NifS-directed assembly of a transient [2Fe-2S] cluster within the NifU protein

The Nifs and NifU proteins from Azotobacter vinelandii are required for the full activation of nitrogenase. Nifs is a homodimeric cysteine desulfurase that supplies the inorganic sulfide necessary for formation of the Fe-S cl usters contained with in the nitrogenase component proteins, NifU has been suggested to complement NifS either by mobilizing the Fe necessary for nitr ogenase Fe-S cluster formation or by providing an intermediate Fe-S cluster assembly site. As isolated, the homodimeric NifU protein contains one [2Fe -2S](2+,+) cluster per subunit, which is referred to as the permanent clust er. In this report, we show that NifU is able to interact with Nifs and tha t a second, transient [2Fe-2S] cluster can be assembled within NifU in vitr o when incubated in the presence of ferric ion, L-cysteine, and catalytic a mounts of Nifs. Approximately one transient [2Fe-2S] cluster is assembled p er homodimer, The transient [2Fe-2S] cluster species is labile and rapidly released on reduction. We propose that transient [2Fe-2S] cluster units are formed on NifU and then released to supply the inorganic iron and sulfur n ecessary for maturation of the nitrogenase component proteins. The role of the permanent [2Fe-2S] clusters contained within NifU is not yet known, but they could have a redox function involving either the formation or release of transient [2Fe-2S] cluster units assembled on NifU, Because homologs to both NifU and Nifs, respectively designated IscU and IscS, are found in no n-nitrogen fixing organisms, it is possible that the function of NifU propo sed here could represent a general mechanism for the maturation of Fe-s clu ster-containing proteins.