The Three-dimensional Structure of the Core Domain of Naf Y from Azotobacter vinelandii determined at 1.8-Å Resolution

Dyer, David H.; Rubio, Luis M.; Thoden, James B.; Holden, Hazel M.; Ludden, Paul W.; Rayment, Ivan

doi:10.1074/jbc.m304264200

Cited by 26 publications

(45 citation statements)

References 33 publications

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“…As compared to the β-galactosidase activity obtained from E. coli cotransformants harboring only the pBT and pTRG plasmids (negative control--22.93 + 6.12 Miller Units), the activity obtained from cells harboring pMH6002 and pMH5002 was much higher (49.7 + 9.11 Miller Units), indicating a protein-protein interaction between NifB and NifX ( Table 2). Since the NifB and NafY proteins share many similarities between their amino acid sequences and since NafY was shown to bind to the apodinitrogenase [35] , we also examined the ability of NifB and NifK to interact with each other and found that the β-galactosidase activity of E. coli cotransformants harboring the plasmids pMH6002 (λCI:NifB) and pBG1716 (RNAP:NifK) was 20.47 + 4.78 Miller Units, suggesting lack of interaction between NifB and NifK.…”

Section: Interactionmentioning

confidence: 99%

“…an N-terminal (Met1 to Leu98) domain and a Cterminal (Glu99 to Ser232) 'core' domain [35] . The NafY core domain was shown to be capable of binding the FeMoco of nitrogenase but unable to bind to apodinitrogenase in the absence of the N-terminal domain [35] .…”

Section: Interaction Of Nafy With Nifxmentioning

confidence: 99%

“…Eventhough a deletion in the nifX gene did not show a significant effect on the nitrogenase activity in vivo, the purified NifX protein was shown to be important in the stimulation of FeMoco synthesis in an in vitro FeMoco synthesis reaction [10,34] . The NifX, NifB, NifY and NafY are most likely related proteins, owing to their overall sequence similarity and the presence of a FeMoco binding 'core domain' in all of them [35] . This conserved family of proteins has a β 1 −β 2 −β 3 −α 1 −α 2 −β 4 −α 3 −β 5 −α 4 −α 5 fold in its core domain, a polarized surface charge distribution and shows considerable similarity with the ribonuclease H family [35] .…”

Section: Introductionmentioning

confidence: 99%

“…The NifX, NifB, NifY and NafY are most likely related proteins, owing to their overall sequence similarity and the presence of a FeMoco binding 'core domain' in all of them [35] . This conserved family of proteins has a β 1 −β 2 −β 3 −α 1 −α 2 −β 4 −α 3 −β 5 −α 4 −α 5 fold in its core domain, a polarized surface charge distribution and shows considerable similarity with the ribonuclease H family [35] . In NifB, the NIFX SMBD is fused with a biotin-synthase-like, metal-dependent catalytic domain and in the archaeal MTH1172 protein, it is fused to a cation transporter [33] .…”

Section: Introductionmentioning

confidence: 99%

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The NifX Protein is Involved in the Final Stages of FeMo-cofactor Transport to the MoFe Protein

Lahiri¹,

Cole²,

Pulakat³

et al. 2007

American J. of Biochemistry and Biotechnology

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Abstract:The nitrogenase enzyme catalyzes the reduction of dinitrogen to ammonia and is composed of the Fe and MoFe proteins. The iron molybdenum cofactor (FeMo-co) of the MoFe protein is the site of active substrate reduction. The NifX protein has also been suggested to have a role in the FeMo-co synthesis, although its exact role is still open to investigation. We attempted to understand the role of NifX by determining the specific interactions it may have with other Nif proteins involved in FeMo-co synthesis, such as NifD, NifK, NifN, NifDK and NifH. Using the BacterioMatch Two-Hybrid System, a translationally fused construct of NifX with the N-terminal α-RNAP of the pTRG target vector was made and its interaction was tested with the NifDK fusion protein, translationally fused to the λCI of the pBT vector. The strength of the interaction, as determined by measuring the β-galactosidase activity, demonstrated that direct protein-protein interaction exists between NifDK and NifX proteins; the extent of interaction between NifK and NifX proteins was much higher than between NifD and NifX, when individually tested; also, reduced interaction was found between NifH and NifX.

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Section: Interactionmentioning

confidence: 99%

Section: Interaction Of Nafy With Nifxmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The NifX Protein is Involved in the Final Stages of FeMo-cofactor Transport to the MoFe Protein

Lahiri¹,

Cole²,

Pulakat³

et al. 2007

American J. of Biochemistry and Biotechnology

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“…64 The "scaffolding" proteins serve more the purpose of assembling and stabilizing a complex metal cofactor prior to delivery to a target protein. Thus far, proteins involved in Fe-S cluster assembly, 26,65 heme transport, 66,67 and a putative protein implicated in the biosynthesis of molybdenum cofactor 68 and delivery have been structurally characterized. The differences in struc- …”

Section: Cast Of Characters In Iron-sulfur Cluster Biosynthesismentioning

confidence: 99%

Iron-Sulfur Cluster Biosynthesis

Agar¹,

Dean²,

Johnson³

Biochemistry and Physiology of Anaerobic Bacteria

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Iron-sulfur clusters are among the most complex metal-containing prosthetic centers in biology. Most if not all of the proteins involved in the biosynthesis of "simple" Fe-S clusters have been identified. The structural and functional chemistry of these proteins has been the subject of intense research efforts, and many of the key details are now understood in structural and mechanistic detail. The fact that Fe-S cluster-binding proteins can be reconstituted in vitro with no accessory proteins provides an important indicator of the intracellular roles for many proteins on the Fe-S cluster assembly pathway. Indeed, such proteins are more correctly viewed as carrier proteins, rather than as catalysts for the reaction, that both avoid the toxicity associated with free iron and sulfide and allow delivery at lower intracellular concentrations of these species. The IscU (or ISU) family of proteins serves a key role as scaffolding proteins on which [2Fe-2S] building blocks are assembled prior to transfer to final apo target proteins. IscU in particular exhibits highly unusual conformational flexibility that appears critical to its function.

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