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Query: EC:1.7.1.4 (nitrite reductase)
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Seventeen strains of the new species Bacillus azotoformans were isolated by enrichment culture in peptone broth inoculated with pasteurized soil and then incubated under N2O at 32 degrees C. The bacterium is a Gram-negative rod, motile with peritrichous flagella, which produces oval spores without exosporia in swollen sporangia. However, the cells have thick walls, mesosomes, and persistent septa characteristic of Gram-positive bacteria. The bacterium lacks fermentative activity, does not attack carbohydrates, has complex growth requirements, and will grow anaerobically only if one of the following electron acceptors is present: NO3-, NO2-, N2O, S4O6--, or fumarate. Nitrate, nitrite, and nitrous oxide are denitrified with the production of N2. The microorganism is mesophilic, gives a positive oxidase reaction, synthesizes a type c cytochrome, and does not hydrolyse gelatin, starch, or "Tween 80." Poly-beta-hydroxybutyric acid is snythesized when the bacterium is grown in a medium containing DL-3-hydroxybutyrate. The following enzymes are present: nitrate reductase A, respiratory nitrite reductase, tetrathionate and fumarate reductases, and L-glutamate dehydrogenase. The following enzymes are absent: thiosulfate reductase, urease, lecithinase, arginine dihydrolase, phenylalanine deaminase, and catalase. For the 17 strains, the mean value of the G = C percent of the DNA is 39.8 +/- 1.2. All the strains are highly similar.
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PMID:[Morphological, physiological and taxonomic studies of Bacillus azotoformans]. 65 12

Chlorate resistant spontaneous mutants of Azospirillum spp. (syn. Spirillum lipoferum) were selected in oxygen limited, deep agar tubes with chlorate. Among 20 mutants from A. brasilense and 13 from A. lipoferum all retained their functional nitrogenase and 11 from each species were nitrate reductase negative (nr-). Most of the mutants were also nitrite reductase negative (nir-), only 3 remaining nir+. Two mutants from nr+ nir+ parent strains lost only nir and became like the nr+ nir- parent strain of A. brasilense. No parent strain or nr+ mutant showed any nitrogenase activity with 10 mM NO3-. In all nr- mutants, nitrogenase was unaffected by 10 mM NO3-. Nitrite inhibited nitrogenase activity of all parent strains and mutants including those which were nir-. It seems therefore, that inhibition of nitrogenase by nitrate is dependent on nitrate reduction. Under aerobic conditions, where nitrogenase activity is inhibited by oxygen, nitrate could be used as sole nitrogen source for growth of the parent strains and one mutant (nr- nir-) and nitritite of the parent strains and 10 mutants (all types). This indicates the loss of both assimilatory and dissimilatory nitrate reduction but only dissimilatory nitrite reduction in the mutants selected with chlorate.
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PMID:Nitrate and nitrite reductase negative mutants of N2-fixing Azospirillum spp. 69 99

The denitrifying capacity of 15 strains of Bacillus licheniformis was evaluated. In general, N2 production by the cultures on complex media containing NO3- is irregular and quite slow and three of the strains never produce gas. Bacillus licheniformis grows rapidly in anaerobiosis on peptone medium containing NO3- which is reduced to NO2-. None of the strains grow in peptone medium with NO2- or N2O as the respiratory substrate, nor do they grow under an atmosphere of 10% NO-90% N2. Denitrification was studied in cell suspensions using gas chromatography. N2O production from NO3- or NO2- is always weak at best; nitric oxide is reduced to N2O at an appreciable rate. All the strains synthesize nitrate reductase A in anaerobiosis when NO3- is present. In cell extracts, nitrite reductase activity is always negligible or nil with tetramethyl-p-phenylenediamine as an electron donor.
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PMID:[Denitrification by Bacillus licheniformis]. 75 76

The described bacterium was isolated by enrichment culture in peptone broth inoculated with garden soil, pasteurized and then put to incubate under N2O at 32 degrees. It is a Gram-negative rod, motile with peritrichous flagella, and producing oval spores without exosporium in swollen sporangia. However, cells have the thick walls, mesosomes and persistant septa characteristic of Gram-positive bacteria. It lacks fermentative activity, does not attack carbohydrates, has complex growth requirements, and will grow anaerobically only if one of the following electron acceptors is present: NO3, NO2, N2O, S4O6, and fumarate. Nitrate, nitrite, and nitrous oxide are denitrified with production of N2. The microorganism is mesophilic, gives a positive oxidase reaction, synthesizes a type of c cytochrome, and does not hydrolyse gelatin, starch nor "Tween 80". The following enzymes are present: nitrate reductase A, respiratory nitrite reductase, tetrathionate and fumarate reductases, L-glutamate dehydrogenase, and superoxide dismutase. The following enzymes are absent: thiosulfate reductase, urease, lecithinase, arginine dihydrolase, L-alanine dehydrogenase, phenylalanine desaminase, and catalase. The GC% of its DNA is 39. The bacterium described can be considered to be a new species. We propose the name Bacillus azotoformans n. sp.
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PMID:[A new, sporulating, denitrifying, mesophilic bacterium: Bacillus azotoformans N. SP. (author's transl)]. 102 Aug 72

The inhibitory effects of nitrate (NO3-) and nitrite (NO2-) on dissimilatory iron (FE3+) reduction were examined in a series of electron acceptor competition experiments using Shewanella putrefaciens 200 as a model iron-reducing microorganism. S. putrefaciens 200 was found to express low-rate nitrate reductase, nitrite reductase, and ferrireductase activity after growth under highly aerobic conditions and greatly elevated rates of each reductase activity after growth under microaerobic conditions. The effects of NO3- and NO2- on the Fe3+ reduction activity of both aerobically and microaerobically grown cells appeared to follow a consistent pattern; in the presence of Fe3+ and either NO3- or NO2-, dissimilatory Fe3+ and nitrogen oxide reduction occurred simultaneously. Nitrogen oxide reduction was not affected by the presence of Fe3+, suggesting that S. putrefaciens 200 expressed a set of at least three physiologically distinct terminal reductases that served as electron donors to NO3-, NO2-, and Fe3+. However, Fe3+ reduction was partially inhibited by the presence of either NO3- or NO2-. An in situ ferrozine assay was used to distinguish the biological and chemical components of the observed inhibitory effects. Rate data indicated that neither NO3- nor NO2- acted as a chemical oxidant of bacterially produced Fe2+. In addition, the decrease in Fe3+ reduction activity observed in the presence of both NO3- and NO2- was identical to the decrease observed in the presence of NO2- alone. These results suggest that bacterially produced NO2- is responsible for inhibiting electron transport to Fe3+.
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PMID:Effects of nitrate and nitrite on dissimilatory iron reduction by Shewanella putrefaciens 200. 154 35

Nitrate transport and its regulation by oxygen was studied in denitrifying halophilic Pseudomonas stutzeri, strain Zobell, and a Tn-5 transposon nitrite reductase mutant of this organism. The rate of nitrate transport was found to be 130 nanomoles nitrate min-1 mg protein-1 and 150 nanomoles nitrate min-1 mg protein-1 in the wildtype and the nitrite reductase mutant respectively as compared to 26.4 nanomoles nitrate min-1 mg protein-1 in a non-halophilic Pseudomonas stutzeri. Asparagine was found to be the best energy source for nitrate uptake. The ratio of nitrate import to nitrite export was established by measuring extracellular nitrate and nitrite concentrations using HPLC/UV analysis. There was a 1.3:1 (NO3-/NO2-) exchange. High concentrations of nitrate during growth was found to have a negative effect on nitrite metabolism. Oxygen exerted an inhibitory effect on nitrate uptake which was reversible and more pronounced in cells grown on low concentrations of nitrate compared to cells grown at high concentrations of nitrate.
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PMID:Regulation and energization of nitrate transport in a halophilic Pseudomonas stutzeri. 215 8

Various heterotrophic nitrifiers have been tested and found to also be aerobic denitrifiers. The simultaneous use of two electron acceptors (oxygen and nitrate) permits these organisms to grow more rapidly than on either single electron acceptor, but generally results in a lower yield than is obtained on oxygen, alone. One strain, formerly known as "Pseudomonas denitrificans", was grown in the chemostat and shown to achieve nitrification rates of up to 44 nmol NH3 min-1 mg protein-1 and denitrification rates up to 69 nmol NO3(-1) min-1 mg protein-1. Unlike Thiosphaera pantotropha, this strain needed to induce its nitrate reductase. However, the remainder of the denitrifying pathway was constitutive and, like T. pantotropha, "Ps. denitrificans" probably possesses the copper nitrite reductase.
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PMID:Aerobic denitrification in various heterotrophic nitrifiers. 261 86

Heterotrophic nitrification by Alcaligenes faecalis DSM 30030 was not restricted to media containing organic forms of nitrogen. In both peptone-meat extract and defined media with ammonium and citrate as the sole nitrogen and carbon sources, respectively, NO2-, NO3-, NO, and N2O were produced under aerobic growth conditions. Heterotrophic nitrification was not attributable to old or dying cell populations. Production of NO2-, NO3-, NO, and N2O was detectable shortly after cultures started growth and proceeded exponentially during the logarithmic growth phase. NO2- and NO3- production rates were higher for cultures inoculated in media with pH values below 7 than for those in media at alkaline pH. Neither assimilatory nor dissimilatory nitrate or nitrite reductase activities were detectable in aerobic cultures.
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PMID:Heterotrophic nitrification by Alcaligenes faecalis: NO2-, NO3-, N2O, and NO production in exponentially growing cultures. 278 77

Oxygen caused a reversible inhibition (switch-off) of nitrogenase activity in whole cells of four strains of diazotrophs, the facultative anaerobe Klebsiella pneumoniae and three strains of photosynthetic bacteria (Rhodopseudomonas sphaeroides f. sp. denitrificans and Rhodopseudomonas capsulata strains AD2 and BK5). In K. pneumoniae 50% inhibition of acetylene reduction was attained at an O2 concentration of 0.37 microM. Cyanide (90 microM), which did not affect acetylene reduction but inhibited whole-cell respiration by 60 to 70%, shifted the O2 concentration that caused 50% inhibition of nitrogenase activity to 2.9 microM. A mutant strain of K. pneumoniae, strain AH11, has a respiration rate that is 65 to 75% higher than that of the wild type, but its nitrogenase activity is similar to wild-type activity. Acetylene reduction by whole cells of this mutant was inhibited 50% by 0.20 microM O2. Inhibition by CN- of 40 to 50% of the O2 uptake in the mutant shifted the O2 concentration that caused 50% inhibition of nitrogenase to 1.58 microM. Thus, when the respiration rates were lower, higher oxygen concentrations were required to inhibit nitrogenase. Reversible inhibition of nitrogenase activity in vivo was caused under anaerobic conditions by other electron acceptors. Addition of 2 mM sulfite to cell suspensions of R. capsulata B10 and R. sphaeroides inhibited nitrogenase activity. Nitrite also inhibited acetylene reduction in whole cells of the photodenitrifier R. sphaeroides but not in R. capsulata B10, which is not capable of enzymatic reduction of NO2-. Lower concentrations of NO2- were required to inhibit the activity in NO3- -grown cells, which have higher activities of nitrite reductase.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Mechanism of nitrogenase switch-off by oxygen. 354 74

An electron capture gas-chromatographic technique was developed to continuously measure nitrate (NO3-) reduction during in vitro complementation tests with extracts from Pseudomonas aeruginosa mutants deficient in both assimilatory and dissimilatory nitrate reduction as a result of a single genetic mutation. The procedure involves coupling nitrate reduction to nitrous oxide (N2O) evolution via a series of reactions specific to the denitrification pathway. The assay was dependent on nitrate concentration, and optimal activity was obtained with a final concentration of 0.2% potassium nitrate. The reduction exhibited a stoichiometry of 2:1 (NO3-/N2O), and succinate was the best electron source for the reaction, followed by glucose, pyruvate, and malate. The technique can also be used for continuously monitoring nitrate reduction. The optimal nitrite concentration in the nitrite reductase assay was 0.025%. The initial complementation studies of mutant extracts demonstrated that at least two genes are shared between the two nitrate reduction pathways in P. aeruginosa.
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PMID:Gas chromatographic assay for in vitro complementation of Pseudomonas aeruginosa mutants deficient in nitrate reduction. 391 41


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