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:1.7.1.4 (nitrite reductase)
1,847 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A substantially improved purification of Escherichia coli NADH-dependent nitrite reductase was obtained by purifying it in presence of 1 mM-NO2- and 10 microM-FAD. The enzyme was obtained in 20% yield with a maximum specific activity of 1.04 kat . kg-1: more than 95% of this sample subjected to sodium dodecyl sulphate/polyacrylamide-gel electrophoresis migrated as a single band of protein. This highly active enzyme contained one non-covalently bound FAD molecule, and, probably, 5 Fe atoms and 4 acid-labile S atoms per subunit. No FMN, covalently bound flavin or Mo was detected. The spectrum of the enzyme shows absorption maxima at 386, 455, 530 and about 575 nm with a shoulder at 480--490 nm. The Soret-band/alpha-band absorbance ratio is about 4:1. These spectral features are characteristic of sirohaem, apart from the maximum at 455nm, which is attributed to flavin. The enzyme also catalyses the NADH-dependent reduction of horse heart cytochrome c, 2,6-dichlorophenol-indophenol and K3Fe(CN)6. The presence of sirohaem in E. coli nitrite reductase explains the apparent identity of the cysG and nirB gene of E. coli and inability of hemA mutants to reduce nitrite.
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PMID:Prosthetic groups of the NADH-dependent nitrite reductase from Escherichia coli K12. 703 Mar 14

Nitric oxide (NO) reductase is an integral membrane component of the anaerobic respiratory chain of Pseudomonas stutzeri that transforms nitrate to dinitrogen (denitrification). The enzyme catalyzes the reduction of NO to nitrous oxide. The structural genes for the NO reductase complex, norC and norB, were sequenced and their organization established by primer extension and Northern blot analysis. The norCB genes encoding the cytochrome c and cytochrome b subunits of the enzyme are contiguous and transcribed as a single 2.0-kb transcript. The promoter region has a canonical recognition motif for the transcriptional activator protein Fnr, centered at -40.5 nucleotides from the initiation site of transcription. No similarity of the derived gene products to known cytochromes of b- or c-type was found in a data bank search. Post-translational processing of the two subunits was limited to the removal of the terminal methionine to leave an N-terminal serine in either subunit. The mature cytochrome c subunit (16508Da, 145 residues) is predicted to be a bitopic protein with a single membrane anchor. The mature cytochrome b subunit (53006Da, 473 residues) is a putatively polytopic, strongly hydrophobic membrane-bound protein with 12 potential transmembrane segments. Several histidine and proline residues were identified with potentially structural and/or functional importance. Mutational inactivation of NO reductase by deletion of norB or the norCB genes affected strongly the in vivo activity of respiratory nitrite reductase (cytochrome cd1), but to a much lesser extent the expression level of this enzyme. In turn, mutational inactivation of the structural gene for cytochrome cd1, nirS, or loss of in vivo nitrite reduction by mutation of the nirT gene, encoding a presumed tetraheme cytochrome, lowered the expression level of NO reductase to 5-20%, but hardly its catalytic activity. The cellular concentration of NO reductase increased again on restoration of nitrite reduction in the nirS::Tn5 mutant MK202 by complementation with nirS or with the heterologous nirK gene, encoding the Cu-containing nitrite reductase from Pseudomonas aureofaciens. Thus, NO may be required as an inducer for its own reductase. Our results show that the nitrite-reducing system and the NO-reducing system are not operating independently from each other but are interlaced by activity modulation and regulation of enzyme synthesis.
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PMID:Nitric oxide reductase from Pseudomonas stutzeri. Primary structure and gene organization of a novel bacterial cytochrome bc complex. 750 88

The genes for nitric oxide reductase (norCB) from Pseudomonas aeruginosa were identified and sequenced. They are located about 2 kb upstream of nirS, the structural gene for nitrite reductase. norC and norB encode cytochrome c (16 kDa) and cytochrome b (52 kDa) subunits of the enzyme, respectively. norCB is immediately followed by an open reading frame encoding a protein of 612 residues.
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PMID:The structural genes for nitric oxide reductase from Pseudomonas aeruginosa. 771 Oct 73

By using the gene encoding the C-terminal part of the cd1-type nitrite reductase of Pseudomonas stutzeri JM300 as a heterologous probe, the corresponding gene from Paracoccus denitrificans was isolated. This gene, nirS, codes for a mature protein of 63144 Da having high homology with cd1-type nitrite reductases from other bacteria. Directly downstream from nirS, three other nir genes were found in the order nirECF. The organization of the nir gene cluster in Pa. denitrificans is different from the organization of nir clusters in some Pseudomonads. nirE has high homology with a S-adenosyl-L-methionine:uroporphyrinogen III methyltransferase (uro'gen III methylase). This methylase is most likely involved in the heme d1 biosynthesis in Pa. denitrificans. The third gene, nirC, codes for a small cytochrome c of 9.3 kDa having high homology with cytochrome c55X of Ps. stutzeri ZoBell. The 4th gene, nirF, has no homology with other genes in the sequence databases and has no relevant motifs. Inactivation of either of these 4 genes resulted in the loss of nitrite and nitric oxide reductase activities but not of nitrous oxide reductase activity. nirS mutants lack the cd1-type nitrite reductase while nirE, nirC and nirF mutants produce a small amount of cd1-type nitrite reductase, inactive due to the absence of heme d1. Upstream from the nirS gene the start of a gene was identified which has limited homology with nosR, a putative regulatory gene involved in nitrous oxide reduction. A potential FNR box was identified between this gene and nirS.
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PMID:Isolation, sequencing and mutational analysis of a gene cluster involved in nitrite reduction in Paracoccus denitrificans. 774 27

The complete amino acid sequence of cytochrome c-552 derived from the chemoautotrophic ammonia-oxidizing bacterium Nitrosomonas europaea was determined. The cytochrome consisted of 81 amino acid residues, and its molecular weight was calculated to be 9098 including heme c. Although the sequence of cytochrome c-552 was highly homologous to those of cytochromes c-551, which were known as the electron-donating components to dissimilatory nitrite reductase in pseudomonads, cytochrome c-552 differed from cytochrome c-551 in two points: (1) the sequence of cytochrome c-552 was shorter by two amino acid residues than that of cytochrome c-551 at the N-terminus and (2) one amino acid insertion was present in cytochrome c-552.
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PMID:The amino acid sequence of Nitrosomonas europaea cytochrome c-552. 776 24

A copper-containing nitrite reductase (Cu-NiR) was purified to homogeneity from the denitrifying fungus Fusarium oxysporum. The enzyme seemed to consist of two subunits with almost the same M(r) value of 41,800 and contains two atoms of copper per subunit. The electron paramagnetic resonance spectrum showed that both type 1 and type 2 copper centers are present in the protein, whereas the visible absorption spectrum exhibited a sole and strong absorption maximum at 595 nm, causing a blue but not green color. The reaction product due to the Cu-NiR was mainly nitric oxide (NO), whereas a stoichiometric amount of nitrous oxide (N2O) was formed when cytochrome P-450nor was further added to the assay system. Therefore, the denitrifying (N2O forming) nitrite reductase activity that we had detected in the cell-free extract of the denitrifying cells (Shoun, H., and Tanimoto, T. (1991) J. Biol. Chem. 266, 11078-11082) could be reconstituted upon combination of the purified Cu-NiR and P-450nor. The Km for nitrite and specific activity at pH 7.0 were estimated as 49 microM and 447 mumol NO.min-1.mg protein-1, respectively. Its activity was strongly inhibited by cyanide, carbon monoxide, and diethyldithiocarbamate, whereas enormously restored by the addition of cupric ions. An azurin-like blue copper protein (M(r) = 15,000) and a cytochrome c were also isolated from the same fungus, both of which together with cytochrome c of the yeast Saccharomyces cerevisiae were effective in donating electrons to the fungal Cu-NiR. The result suggested that the physiological electron donor of the Cu-NiR is the respiratory electron transport system. The intracellular localization of Cu-NiR was investigated, and it was suggested that the Cu-NiR localizes in an organelle such as mitochondrion. These findings showed the identity in many aspects between the fungal nitrite reductase and bacterial dissimilatory Cu-NiRs. This is the first isolation of dissimilatory NiR from a eukaryote.
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PMID:The copper-containing dissimilatory nitrite reductase involved in the denitrifying system of the fungus Fusarium oxysporum. 787 66

Monomeric nitrite reductase in an active form has been prepared by controlled succinylation of the dimeric native enzyme of Pseudomonas aeruginosa and subsequent purification. The monomeric enzyme has an optical spectrum indistinguishable from that of the native enzyme. On the other hand, circular dichroic spectra in the heme and peptide absorption regions show differences with respect to the dimer that indicate that the chemical modification and/or the dissociation into monomers somewhat perturb the chromophores' environment and the secondary structure. The (negatively charged) monomer is unable to oxidize its physiological substrates, azurin and cytochrome c551. This loss of activity is not due to monomerization, but is linked to the total net charge of the succinylated molecule, which interestingly enough acquires the ability to oxidize efficiently eukaryotic cytochrome c (which is not a substrate of the native dimeric enzyme). Stopped-flow studies show that the reduced monomer reacts with oxygen with a kinetic pattern similar to that shown by the dimeric enzyme. However, a higher reaction rate in the bimolecular binding of oxygen and a much higher oxygen affinity than for the native enzyme are observed. The evidence reported in this paper indicates that the dimeric state of Pseudomonas nitrite reductase is not a prerequisite for the ferrocytochrome c-oxygen oxidoreductase activity of this enzyme.
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PMID:Monomeric Pseudomonas aeruginosa nitrite reductase: preparation, characterization, and kinetic properties. 787 36

The nirC and nirF genes were identified downstream from nirSM, the structural genes encoding nitrite reductase (NIR) and cytochrome c-551 from Pseudomonas aeruginosa (Pa). The nirC gene encodes a probable c-type cytochrome with a signal sequence for membrane translocation. The nirF gene codes for a protein of 392 amino acids. A nirF mutant of Pa, constructed by marker exchange mutagenesis, synthesized an inactive NIR protein whose activity was restored by adding purified heme d1. The mutant strain produced an active NIR, when it was transformed by a broad-host-range plasmid carrying nirF. These results showed that the product of nirF was essential for the biosynthesis of heme d1 in Pa.
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PMID:Sequencing and characterization of the downstream region of the genes encoding nitrite reductase and cytochrome c-551 (nirSM) from Pseudomonas aeruginosa: identification of the gene necessary for biosynthesis of heme d1. 856 17

It has been suggested that two groups of Escherichia coli genes, the ccm genes located in the 47-min region and the nrfEFG genes in the 92-min region of the chromosome, are involved in cytochrome c biosynthesis during anaerobic growth. The involvement of the products of these genes in cytochrome c synthesis, assembly and secretion has now been investigated. Despite their similarity to other bacterial cytochrome c assembly proteins, NrfE, F and G were found not to be required for the biosynthesis of any of the c-type cytochromes in E. coli. Furthermore, these proteins were not required for the secretion of the periplasmic cytochromes, cytochrome C550 and cytochrome C552, or for the correct targeting of the NapC and NrfB cytochromes to the cytoplasmic membrane. NrfE and NrfG are required for formate-dependent nitrite reduction (the Nrf pathway), which involves at least two c-type cytochromes, cytochrome C552 and NrfB, but NrfF is not essential for this pathway. Genes similar to nrfE, nrfF and nrfG are present in the E. coli nap-ccm locus at minute 47. CcmF is similar to NrfE, the N-terminal region of CcmH is similar to NrfF and the C-terminal portion of CcmH is similar to NrfG. In contrast to NrfF, the N-terminal, NrfF-like portion of CcmH is essential for the synthesis of all c-type cytochromes. Conversely, the NrfG-like C-terminal region of CcmH is not essential for cytochrome c biosynthesis. The data are consistent with proposals from this and other laboratories that CcmF and CcmH form part of a haem lyase complex required to attach haem c to C-X-X-C-H haem-binding domains. In contrast, NrfE and NrfG are proposed to fulfill a more specialised role in the assembly of the formate-dependent nitrite reductase.
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PMID:The role of the genes nrf EFG and ccmFH in cytochrome c biosynthesis in Escherichia coli. 884 53

The assembly of anaerobically induced electron transfer chains in Escherichia coli strains defective in periplasmic disulphide bond formation was investigated. Strains deficient in DsbA, DsbB or DipZ (DsbD) were unable to catalyse formate-dependent nitrite reduction (Nrf activity) or synthesize any of the known c-type cytochromes. The Nrf+ activity and cytochrome c content of mutants defective in DsbC, DsbE or DsbF were similar to those of the parental, wild-type strain. Neither DsbC expressed from a multicopy plasmid nor a second mutation in dipZ (dsbD) was able to compensate for a dsbA mutation by restoring nitrite reductase activity and cytochrome c synthesis. In contrast, only the dsbB and dipZ (dsbD) strains were defective in periplasmic nitrate reductase activity, suggesting that DsbB might fulfil an additional role in anaerobic electron transport. Mutants defective in dipZ (dsbD) were only slightly more sensitive to Cu++ ions at concentrations above 5 mM than the parental strain, but strains defective in DsbA, DsbB, DsbC, DsbE or DsbF were unaffected. These results are consistent with our earlier proposals that DsbA, DsbB and DipZ (DsbD) are part of the same pathway for ensuring that haem groups are attached to the correct pairs of cysteine residues of apocytochromes c in the E. coli periplasm. However, neither DsbE nor DsbF are essential for the reduction of DipZ (DsbD).
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PMID:Effects of mutations in genes for proteins involved in disulphide bond formation in the periplasm on the activities of anaerobically induced electron transfer chains in Escherichia coli K12. 900 92


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