactions Studied by Sephacryl Column Chromatography

Abstract

Experiments were designed to manipulate only one of the factors known to be important in maintaining the integrity of casein micelles while holding all of the other factors constant. Casein micelles were dissociated either by reducing the content of colloidal calcium phosphate or by adding κ-casein at a constant free calcium ion concentration, pH, and ionic strength and, in the latter case, also at a constant concentration of colloidal calcium phosphate. Size fractionation of casein micelles and partially dissociated casein micelles was achieved by gel filtration chromatography using an eluent that had an ionic strength and concentrations of free calcium and magnesium ions comparable with those of milk and was saturated with respect to colloidal calcium phosphate. Dissociation by selective reduction of colloidal calcium phosphate produced particles the average size of which changed during dialysis, but at no time was there an accumulation of the putative submicelles. Similarly, the addition of κ-casein, even in great excess, did not completely reverse the suggested process of association by submicelles. Surprisingly, in an attempt to speed the dissociation of casein micelles by urea addition, it was observed that the micelles could form a temperature-dependant precipitate at 3.5 to 4.0 M. None of the submicellar models of casein micelle structure adequately describe these controlled dissociation experiments.