Investigation on the binding of TNS to centrin, an EF-hand protein

The interaction between 2-p-toluidinylnaphthalene-6-sulfonate (TNS) and ciliate Etiplotes Octocarinatits centrin (Cen) has been studied by fluorescence spectroscopy. The binding constants of TNS with Cen were measured at different temperature in the 0.01 M Hepes, pH 7.4. The binding process is exothermic and involves a positive entropy change. The negative value of enthalpy predominately contributes to the negative free energy of binding between TNS and Cen. The salt (KCl) increases the association constant of TNS and Cen. These results and resonance light scattering experiment suggest that the binding force between TNS and Cen is hydrophobic. The distance (r) between TNS and tryptophan of mutant G115W, which sheds more insight into the binding of TNS to Cen, was determined as 4.85 nm based on Forster non-radiative energy transfer theory. (C) 2007 Elsevier B.V. All rights reserved.