Evidence that MgATP accelerates primary electron transfer in a Clostridiumpasteurianum Fe protein-Azotobacter vinelandii MoFe protein nitrogenase tight complex

The nitrogenase catalytic cycle involves binding of the iron (Fe) protein t o the molybdenum-iron (MoFe) protein, transfer of a single electron from th e Fe protein to the MoFe protein concomitant with the hydrollysis of at lea st two MgATP molecules, followed by dissociation of the two proteins. Earli er studies found that combining the Fe protein isolated from the bacterium Clostridium pasteurianum with the MoFe protein isolated from the bacterium Azotobacter vinelandii resulted in an inactive, nondissociating Fe protein- MoFe protein complex. In the present work, it is demonstrated that primary electron transfer occurs within this nitrogenase tight complex in the absen ce of MgATP (apparent first-order rate constant k = 0.007 s(-1)) and that M gATP accelerates this electron transfer reaction by more than 10,000-fold t o rates comparable to those observed within homologous nitrogenase complexe s (k = 100 s(-1)). Electron transfer reactions were confirmed by EPR spectr oscopy. Finally, the midpoint potentials (E-m) for the Fe protein [4Fe-4S]( 2+/+) cluster and the MoFe protein p(2+/N) cluster were determined for both the uncomplexed and complexed proteins and with or without MgADP. Calculat ions from electron transfer theory indicate that the measured changes in E- m are not likely to be sufficient to account for the observed nucleotide-de pendent rate accelerations for electron transfer.