Calcium binding by alpha-lactalbumin in human milk and bovine milk.

The metal-binding property of alpha-lactalbumin (alpha-LA) in human milk was studied and compared to that of bovine milk alpha-LA. Gel filtration on Sephadex G-75 at physiological pH and ionic strength separated alpha-LA in human and bovine milk from most other proteins. The only metal ion associated with alpha-LA under these conditions was Ca2+. Minor protein contaminants were removed by ion-exchange chromatography, and the ratio of Ca2+:alpha-LA was determined in the isolated protein preparations. Concentrations of alpha-LA in mature human milk were between 1.03 and 1.57 mg/ml; the Ca2+ concentration bound to alpha-LA varied, yielding a molar ratio of Ca2+:alpha-LA of approximately 1:1 (0.82-1.41 mol Ca2+/mol alpha-LA) in mature milk. Gel filtration with excess Ca2+ in the running buffer showed that there is another weaker binding site for Ca2+, but this binding does not occur under physiological conditions. Less Ca2+ was bound to bovine alpha-LA (0.6-0.9 mol Ca2+/mol alpha-LA) than to human alpha-LA. Calcium binding was abolished at pH 3.0 and resulted in a substantial increase in the hydrodynamic radius of alpha-LA. Reconstitution of human alpha-LA with Ca2+ and other divalent cations at native pH (6.8) and ionic strength showed a binding specific for Ca2+. Since only 1% of calcium from human milk and 0.15% from bovine milk is alpha-LA bound, alpha-LA is probably unimportant with respect to calcium nutrition of the infant. However, the metal binding of alpha-LA may have a biological significance through its role in the lactose synthase complex.