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1.
Two strains ofRhizobium, cowpeaRhizobium 32H1 andRhizobium japonicum CB 1809, showed a marked stimulation in growth on addition of formate to the minimal medium containing nitrate as the sole
source of nitrogen. The amount of accumulated nitrite and specific nitrate reductase activity was much higher in cultures
supplemented with formate than in the control medium. In contrast, growth, consumption of nitrite and specific nitrite reductase
activity in minimal medium + nitrite was greatly reduced by the addition of formate. A chlorate resistant mutant (Chl-16) was isolated spontaneously which contained a nitrite reductase which was not inhibited by formate. The results suggest
that formate serves as an electron donor for nitrate reductase and inhibits nitrite assimilation inRhizobium 相似文献
2.
3.
Paracoccus halodenitrificans produced a membrane-associated nitrite reductase. Spectrophotometric analysis showed it to be associated with a cd-cytochrome and located on the inner side of the cytoplasmic membrane. When supplied with nitrite, membrane preparations produced nitrous oxide and nitric oxide in different ratios depending on the electron donor employed. The nitrite reductase was maximally active at relatively low concentrations of sodium chloride and remained attached to the membranes at 100 mM sodium chloride. 相似文献
4.
Rhodopseudomonas sphaeroides f. denitrificans grown photosynthetically with NO
3
-
under anaerobic conditions accumulated NO
2
-
in the culture medium. In washed cells succinate, lactate, fumarate, citrate and malate, were effective electron donors for the reduction of NO
3
-
, NO
2
-
and N2O to N2 gas. Nitrate reductase was inhibited by amytal and potassium thocyanate. Nitrite reductase activity was severely restricted by potassium cyanide, sodium diethyldithiocarbamate, Amytal and 2-n-heptyl-4-hydroxyquinoline-N-oxide whereas N2O reductase was inhibited by NaN3, C2H2 and KCNS. Cells incubated with either K15NO3 or K15NO2 produced 15N2O and 15N2. A stoichiometry of 2:1 was recorded for the reduction of either NO
3
-
or NO
2
-
to N2O and N2 and for N2O to N2 it was 1:1.Abbreviations BVH
reduced benzyl viologen
- MVH
reduced methyl viologen
- HOQNO
2-n-heptyl-4-hydroxyquinoline-N-oxide
- CCCP
carbonyl cyanide-m-chlorophenyl-hydrazone
- DIECA
diethyl dithiocarbamate
- KCN
potassium cyanide 相似文献
5.
Application of nitrate leads to an induction of nitrate reductase (NR; EC 1.6.6.1) and nitrite reductase (NIR; EC 1.7.7.1) in the cotyledons of dark-grown mustard (Sinapis alba L.) seedlings, and this induction can strongly be promoted by a far-red-light pretreatment — operating through phytochrome — prior to nitrate application. This light treatment is almost ineffective — as far as enzyme appearance is concerned — if no nitrate is given. When nitrate is applied, the stored light signal potentiates the appearance of NR and NIR in darkness, even in the absence of active phytochrome, to the same extent as continuous far-red light. This action of previously stored light signal lasts for approx. 12 h.Storage of the light signal was measured for NR and NIR. The process shows enzyme-specific differences. Storage occurs in the absence as well as in the presence of nitrate, i.e. irrespective of whether or not enzyme synthesis takes place. The kinetics of signal transduction and signal storage indicate that the formation and action of the stored signal are a bypass to the process of direct signal transduction. Signal storage is possibly a means of enabling the plant to maintain the appropriate levels of NR and NIR during the dark period of the natural light/dark cycle.Abbreviations cD
continuous darkness
- cFR
continuous far-red light
- D
darkness
- FR
far-red light
- NIR
nitrite reductase (EC 1.7.7.1)
- NR
nitrate reductase (EC 1.6.6.1)
- Pfr
phytochrome (far-red absorbing)
- Pr
phytochrome (red absorbing)
- R
red light
- RG9-light
long wavelength far-red light obtained with RG9 glass filter
-
- Ptot
total phytochrome (Pr+Pfr)
Professor Wilhelm Nultsch mit guten Wünschen zum 60. Geburtstag 相似文献
6.
Grant Michael A. Cronin Sonja E. Hochstein Lawrence I. 《Archives of microbiology》1984,140(2-3):183-186
Membranes prepared from Paracoccus halodenitrificans reduced nitrite or nitric oxide to nitrous oxide. Extraction of these membranes with the detergent CHAPSO [3-(3-cholamidopropyldimethylammonio)-1-(2-hydroxy-1-propanesulfonate)], followed by ammonium sulfate fractionation of the solubilized proteins, resulted in the separation of nitrite and nitric oxide reductase activities. The fraction containing nitrite reductase activity spectrally resembled a cd-type cytochrome. Several cytochromes were detected in the nitric oxide reductase fraction. Which, if any, of these cytochromes is associated with the reduction of nitric oxide is not clear at this time.Abbreviations PMS
phenazine methosulfate
- HEPES
N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid
- CHAPSO
3-(3-cholamidopropyl-dimethylammonio)-1-(2-hydroxy-1-propanesulfonate)
- NH buffer
150 mM NaCl-50 mM
- HEPES
pH 7.5; octylglucoside, octyl--d glucopyranoside
- NIR
intrite reductase (nitrite to nitric oxide)
- NOR
nitric oxide reductase (nitric oxide to nitrous oxide) 相似文献
7.
8.
Molybdenum is required for induction of nitrate reductase and of NAD-linked formate dehydrogenase activities in suspensions of wild type Paracoccus denitrificans; tungsten prevents the development of these enzyme activities. The wild type forms a membrane protein M
r150,000 when incubated with tungsten and inducers of nitrate reductase and this is presumed to represent an inactive form of the enzyme. Suspensions of mutant M-1 did not develop nitrate reductase or formate dehydrogenase activities but the membrane protein M
r150,000 was formed under all conditions tested, including without inducers and without molybdenum. Analysis of membranes, solubilized with deoxycholate, by polyacrylamide gel electrophoresis under nondenaturing conditions showed that the mutant protein had similar electrophoretic mobility to the active nitrate reductase formed by the wilde type. Autoradiography of preparations from cells incubated with 55Fe showed that the mutant and wild type proteins contained iron. However, in similar experiments with 99Mo, incorporation of molybdenum into the mutant protein was not detectable.We conclude that mutant M-1 is defective in one or more steps required to process molybdenum for incorporation into molybdoenzymes. This failure affects the normal regulation of nitrate reductase protein with respect to the role of inducers.Non-Standard Abbreviations DOC
deoxycholate
- PAGE
polyacrylamide gel electrophoresis
- SDS
sodium dodecyl sulfate 相似文献
9.
Three nitrate reductase activities were detected in Alcaligenes eutrophus strain H16 by physiological and mutant analysis. The first (NAS) was subject to repression by ammonia and not affected by oxygen indicating a nitrate assimilatory function. The second (NAR) membrane-bound activity was only formed in the absence of oxygen and was insensitive to ammonia repression indicating a nitrate respiratory function. The third (NAP) activity of potential respiratory function occurred in the soluble fraction of cells grown to the stationary phase of growth. In contrast to NAR and NAS, expression of NAP did not require nitrate for induction and was independent of the rpoN gene product. Genes for the three reductases map at different loci. NAR and NAS are chromosomally encoded whereas NAP is a megaplasmid-borne activity in A. eutrophus. 相似文献
10.
Light inhibited each step of the denitrification process in whole cells of Rhodobacter sphaeroides forma sp. denitrificans. This inhibition by light was prevented in the presence of exogenous electron donors like N,N,N,N-tetramethyl-p-phenylenediamine (TMPD) plus ascorbate or in the presence of an uncoupler (carbonyl cyanide m-chlorophenylhydrazone). Addition of myxothiazol restored the inhibition by light in uncoupled cells. Measurements of light-induced absorbance changes under these conditions showed that this inhibition is due, for the steps of reduction of nitrite to dinitrogen, to the photooxidation of cytochromes c
1 plus c
2 and not due to the photoinduced membrane potential. Moreover, the presence of oxygen inhibited almost all of the reduction of nitrate and nitrous oxide but only 70% of the reduction of nitrite to nitrous oxide. These inhibitions were overcome in the presence of TMPD plus ascorbate. This implies that the inhibition in presence of oxygen was due to a diversion of the reducing power from the denitrifying chain to the respiratory chain. It was concluded from this series of experiments that the reduction of nitrate to nitrite is inhibited when the ubiquinone pool is partly oxidized and that nitrite and nitrous oxide reductions are inhibited when cytochromes c
1 plus c
2 are oxidized by photosynthesis or respiration.Abbreviations
R
Rhodobacter
- TMPD
N,N,N,N-tetramethyl-p-phenylenediamine
- HOQNO
2-n-heptyl-4-hydroxyquinoline N-oxide
- CCCP
carbonyl cyanide m-chlorophenylhydrazone
- cytochrome c
1
cytochrome c
2 plus cytochrome c
1 相似文献
11.
An experimental system has been devised for induction of nitrate reductase in suspensions of wild type Paracoccus denitrificans incubated with limited aeration in the presence of azide, nitrate or nitrite. Azide promoted maximum synthesis of enzyme, accompanied by formation of excess b-type cytochrome; the level of enzyme attained with nitrate was less and c-type cytochrome predominated in the membrane. The nitrate reductase was solubilized with deoxycholate from membranes of azide-induced cells and was identified as a major polypeptide M
r
=150,000 by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Mutants strains lacking nitrate reductase activity were isolated on the basis of resistance to chlorate and mutant M-1 was examined in detail. When incubated in the cell suspension system M-1 formed a membrane protein M
r
=150,000 similar to that attributed to nitrate reductase in the wild type. Maximum formation of the protein by M-1 occurred without inducer and it was accompanied by synthesis of excess b-type cytochrome. The observations with wild type and M-1 indicate that nitrate reductase protein and b-type cytochrome are coregulated and that the active enzyme has a role in regulating its own synthesis.Non-standard Abbreviations SDS
sodium dodecyl sulphate
- PAGE
polyacrylamide gel electrophoresis
- DOC
sodlum deoxycholate 相似文献
12.
Summary We investigated the inducibility of nitrate reductase (NR; EC 1.6.6.1), nitrite reductase (NiR; EC 1.7.7.1), and glutamine synthetase (GS; EC 6.3.1.2) isoforms in cotyledons of 7-day-old seedlings of sunflower (Helianthus annuus L.) in relation to light, nitrogen source (NO
3
–
, NO
2
–
or NH
4
+
), and the involvement of plastids. Nitrate was absolutely (and specifically) required for NR induction, and stimulated more effectively than NO
2
–
or NH
4
+
the synthesis of NiR and chloroplastic GS (GS2) over the constitutive levels present in N-free-grown seedlings. In vivo inhibition of NR activity by tungsten application to seedlings and measurements of tissue NO
3
–
concentration indicate that NO
3
–
-dependent enzyme induction is elicited by NO
3
–
per se and not by a product of its assimilatory reduction, e.g., NO
2
–
or NH
4
+
. In the presence of NO
3
–
, light remarkably enhanced the appearance of NR, NiR, and GS2, while the activity of the cytosolic GS isoform (GS1) was adversely affected. Cycloheximide suppressed much more efficiently than chloramphenicol the light- and NO
3
–
-dependent increase of GS2 activity, indicating that sunflower chloroplastic GS is synthesized on cytoplasmic 80S ribosomes. When the plastids were damaged by photooxidation in cotyledons made carotenoid-free by application of norflurazon, the positive action of light and NO
3
–
on the appearance of NR, NiR, and GS2 isoform was greatly abolished. Therefore, it is suggested that intact chloroplasts are required for the inductive effect of light and NO
3
–
and/or for the accumulation of newly formed enzymes in the organelle.Abbreviations CAP
chloramphenicol
- CHX
cycloheximide
- GS
glutamine synthetase
- GS1
cytosolic GS
- GS2
plastidic (chloroplastic) GS
- NF
norflurazon
- NiR
nitrite reductase
- NR
nitrate reductase 相似文献
13.
Pseudomonas oxalaticus was grown in carbon- and energy-limited continuous cultures either with oxalte or formate or with mixtures of these substrates. During growth on the mixtures, simultaneous utilization of the two substrates occurred at all dilution rates tested. Under these conditions oxalate repressed the synthesis of ribulosebisphosphate carboxylase. The degree of this repression was dependent on the dilution rate and the ratio of oxalate and formate in the medium reservoir. At a fixed oxalate/formate ratio repression was greatest at intermediate dilution rates, whereas derepression occurred at both low and high dilution rates. Progressive depression of ribulosebisphosphate carboxylase synthesis and of autotrophic CO2 fixation at low dilution rates was attributed to the decreasing concentration of intracellular repressor molecule(s), parallel to the decreasing concentration of the growth-limiting substrates in the culture. To account for the derepression at higher dilution rates, it is proposed that the rate of oxalyl-CoA production from oxalate limits the supply of metabolic intermediates and that additional energy and reducing power generated from formate drains the pools of metabolic intermediates sufficiently to lower the intracellular concentration of the repressor(s). During growth of Pseudomonas oxalaticus on the heterotrophic substrate oxalate alone, at dilution rates below 10% of the maximum specific growth rate, derepression of ribulosebisphosphate carboxylase synthesis and of autotrophic CO2 fixation was observed to a level which was 50% of that observed during growth on formate alone at the same dilution rate. It is concluded that in Pseudomonas oxalaticus the synthesis of enzymes involved in autotrophic CO2 fixation via the Calvin cycle is regulated by a repression/derepression mechanism and that the contribution of autotrophic CO2 fixation to the biosynthesis of cell material in this organism is mainly controlled via the synthesis of these enzymes.Abbreviations RuBPCase
ribulosebisphosphate carboxylase
- PMS
phenazine methosulphate
- DCPIP
2,6-dichlorophenolindophenol
- FDH
formate dehydrogenase
- SR
concentration of growth-limiting substrate in reservoir 相似文献
14.
Effects of artificial electron donors to deliver reducing power on enzymic denitrification were investigated using nitrate
reductase and nitrite reductase obtained fromOchrobactrum antropi. The activity of nitrite reductase in the soluble portion was almost the same as that in the precipitated portion of the
cell extract. Nitrate removal efficiency was higher with benzyl viologen than with methyl viologen or NADH as an artificial
electron donor. The turn-over numbers of nitrate and nitrite reductase were 14.1 and 1.9 μmol of nitrogen reduced/min·mg cell
extracts, respectively when benzyl viologen was used as an electron donor. 相似文献
15.
Growth of Pseudomonas oxalaticus in carbon- and energy-limited continuous cultures with mixtures of acetate and formate resulted in the simultaneous utilization of both substrates at all dilution rates tested. During growth on these mixtures, acetate repressed the synthesis of ribulosebisphosphate carboxylase. The degree of this repression was dependent on the dilution rate and on the ratio of acetate and formate in the medium reservoir. At fixed acetate and formate concentrations in the inflowing medium of 30 and 100 mM, respectively, and dilution rates above 0.10h-1, the severe repression of autotrophic enzymes resulted in a marked increase in bacterial dry weight compared to the growth yield of the organisms on the two substrates separately. Also, at these dilution rates a significant increase in isocitrate lyase activity was observed in the cells as compared to growth on acetate alone. This indicated that under these conditions more acetate was assimilated and less dissimilated since acetate was partly replaced by formate as the energy source. When formate was added to the reservoir of an acetate-limited culture (SR=30 mM), derepression of RuBPCase synthesis was observed at formate concentrations of 50 mM and above. Below this concentration formate only served as an energy source for acetate assimilation; when its concentration was increased above 50 mM a progressively increasing contribution of carbon dioxide fixation to the total carbon assimilation was observed as the activity of RuBPCase in the cells increased. It is concluded that in Pseudomonas oxalaticus the synthesis of enzymes involved in autotrophic carbon dioxide fixation via the Calvin cycle is regulated by a repression/derepression mechanism.Abbreviations RuBPCase
ribulosebisphosphate carboxylase
- PMS
phenazine methosulphate
- DCPIP
2,6-dichlorophenol-indophenol
- FDH
formate dehydrogenase
- SR
concentration of growth-limiting substrate in reservoir 相似文献
16.
Elisabetta Zennaro Ilaria Ciabatti Francesca Cutruzzola Rosanna D'Alessandro Maria Chiara Silvestrini 《FEMS microbiology letters》1993,109(2-3):243-250
Abstract The expression of nitrite reductase has been tested in a wild-type strain of Pseudomonas aeruginosa (Pao1) as a function of nitrate concentration under anaerobic and aerobic conditions. Very low levels of basal expression are shown under non-denitrifying conditions (i.e. absence of nitrate, in both aerobic and anaerobic conditions); anaerobiosis is not required for high levels of enzyme production in the presence of nitrate. A Pseudomonas aeruginosa strain, mutated in the nitrite reductase gene, has been obtained by gene replacement. This mutant, the first of this species described up to now, is unable to grow under anaerobic conditions in the presence of nitrate. The anaerobic growth can be restored by complementation with the wild-type gene. 相似文献
17.
Isolation and characterisation of a strain of Pseudomonas putida that can express a periplasmic nitrate reductase 总被引:2,自引:0,他引:2
A strain of Pseudomonas putida that can express a nitrate reductase that is located in the periplasmic compartment was isolated from freshwater. The enzyme was active in vivo during arginine fermentation and at the onset of oxygen limitation in batch cultures. The activity of the enzyme increased the yield of bacteria following fermentative growth under anoxic conditions with arginine, but nitrate reduction did not support growth on nonfermentable carbon substrates under anoxic conditions. Cells expressing the periplasmic nitrate reductase were capable of reducing nitrate in the presence of oxygen. Nitrate reduction under oxic conditions was clearly coupled to a respiratory electron transport chain because: (1) the process was sensitive to the respiratory inhibitors rotenone and 2-n-heptyl-4-hydroxyquinoline N-oxide, and (2) membrane-bound and periplasmic cytochromes were involved. This is the first report of the presence of a periplasmic nitrate reductase in a member of the proteobacteria. 相似文献
18.
Nitrite reductase (NIR; EC 1.7.7.1) is a central enzyme in nitrate assimilation and is localized in plastids. The present study concerns the regulation of the appearance of NIR in cotyledons of the mustard (Sinapis alba L.) seedling. It was shown that light exerts its positive control over the nitrate-mediated induction of NIR via the farred-absorbing form of phytochrome. Without nitrate the light effect cannot express itself; even though the light signal is accumulated in the cotyledons it remains totally cryptic in the absence of nitrate. Moreover, it was recognised that intact plastids are important in the control of the appearance of NIR. If the plastids are damaged by photooxidation the action of nitrate and phytochrome on NIR appearance is abolished. The appearance of nitrate reductase (NR; EC 1.6.6.1) responds similarly to photooxidative damage even though this enzyme is cytosolic. While the data strongly indicate that some plastidic signal is a prerequisite for the nitrate-induced and phytochrome-modulated appearance of NIR and NR, the possibility could not be ruled out that photooxidative damage affects the accumulation of NIR in the organelle.Abbreviations c
continuous
- D
darkness
- FR
far-red light
- NADP-GPD
NADP-dependent glyceraldehyde-3-phosphate dehydrogenase (EC 1.2.1.1.13)
- NF
Norflurazon
- NIR
nitrite reductase (EC 1.7.7.1.)
- NR
nitrate reductase (EC 1.6.6.1)
- Pfr
phytochrome (far-red light obtained with RG9 glass filter
- R
red light
- RG9-light
long wavelenght far-red light obtained with RG9 glass filter
- RuBPCase
ribulose-1,5-bisphosphate carboxylase (EC 4.1.1.39)
- WL
white light
- WLs
strong white light (28 W m-2) 相似文献
19.
Nitrate-induced and phytochrome-modulated appearance of nitrate reductase (NR; EC 1.6.6.1) and nitrite reductase (NIR; EC 1.7.7.1) in the cotyledons of the mustard (Sinapis alba L.) seedling is strongly affected by externally supplied ammonium (NH
4
+
). In short-term experiments between 60 and 78 h after sowing it was found that in darkness NH
4
+
—simultaneously given with NO
3
-
—strongly inhibits appearance of nitrate-inducible NR and NIR whereas in continuous far-red light—which operates exclusively via phytochrome without significant chlorophyll formation —NH
4
+
(simultaneously given with NO
3
-
) strongly stimulates appearance of NR. The NIR levels are not affected. This indicates that NR and NIR levels are regulated differently. In the absence of external NO
3
-
appearance of NR is induced by NH4 in darkness as well as in continuous far-red light whereas NIR levels are not affected. On the other hand, in the absence of external NO
3
-
, exogenous NH
4
+
strongly inhibits growth of the mustard seedling in darkness as well as in continuous far-red light. This effect can be abolished by simultaneously supplying NO
3
-
. The adverse effect of NH
4
+
on growth (NH
4
+
-toxicity) cannot be attributed to pH-changes in the medium since it was shown that neither the growth responses nor the changes of the enzyme levels are related to pH changes in the medium. Non-specific osmotic effects are not involved either.Abbreviations c
continuous
- D
darkness
- FR
far-red light
- NIR
nitrite reductase (EC 1.7.7.1)
- NR
nitrate reductase (EC 1.6.6.1) 相似文献
20.
Strain HUUG25 ofParacoccus denitrificans has been frequently thought to be devoid of allc-type cytochromes. We show here by means of enzymological and immunological techniques that the mutant synthesizes active
nitrite reductase (cytochromecd
1) upon prolonged exposure to microoxic conditions. The synthesis occurred faster in the presence of exogenous hemin. The time
pattern of 5-aminolevulinate synthase activity was also altered by the mutation. These findings suggest a defective regulation
of heme supply to the site of nitrite reductase assembly in the periplasm. 相似文献