Nitrogenase: the reaction between the Fe protein and bathophenanthrolinedisulfonate as a probe for interactions with MgATP.

The reaction between the Fe(II) chelating agent, bathophenanthrolinedisulfonate, and the iron-sulfur cluster in the Fe protein of nitrogenase from Clostridium pasteurianum has been studied. This reaction is greatly accelerated by the presence of MgATP. Analysis of the relationship between reaction rate and concentration of MgATP supports a model in which both of two binding sites for MgATP on the Fe protein must be occupied before the protein undergoes a conformational change, allowing the iron-sulfur site to react rapidly with chelator. This model is also consistent with presently available data on equilibrium binding of MgATP to the Fe protein. MgADP inhibits the effect of MgATP on the chelator reaction in a manner which suggests that MgADP binds strongly to one of the MgATP sites and more weakly to the other. Loss of enzymic activity due to exposure to O2 or 0 degrees C is accompanied by a decrease in the ATP-specific chelator reaction. Hence, this reaction was used to estimate the concentration of active iron-sulfur centers for the purpose of computing the extinction coefficient of the Fe protein, giving the value delta epsilon 430nm(ox-red) = 6600 M-1 cm-1.