The Effect of the Redox Potential on the Activity of the Nitrogenase and on the Fe‐Protein of Azotobacter vinelandi

Abstract

The redox potential dependence on the nitrogenase reaction catalyzed by different nitrogenase cornplexcs from Azotohacter vinelundii has been studied by two methods. The redox potential was set with the redox couplc SO2−/SO2−3 and the effect on the nitrogenase activity was determined. The oxidation of photochemically reduced low‐potential electron carriers by nitrogenase was followed spectrophotometrically. Simultaneously the nitrogenase activity was estimated by the production of hydrogen. It was found that when component I1 of the nitrogenase is in redox equilibrium with the electron donor, the nitrogenase activity is maximum and independent of the redox potential up to −440 mV. At higher potentials the nitrogenase activity declines and no significant activity was detectable at potentials above −350 mV. The effect of the redox potential on the oxidation reduction state of the 4Fe‐4S cluster of Azotohacter nitrogenase component I1 was studied by electron paramagnetic resonance spectroscopy. Without adenine nucleotides present, component I1 of the nitrogenase shows a midpoint potential of −393 mV and the oxidation reduction reaction is characterized by the transfer of two electrons per redox step. In the presence of MgATP the midpoint potential of component I1 of the nitrogenase undergoes a negative shift of 42 mV. The curve fitting the experimental points is also characteristic of a two‐electron redox step. In contrast, it is found that in the presence of MgADP, the curve fitting to the experimental points is characterized by a one‐electron step with a midpoint potential of −473 mV. It is demonstrated that the nitrogenase activity correlates with the oxidation reduction state of component II of the nitrogenase without adenine nucleotides bound. The implications of this observation, indicating a sequential reaction of the reduced Fe protein with the Mo‐Fe protein, for thc present kinetic models of nitrogenase will be discussed. Copyright © 1982, Wiley Blackwell. All rights reserved