Spectral study on the binding of gadolinium ions with apoovotransferrin

Abstract

The binding of Gd 3+ 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 Gd 3+ 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) × 10 4 cm −1 M −1. The apparent binding constants for the complexes of Gd 3+ with apoovotransferrin are log K 1 = 7.61 ± 0.14 and log K 2 = 4.96 ± 0.26. A very large conformational change of apoovotransferrin appears when Gd 3+ is bound to the N-terminal binding site. When Gd 3+ is bound to C-terminal binding site there is less conformational change.