Fully Active Fe‐Protein of the Nitrogenase from Azotobacter vinelandii Contains at least Eight Iron Atoms and Eight Sulphide Atoms per Molecule

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Abstract
The Fe-protein of the A. vinelandii nitrogenase enzyme complex contians a variable Fe and sulfide content. The Fe and sulfide content of the protein is dependent upon the specific activity. Up to a specific activity of 1000 nmol C2H4 produced .cntdot. min-1 .cntdot. mg Av2-1, 3 Fe and 3 sulfide atoms/molecule Av2 are found. At specific activities above 1000 nmol C2H4 produced .cntdot. min-1 .cntdot. mg Av2-1, a linear relationship between specific activity and Fe and sulfide content of Av2 is found. The maximum values found are 8.8 Fe atoms and 8.6 sulfide atoms/molecule at a specific activity of 2250 nmol C2H4 produced .cntdot. min-1 .cntdot. mg Av2-1. The experimental molar absorption coefficients at 430 nm of the oxidized and reduced forms depend on the specific activity. The highest values found are 15.9 and 9.1 mM-1 cm-1, respectively. Since occasionally the preparations with specific activities around 3000 nmol .cntdot. min-1 .cntdot. mg-1 are isolated which contain more than 10 Fe atoms and 11 sulfide atoms /molecule, it cannot be excluded that under certain physiological conditions Av2 contains even more than 2 [4 Fe-4S] clusters. The addition of MgATP induces a conformational change in the Fe-protein which results in a higher reactivity with Fe chelators. Irrespective of the specific activity, the amount of Fe extracted from the protein after addition of MgATP never exceeds 4 atoms/molecule. The results are discussed with respect to the present molecular model of the Fe-protein.