title: NifS-directed assembly of a transient [2Fe-2S] cluster within the NifU protein.
abstract: 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 clusters contained within the nitrogenase component proteins. NifU has been suggested to complement NifS either by mobilizing the Fe necessary for nitrogenase 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 cluster. In this report, we show that NifU is able to interact with NifS and that a second, transient [2Fe-2S] cluster can be assembled within NifU in vitro when incubated in the presence of ferric ion, L-cysteine, and catalytic amounts of NifS. Approximately one transient [2Fe-2S] cluster is assembled per 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 necessary 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 non-nitrogen fixing organisms, it is possible that the function of NifU proposed here could represent a general mechanism for the maturation of Fe-S cluster-containing proteins.
authors.affiliation: Department of Biochemistry, Virginia Tech, Blacksburg, VA 24061-0346, USA.
authors.forename: P
authors.initials: P
authors.lastname: Yuvaniyama
authors.forename: J N
authors.initials: JN
authors.lastname: Agar
authors.forename: V L
authors.initials: VL
authors.lastname: Cash
authors.forename: M K
authors.initials: MK
authors.lastname: Johnson
authors.forename: D R
authors.initials: DR
authors.lastname: Dean
mesh_terms: D019943:Amino Acid Substitution; D016948:Azotobacter vinelandii / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D007501:Iron / Q000737:chemistry; D007506:Iron-Sulfur Proteins / Q000737:chemistry*; D046911:Macromolecular Substances; D010084:Oxidation-Reduction; D013056:Spectrophotometry, Ultraviolet; D013059:Spectrum Analysis, Raman
chemical_list: D001426:Bacterial Proteins; D007506:Iron-Sulfur Proteins; D046911:Macromolecular Substances; C088637:NifU protein, Bacteria; C080001:nifS protein, Bacteria; D007501:Iron