Products of the iron-molybdenum cofactor-specific biosynthetic genes, nifE and nifN, are structurally homologous to the products of the nitrogenase molybdenum-iron protein genes, nifD and nifK

Brigle, Kevin E.; Weiss, Mary C.; Newton, William E.; Dean, Dennis R.

doi:10.1128/jb.169.4.1547-1553.1987

Cited by 168 publications

(112 citation statements)

References 24 publications

(12 reference statements)

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“…Homology modeling and sequence alignments suggest that this type of cluster would be easily accommodated in the NifEN polypeptide environment, which is predicted to be very open with less-bulky amino acids than those found in the MoFe protein environment (29). The lone cysteine residue in this region of NifE, Cys-E250, is homologous to Cys-␣275 in the MoFe protein primary sequence and is therefore believed to serve as the ligand to the terminal iron atom of the NifEN-bound precursor in a manner similar to that observed between Cys-␣275 and FeMoco in the MoFe protein (20,29).…”

Section: Resultsmentioning

confidence: 99%

“…NifEN is a heterotetramer that shares significant sequence homology with the MoFe protein, including several of the residues surrounding both the P-cluster and FeMoco binding sites (20). Two different types of ironsulfur clusters, neither of which contain molybdenum, have been identified on NifEN: a permanent cluster and a transient FeMoco precursor (19,21).…”

Section: Resultsmentioning

confidence: 99%

“…In the 8Fe and 7Fe models for the NifEN-bound FeMoco precursor, sulfur is shown as the ligand to one of the terminal iron atoms because the Cys residue that ligates FeMoco in the MoFe protein is conserved in the NifE primary sequence (20). The ligand at the opposite terminal iron atom is not given because it could be any number of light-atom ligands, including Asn, which replaces the FeMoco His ligand in the NifE sequence (20). sity of the short-range iron-iron scattering precludes a six-iron cluster.…”

Section: Resultsmentioning

confidence: 99%

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Structural insights into a protein-bound iron-molybdenum cofactor precursor

Corbett

Fay

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

102

107

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The iron-molybdenum cofactor (FeMoco) of the nitrogenase MoFe protein is a highly complex metallocluster that provides the catalytically essential site for biological nitrogen fixation. FeMoco is assembled outside the MoFe protein in a stepwise process requiring several components, including NifB-co, an iron-and sulfurcontaining FeMoco precursor, and NifEN, an intermediary assembly protein on which NifB-co is presumably converted to FeMoco. Through the comparison of Azotobacter vinelandii strains expressing the NifEN protein in the presence or absence of the nifB gene, the structure of a NifEN-bound FeMoco precursor has been analyzed by x-ray absorption spectroscopy. The results provide physical evidence to support a mechanism for FeMoco biosynthesis. The NifEN-bound precursor is found to be a molybdenum-free analog of FeMoco and not one of the more commonly suggested cluster types based on a standard [4Fe-4S] architecture. A facile scheme by which FeMoco and alternative, non-molybdenum-containing nitrogenase cofactors are constructed from this common precursor is presented that has important implications for the biosynthesis and biomimetic chemical synthesis of FeMoco.biosynthesis ͉ extended x-ray absorption fine structure ͉ nitrogenase ͉ x-ray absorption spectroscopy

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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Structural insights into a protein-bound iron-molybdenum cofactor precursor

Corbett

Fay

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

102

107

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“…The conserved regions include an ATPase motif and three cysteine residues, two of which are thought to be involved in chelating a 4Fe-4S cluster (51). Like BchY and BchZ of R. capsulatus (15), cysteine residues of BchY (55th, 80th, and 138th) and BchZ (35th) of R. sphaeroides also appear to be conserved at similar positions compared with those possibly responsible for the chelation of two 4Fe-4S clusters of NifE-NifN complex, another NifDK-like protein, which is involved in MoFe cofactor biosynthesis (52,53). Moreover, the co-purification of BchY-BchZ of R. capsulatus supported a nitrogenase model of COR (15).…”

Section: Suppressor Mutations Rescuing the Inability Of Sodb1 To Growmentioning

confidence: 98%

Superoxide Generation by Chlorophyllide a Reductase of Rhodobacter sphaeroides

Kim

Lee

et al. 2008

Journal of Biological Chemistry

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Chlorophyllide a reductase of Rhodobacter sphaeroides, which were reconstituted with the purified subunits of BchX, BchY, and BchZ, reduced ring B of chlorophyllide a using NADH under anaerobic conditions. Interestingly, suppressor mutations rescuing the inability of R. sphaeroides Fe-SOD mutant to grow in succinate-based minimal medium were predominantly mapped to BchZ subunit of chlorophyllide a reductase. The enzyme is labile in the presence of O 2 . However, it generates superoxide at low O 2 . The enzymes reconstituted with BchX, BchY, and the mutein subunit of BchZ from suppressor mutants showed less activity not only for chlorophyllide a reduction but also for superoxide generation compared with the enzyme reconstituted with the wild-type subunits. BchX, which contains FMN, and BchY are iron-sulfur proteins, whereas BchZ is a hemoprotein containing b-type heme. Neither chlorophyllide a reduction nor superoxide generation was observed with the enzyme reconstituted with the wild-type subunits of BchX and BchY, and the apo-subunit of BchZ that had been refolded without heme, in which FMN of BchX was fully reduced. Thus, superoxide is generated not from FMN of BchX but from heme of BchZ. Consistently, the heme of BchZ muteins was half-reduced in its redox state compared with that of wild-type BchZ.Rhodobacter sphaeroides, a facultative photosynthetic bacterium, contains two SODs 3 ; CuZn-SOD is detected only under the conditions where photosynthetic complexes are formed (1), whereas Fe-SOD is constitutively expressed, although its activity of the aerobically grown cell nearly doubled as compared with that of the anaerobically grown cell. The role of CuZn-SOD in protecting the photoheterotrophic cells from periplasmic superoxide upon exposure to O 2 was proposed (1). A cambialistic SOD using manganese or iron is the only SOD found in Rhodobacter capsulatus and is essential for cell viability (2, 3). Oxidative stress defense in R. sphaeroides and R. capsulatus is also mediated by catalases as well as by glutathione-glutaredoxin and thioredoxin systems that act as thiol-disulfide redox buffer to reduce the protein thiols that were oxidized (4, 5). Mutations in glutathione-glutaredoxin and thioredoxin systems lowered the formation of the photosynthetic complex (4, 5).The formation of photosynthetic complexes of R. sphaeroides is redox-dependent. Lowering oxygen tension induces the formation of intracytoplasmic membrane housing the photosynthetic complexes of R. sphaeroides. Light captured by B800 -850 and B875 LH complexes is transferred to reaction center complex, where the redox reactions are initiated to convert light energy into ATP and reducing power. The oxygenregulated expression of apoproteins of LH and reaction center complexes, which are encoded by puc, puf, and puh operons, was elucidated (for reviews see Refs. 6 and 7).The expression of several enzymes for Bchl a synthesis is also subject to anaerobic induction (8 -10). Protoporphyrin IX, which is synthesized from 5-aminolevulinic acid, is a common in...

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“…Biosynthesis of FeMoco presumably starts with the production of the Fe͞S core by NifB (encoded by nifB) (19,20), which is then transferred to, and further processed on, the ␣ 2 ␤ 2 tetrameric NifEN protein (17,21). Sequence similarity between the respective subunit-encoding genes led to the proposal that NifE and NifN form a structurally homologous complex to the MoFe protein (22,23) and that, by analogy, NifEN also contains two types of metal cluster sites, one corresponding to the P-cluster site and the other to the FeMoco site of the MoFe protein (17,21). The P-cluster analog was identified as a [4Fe-4S] cluster likely coordinated by conserved Cys residues at the NifE-NifN interface (21).…”

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confidence: 99%

FeMo cofactor maturation on NifEN

Corbett

Fay

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

163

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FeMo cofactor (FeMoco) biosynthesis is one of the most complicated processes in metalloprotein biochemistry. Here we show that Mo and homocitrate are incorporated into the Fe͞S core of the FeMoco precursor while it is bound to NifEN and that the resulting fully complemented, FeMoco-like cluster is transformed into a mature FeMoco upon transfer from NifEN to MoFe protein through direct protein-protein interaction. Our findings not only clarify the process of FeMoco maturation, but also provide useful insights into the other facets of nitrogenase chemistry. (9), which is located at the ␣␤-interface and ligated to six protein residues; and the [Mo-7Fe-9S-X-homocitrate] (the identity of X is unknown but is considered to be C, O, or N; ref. 10) FeMo cofactor (FeMoco), which is situated within the ␣-subunit and bound to only two protein residues and an exogenous homocitrate ligand. Both P-cluster and FeMoco are composed of smaller substructures: the P-cluster comprises two subclusters that share a 6 -sulfide (9) and FeMoco consists of [Mo-3Fe-3S] and [4Fe-3S] subcubanes that are bridged by three 2 -sulfides and share a central 6 -light atom (10). These metal clusters are essential for nitrogenase reaction, a process that involves ATP-dependent electron transfer from the [4Fe-

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Products of the iron-molybdenum cofactor-specific biosynthetic genes, nifE and nifN, are structurally homologous to the products of the nitrogenase molybdenum-iron protein genes, nifD and nifK

Cited by 168 publications

References 24 publications

Structural insights into a protein-bound iron-molybdenum cofactor precursor

Structural insights into a protein-bound iron-molybdenum cofactor precursor

Superoxide Generation by Chlorophyllide a Reductase of Rhodobacter sphaeroides

FeMo cofactor maturation on NifEN

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