Spectral study on the binding of gadolinium ions with apoovotransferrin

The binding of Gd3+ ion to apoovotransferrin (apoOTf) was monitored by means of UV difference spectra in 0.01 M Hepes, pH 7.4 at 25 °C. Used 2-p-toluidinylnaphthalene-6-sulfonate (TNS) as fluorescence probe the conformational changes of protein were studied while gadolinium ions bound to apoOTf. The results show that Gd3+ binding produces peaks at 244 and 294 nm that is the characteristic of binding at the apoOTf specific metal-binding sites. At 244 nm the molar absorptivity of Gd-apoOTf complex is (1.99 ± 0.17) × 104 cm−1 M−1. The apparent binding constants for the complexes of Gd3+ with apoovotransferrin are log K1 = 7.61 ± 0.14 and log K2 = 4.96 ± 0.26. A very large conformational change of apoovotransferrin appears when Gd3+ is bound to the N-terminal binding site. When Gd3+ is bound to C-terminal binding site there is less conformational change.