Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.7.1.2 (nitrate reductase)
3,861 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Seven genes, napKEFDABC, encoding the periplasmic nitrate reductase system were cloned from the denitrifying phototrophic bacterium Rhodobacter sphaeroides f. sp. denitrificans IL106. Two transmembrane proteins, NapK and NapE, an iron-sulfur protein NapF, a soluble protein NapD, a catalytic subunit of nitrate reductase precursor NapA, a soluble c-type diheme cytochrome precursor NapB, and a membrane-anchored c-type tetraheme cytochrome NapC were deduced as the gene products. Every mutant in which each nap gene was disrupted by omega-cassette insertion lost nitrate reductase activity as well as the ability of cells to grow with nitrate under anaerobic-dark conditions. A transconjugant of the napD-disrupted mutant with a plasmid bearing the napKEFDABC genes recovered both nitrate reductase activity and nitrate-dependent anaerobic-dark growth of cells. Denitrification activity, which was not observed in the napD mutant, was also restored by the conjugation. These results indicate that the periplasmic nitrate reductase encoded by the napKEFDABC genes is the enzyme responsible for denitrification in this phototroph, although the presence of a membrane-bound nitrate reductase has been reported in the same strain.
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PMID:Involvement in denitrification of the napKEFDABC genes encoding the periplasmic nitrate reductase system in the denitrifying phototrophic bacterium Rhodobacter sphaeroides f. sp. denitrificans. 1022 38

The seven nap genes at minute 47 on the Escherichia coli K-12 chromosome encode a functional nitrate reductase located in the periplasm. The molybdoprotein, NapA, is known to be essential for nitrate reduction. We now demonstrate that the two c-type cytochromes, the periplasmic NapB and the membrane-associated NapC, as well as a fourth polypeptide, NapD, are also essential for nitrate reduction in the periplasm by physiological substrates such as glycerol, formate and glucose. None of the three iron-sulphur proteins, NapF, NapG or NapH, are essential, irrespective of whether the bacteria are grown anaerobically in the presence of nitrate or fumarate as a terminal electron acceptor, or by glucose fermentation. Mutation of napD resulted in the total loss of Methyl Viologen-dependent nitrate reductase activity of the molybdoprotein, NapA, consistent with an earlier suggestion by others that NapD might be required for post-translational modification of NapA.
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PMID:Essential roles for the products of the napABCD genes, but not napFGH, in periplasmic nitrate reduction by Escherichia coli K-12. 1054 35

Nitrate reduction in the dissimilatory iron-reducing bacterium Geobacter metallireducens was investigated. Nitrate reductase and nitrite reductase activities in nitrate-grown cells were detected only in the membrane fraction. The apparent K(m )values for nitrate and nitrite were determined to be 32 and 10 microM, respectively. Growth on nitrate was not inhibited by either tungstate or molybdate at concentrations of 1 mM or less, but was inhibited by both at 10 and 20 mM. Nitrate and nitrite reductase activity in the membrane fraction was not, however, affected by dialysis with 20 mM tungstate. An enzyme complex that exhibited both nitrate and nitrite reductase activity was solubilized from membrane fractions with CHAPS and was partially purified by preparative gel electrophoresis. It was found to be composed of four different polypeptides with molecular masses of 62, 52, 36, and 16 kDa. The 62-kDa polypeptide [a low-midpoint potential (-207 mV), multiheme cytochrome c] exhibited nitrite reductase activity under denaturing conditions. No molybdenum was detected in the complex by plasma-emission mass spectrometry.
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PMID:A heme-C-containing enzyme complex that exhibits nitrate and nitrite reductase activity from the dissimilatory iron-reducing bacterium Geobacter metallireducens. 1055 Apr 73

We have conducted the genetic analysis of fermentative nitrate reduction in Clostridium perfringens, a strict anaerobic bacterium. Nitrate reductase (NarA) was purified from the cytoplasmic fraction of the organism. Using a degenerate primer designed from its N-terminal amino acid sequence, a 9.5 kb fragment containing seven ORFs was cloned. The molecular mass and N-terminal amino acid sequence predicted from the nucleotide sequence of ORF 4 coincided with those determined for the purified NarA, indicating that ORF 4 corresponds to a narA gene. ORFs 5 and 6 encode a 15.4 kDa ferredoxin-like protein containing four iron-sulfur clusters and a 45 kDa protein homologous to NADH oxidase, respectively. Analyses involving primer extension and Northern blotting revealed that these three ORFs are transcribed as a polycistronic message. The ORF 5- and ORF 6-encoded proteins were shown by immunoblotting to be synthesized by cells grown in the presence of nitrate. Thus, these two proteins are likely to function as electron-transfer components in nitrate reduction in C perfringens. The 9.5 kb fragment and a downstream region of 6.1 kb do not contain any genes involved in nitrate uptake or nitrite reduction. Instead, all 5 ORFs downstream of ORF 6 are homologous to genes reported for molybdopterin biosynthesis, unlike the genomic organization already determined for the respiratory and assimilatory nitrate-reduction systems. The evolutionary relationships between these two nitrate-reduction systems and the fermentative one based on the results of comparative genetic analysis are discussed.
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PMID:Analysis of genes involved in nitrate reduction in Clostridium perfringens. 1062 36

The periplasmic selenate reductase (Ser) of Thauera selennatis is a component of the electron transport chain catalyzing selenate reduction with acetate as the electron donor (i.e., selenate respiration). The purified enzyme consists of three subunits (SerA, SerB and SerC). Using transposon (i.e., Tn5) mutagenesis selenate reductase mutants were isolated. Junction fragments of DNA adjacent to the integrated Tn5 were used, together with oligonucleotides derived from the N-termini of SerA and SerB, to clone from a gene bank a DNA fragment that contained the corresponding genes. After sequencing, serA, serB and serC were identified by sequence comparison with the N-termini of the three subunits. The genes are arranged in the order serA, serB, serC; a fourth open reading frame (serD) in between, but overlapping serB and serC, is also present. The serA gene product contains an apparent leader peptide with a twin-arginine motif. The remainder of the translated amino acid sequence is similar to that of a number of prokaryotic molybdenum-containing enzymes (e.g., nitrate reductases and formate dehydrogenases of Escherichia coli). The serB gene product contains four cysteine clusters and is similar to various iron-sulfur protein subunits. The serC gene product contains a putative Sec-dependent leader peptide, but there are no similarities between the remainder of the translated protein and other protein subunits. The SerC contains two histidine and four methionine residues, and these may noncovalently bind heme b--which is a component of the active selenate reductase. The serD gene product encodes a putative protein that shows no significant sequence similarities to other proteins. However, the location of the serD within the other ser genes is similar to that of narJ within the E. coli narGHJI operon (nitrate reductase A); thus suggesting that the role of SerD may be similar to that of NarJ, which is a system-specific chaperone protein.
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PMID:Cloning and sequencing of the genes encoding the periplasmic-cytochrome B-containing selenate reductase of Thauera selenatis. 1082 93

The molybdopterin cofactor (MoCF) is required for the activity of a variety of oxidoreductases. The xanthine oxidase class of molybdoenzymes requires the MoCF to have a terminal, cyanolysable sulphur ligand. In the sulphite oxidase/nitrate reductase class, an oxygen is present in the same position. Mutations in both the ma-l gene of Drosophila melanogaster and the hxB gene of Aspergillus nidulans result in loss of activities of all molybdoenzymes that necessitate a cyanolysable sulphur in the active centre. The ma-l and hxB genes encode highly similar proteins containing domains common to pyridoxal phosphate-dependent cysteine transulphurases, including the cofactor binding site and a conserved cysteine, which is the putative sulphur donor. Key similarities were found with NifS, the enzyme involved in the generation of the iron-sulphur centres in nitrogenase. These similarities suggest an analogous mechanism for the generation of the terminal molybdenum-bound sulphur ligand. We have identified putative homologues of these genes in a variety of organisms, including humans. The human homologue is located in chromosome 18.q12.
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PMID:Comparison of the sequences of the Aspergillus nidulans hxB and Drosophila melanogaster ma-l genes with nifS from Azotobacter vinelandii suggests a mechanism for the insertion of the terminal sulphur atom in the molybdopterin cofactor. 1102 94

Significant recent advances have been made in studies of the major dissimilatory nitrate reductase (NarGHI) of Escherichia coli. This enzyme is a complex iron-sulfur ([Fe-S]) molybdoenzyme that oxidizes menaquinol or ubiquinol at a periplasmically oriented Q-site (Qp site), and reduces nitrate at a cytoplasmically-oriented molybdo-(bismolybdopterin guanine dinucleotide) (Mo-bisMGD) cofactor. The Qp site, as well as two hemes, termed bL and bH, are localized in a hydrophobic diheme cytochrome b(Narl) that: (i) provides a conduit for electron-transfer from the periplasmically-oriented Qp-site; (ii) provides a membrane anchoring functionality for the membrane-extrinsic subunits (NarGH) that coordinate the Mo-bisMGD (NarG) and four [Fe-S] clusters (NarH); and (iii) helps ensure the separation of sites of H+-yielding and H+-consuming reactions such that enzyme turnover leads to the generation of a proton-electrochemical potential across the cytoplasmic membrane. This minireview focuses on recent advances and future prospects for the diheme cytochrome b subunit (Narl) of NarGHI.
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PMID:The diheme cytochrome b subunit (Narl) of Escherichia coli nitrate reductase A (NarGHI): structure, function, and interaction with quinols. 1132 83

Respiratory reduction of nitrate to nitrite is the first key step in the denitrification process that leads to nitrate loss from soils. In Paracoccus pantotrophus, the enzyme system that catalyzes this reaction is encoded by the narKGHJI gene cluster. Expression of this cluster is maximal under anaerobic conditions in the presence of nitrate. Upstream from narK is narR, a gene encoding a member of the FNR family of transcriptional activators. narR is transcribed divergently from the other nar genes. Mutational analysis reveals that NarR is required for maximal expression of the membrane-bound nitrate reductase genes and narK but has no other regulatory function related to denitrification. NarR is shown to require nitrate and/or nitrite is order to activate gene expression. The N-terminal region of the protein lacks the cysteine residues that are required for formation of an oxygen-sensitive iron-sulfur cluster in some other members of the FNR family. Also, NarR lacks a crucial residue involved in interactions of this family of regulators with the sigma(70) subunit of RNA polymerase, indicating that a different mechanism is used to promote transcription. narR is also found in Paracoccus denitrificans, indicating that this species contains at least three FNR homologues.
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PMID:Maximal expression of membrane-bound nitrate reductase in Paracoccus is induced by nitrate via a third FNR-like regulator named NarR. 1137 24

The nitrate reductase of the hyperthermophilic archaeon Pyrobaculum aerophilum was purified 137-fold from the cytoplasmic membrane. Based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, the enzyme complex consists of three subunits with apparent molecular weights of 130,000, 52,000, and 32,000. The enzyme contained molybdenum (0.8-mol/mol complex), iron (15.4-mol/mol complex) and cytochrome b (0.49-mol/mol complex) as cofactors. The P. aerophilum nitrate reductase distinguishes itself from nitrate reductases of mesophilic bacteria and archaea by its very high specific activity using reduced benzyl viologen as the electron donor (V(max) with nitrate, 1,162 s(-1) (326 U/mg); V(max) with chlorate, 1,348 s(-1) (378 U/mg) [assayed at 75 degrees C]). The K(m) values for nitrate and chlorate were 58 and 140 microM, respectively. Azide was a competitive inhibitor and cyanide was a noncompetitive inhibitor of the nitrate reductase activity. The temperature optimum for activity was > 95 degrees C. When incubated at 100 degrees C, the purified nitrate reductase had a half-life of 1.5 h. This study constitutes the first description of a nitrate reductase from a hyperthermophilic archaeon.
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PMID:Properties of a thermostable nitrate reductase from the hyperthermophilic archaeon Pyrobaculum aerophilum. 1154 9

Dissimilatory nitrate reductase (Nar) was solubilized and partially purified from the large particle (mitochondrial) fraction of the denitrifying fungus Fusarium oxysporum and characterized. Many lines of evidence showed that the membrane-bound Nar is distinct from the soluble, assimilatory nitrate reductase. Further, the spectral and other properties of the fungal Nar were similar to those of dissimilatory Nars of Escherichia coli and denitrifying bacteria, which are comprised of a molybdoprotein, a cytochrome b, and an iron-sulfur protein. Formate-nitrate oxidoreductase activity was also detected in the mitochondrial fraction, which was shown to arise from the coupling of formate dehydrogenase (Fdh), Nar, and a ubiquinone/ubiquinol pool. This is the first report of the occurrence in a eukaryote of Fdh that is associated with the respiratory chain. The coupling with Fdh showed that the fungal Nar system is more similar to that involved in the nitrate respiration by Escherichia coli than that in the bacterial denitrifying system. Analyses of the mutant species of F. oxysporum that were defective in Nar and/or assimilatory nitrate reductase conclusively showed that Nar is essential for the fungal denitrification.
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PMID:Nitrate reductase-formate dehydrogenase couple involved in the fungal denitrification by Fusarium oxysporum. 1192 96


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