Binding Strength of Calcium Ion to Bovine α-Lactalbnmin1

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Abstract
A Ca2+-sensitive electrode was used for determination of the binding strength of Ca2+ to bovine α-lactalbumin in 60 mM Tris buffer (pH 7.8–8.5) in the presence of various concentrations of NaCl. The dependence of the apparent binding constant on the concentration of NaCl was consistent with competitive binding of Ca2+ and Na+, and the binding constants of Ca2+ and Na+ were found to be 2.2 (±0.5)× 107M−1 and 99 (±33) M−1, respectively, at 37°C and pH 8.0. The temperature dependence of the binding constant of Ca2+ was examined between 30 and 45°C; extrapolation of the dependence led to a binding constant of ˜1 × 107 M−1 at pH 8.4 and 25°C. The electrostatic contribution and conformational effect of the protein were also taken into consideration, and the intrinsic binding constant of Ca2+ to native a-lactalbumin was calculated to be (1.2–1.5) × 1010 M−1 at 37°C and pH 8.0.