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  • 1
    ISSN: 1432-072X
    Keywords: Nitrogenase regulation ; Glutamine synthetase ; Ammonia switch-off ; Rhodopseudomonas palustris
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Nitrogenase activity in Rhodopseudomonas palustris is subject to a rapid switch-off in response to exogenous ammonia. When cells were grown on limiting nitrogen and eventually became nitrogen deficient, nitrogenase synthesis was fully derepressed but the enzyme was insensitive to ammonia. The transformation of ammonia-sensitive to ammonia-insensitive cells was a slow, but fully reversible process. The switch-off effect in ammonia-sensitive cells paralleled changes in the adenylylation state of glutamine synthetase. Ammonia-insensitive cells, however, showed similar changes in glutamine synthetase activity although nitrogenase activity was unaffected. We conclude that nitrogenase regulation and adenylylation of glutamine synthetase are independent processes, at least under conditions of nitrogen deficiency.
    Type of Medium: Electronic Resource
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  • 2
    ISSN: 1432-072X
    Keywords: Nitrogen fixation ; Nitrogenase regulation ; Glutamine synthetase ; Methionine suofoximine ; Rhodospirillaceae
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Methionine sulfoximine (MSX), an irreversible inhibitor of glutamine synthetase of Rhodopseudomonas palustris restored nitrogenase activity to cells in which nitrogenase had been completely inhibited by ammonia switch-off. After addition of MSX, there was a lag period before nitrogenase activity was fully restored. During this lag, glutamine synthetase activity progressively decreased, and near the time of its complete inhibition, nitrogenase activity resumed. Nitrogenase switch-off by ammonia thus required active glutamine synthetase. Glutamine itself caused nitrogenase inhibition whose reversal by MSX depended on the relative ratio of MSX to glutamine. Unlike ammonia, glutamine inhibited nitrogenase under conditions where glutamine synthetase activity was absent. This indicates that glutamine is the effector molecule in nitrogenase switch-off, for instance by interacting with the enzymatic system for Fe protein inactivation. The effects of glutamine and MSX were also dependent on the culture age. Possible explanation for this and for the competitive effects are a common binding site within the regulatory apparatus for nitrogenase, or, in part, within a common transport system. Some observations with MSX were extended to Rhodopseudomonas capsulata and agreed with those in R. palustris.
    Type of Medium: Electronic Resource
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  • 3
    ISSN: 1432-072X
    Keywords: Key words Cytochrome cd1 ; Nitrite reductase ; Nitrous ; oxide reductase ; Denitrification ; Thiobacillus ; denitrificans ; Pseudomonas stutzeri ; DNA hybridization
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Cytochrome cd 1-nitrite reductase and nitrous oxide reductase of Thiobacillus denitrificans were purified and characterized by biochemical and immunochemical methods. In contrast to the generally soluble nature of the denitrification enzymes, these two enzymes were isolated from the membrane fraction of T. denitrificans and remained active after solubilization with Triton X-100. The properties of the membrane-derived enzymes were similar to those of their soluble counterparts from the same organism. Nitrous oxide reductase activity was inhibited by acetylene. Nitrite reductase and nitrous oxide reductase cross-reacted with antisera raised against the soluble enzymes from Pseudomonas stutzeri. The nirS, norBC, and nosZ genes encoding the cytochrome cd 1-nitrite reductase, nitric oxide reductase, and nitrous oxide reductase, respectively, from P. stutzeri hybridized with genomic DNA from T. denitrificans. Cross-reactivity and similar N-terminal amino acid and gene sequences suggest that the primary structures of the Thiobacillus enzymes are homologous to the soluble proteins from P. stutzeri.
    Type of Medium: Electronic Resource
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  • 4
    ISSN: 1432-072X
    Keywords: Nitrous oxide reduction ; Copper proteins ; Cytochrome patterns ; Denitrification ; Pseudomonas
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Copper is the essential transition element for nitrous oxide respiration in Pseudomonas perfectomarinus. Two novel kinds of copper proteins were detected in this organism. Their distribution was studied under different growth conditions and in other pseudomonads, as well as their association with N2O reduction of intact cells. A low molecular mass copper protein (M r 38,000) with a single absorption band at 340 nm (oxidized form), was found only in P. perfectomarinus and was not required for N2O reduction. N2O respiration was consistently associated with a high molecular mass copper protein (M r 120,000) in P. perfectomarinus, Pseudomonas stutzeri, and in strains of Pseudomonas fluorescens that were capable of this type of respiration. The oxidized protein was violet to pink with absorption bands at 350, 480, 530, 620, and 780 nm. Pseudomonas chlororaphis and Pseudomonas aureofaciens which did not respire with N2O as electron acceptor, did not contain the novel type of copper protein. Cytochrome patterns were compared in these denitrifying pseudomonads to search for the physiological electron carrier to N2O reductase. The content and nature of the soluble c-type cytochromes depended strongly on the species and the particular growth condition.
    Type of Medium: Electronic Resource
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  • 5
    ISSN: 1432-072X
    Keywords: Nucleotide sequence ; Apocytochrome cd 1 ; Heme d 1 incorporation ; Denitrification ; Copper coordination ; Signal peptide ; Escherichia coli
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract The structural gene, nirK, for the respiratory Cu-containing nitrite reductase from denitrifying Pseudomonas aureofaciens was isolated and sequenced. It encodes a polypeptide of 363 amino acids including a signal peptide of 24 amino acids for protein export. The sequence showed 63.8% positional identity with the amino acid sequence of “Achromobacter cycloclastes” nitrite reductase. Ligands for the blue, type I Cu-binding site and for a putative type-II site were identified. The nirK gene was transferred to the mutant MK202 of P. stutzeri which lacks cytochrome cd 1 nitrite reductase due to a transposon Tn5 insertion in its structural gene, nirS. The heterologous enzyme was active in vitro and in vivo in this background and restored the mutationally interrupted denitrification pathway. Transfer of nirK to Escherichia coli resulted in an active nitrite reductase in vitro. Expression of the nirS gene from P. stutzeri in P. aureofaciens and E. coli led to nonfunctional gene products. Nitrite reductase activity of cell extract from either bacterium could be reconstituted by addition of heme d 1, indicating that both heterologous hosts synthesized a cytochrome cd 1 without the d 1-group.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    Springer
    Antonie van Leeuwenhoek 71 (1997), S. 43-58 
    ISSN: 1572-9699
    Keywords: Denitrification ; mosaic gene organization ; nitrous oxide reductase ; nitric oxide reductase ; structural models ; cytochrome c oxidase
    Source: Springer Online Journal Archives 1860-2000
    Topics: Biology
    Notes: Abstract Denitrification is a main branch of the global nitrogen cycle. In the past ten years unravelling the underlying biochemistry and genetics has proceeded at an increasing pace. Fungal denitrification has become a new field. The biochemical investigation of denitrification has culminated in the description of the crystal structures of the two types of nitrite reductases. The N2O reductase shares with cytochrome c oxidase the CuA center as a structurally novel metal site. The cytochrome b subunit of NO reductase has a striking conservation of heme-binding transmembrane segments versus the subunit I of cytochrome c oxidase. Another putative denitrification gene product shows structural relation to the subunit III of the oxidase. N2O reductase and NO reductase may be ancestors of energy-conserving enzymes of the heme-copper oxidase superfamily. More than 30 genes for denitrification are located in a 〉30-kb cluster in Pseudomonas stutzeri, and comparable gene clusters have been identifi ed in Pseudomonas aeruginosa and Paracoccus denitrificans. Genes necessary for nitrite reduction and NO reduction have a mosaic arrangement with very few conserved locations within these clusters and relative to each other.
    Type of Medium: Electronic Resource
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