Reduction potential and thermodynamic parameters of adrenodoxin by the use of an anaerobic thin-layer electrode

An enzymatic system to remove dissolved O2 in solution was applied to an optically transparent thin-layer electrode cell. This combination enabled the easy establishment of anaerobic conditions for the spectroelectrochemical study of O2-sensitive oxidation--reduction enzymes. By the use of the anaerobic cell with a small volume of 80 microliter, the formal reduction potential (E0) of adrenodoxin was found to be -250 +/- 2 mV vs a normal hydrogen electrode at 25 degrees C in a solution containing 0.1 M K-phosphate (pH 7.4) and 0.1 M KCl. Temperature-dependent experiments (5-25 degrees C) yielded the following thermodynamic parameters for the electron-transfer reaction of adrenodoxin: enthalpy (delta H0), -13.6 +/- 0.7 kcal/mol; entropy (S0red-S0ox), -49.6 +/- 2.4 eu. The large negative entropy value is quite unique among iron-sulfur proteins and heme proteins, suggesting that the protein structure changes dramatically upon one-electron reduction.