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PubMedDate not supplied

[The effect of chronic UVB-radiation on the nitrogen-fixing capability of soils in a field experiment].

The effect of chronic UV-B-irradiation on the nitrogen-fixing ability of soil was studied in a field experiment. In the study, LER-40 lamps were used as a source of additional UV-B-irradiation with respect to natural illumination (the irradiation level = 0.34, 0.49 and 0.77 W/m2). Nitrogen-fixation was measured by gas chromatography. The experimental results were treated by bifactorial analysis of variance. It was found that chronic UV-B-irradiation affected the nitrogen-fixing ability of soil during the observation period (June-August). At soil UV-B-irradiation of 0.34 and 0.49 W/m2 the nitrogen-fixation intensity increased by 17 and 9%, respectively, and at soil UV-B-irradiation of 0.77 W/m2 it decreased by 13%, when compared to untreated controls.

Nitrogen Fixation↗
PubMedDate not supplied

[Physiological and agrochemical properties of different symbiotic genotypes of pea (Pisum sativum L.)].

Physiological characters of symbiotic mutants of pea were studied: nodulation, activities of nitrogenase and nitrate reductase, chlorophyll content in leaves and their water-holding capacity, biomass accumulation, and nitrogen forms. The parameters reflecting the genotype state of the macrosymbiont under soil conditions considerably varied. Supernodulation mutants stood out against symbiotic pea genotypes by high contents of chlorophyll and nonprotein nitrogen compounds, high nitrogenase activity, and low nitrate reductase activity. The efficiency of the legume-rhizobium symbiosis was largely mediated by the macrosymbiont genotype. The highest atmospheric nitrogen fixation (50-80%) was observed in the parental pea varieties. Despite the highest nitrogenase activity in the nodules, the supernodulation mutants were inferior to the parental varieties by the nitrogen fixation capacity (40-60%), which was due to their low productivity.

Genotype↗
PubMed1993

Functional organization of the glnB-glnA cluster of Azospirillum brasilense.

The functional organization of the glnB-A cluster of Azospirillum brasilense, which codes for the PII protein and glutamine synthetase, respectively, was studied with the aid of lacZ fusions, deletion mapping, site-directed mutagenesis, and complementation. It was shown previously by mRNA mapping that the cluster contains two tandemly organized promoters, glnBp1 and glnBp2, of the sigma 70 and sigma 54 types, respectively, upstream of glnB and a third unidentified promoter upstream of glnA. Data obtained with lacZ fusions in the wild-type strain confirmed that cotranscription of glnBA and transcription of glnA alone were oppositely regulated by the cell N status. Quantification of promoter activities showed a high level of transcription from glnBp1p2 and a low level from glnAp under conditions of nitrogen limitation. The opposite situation prevails under conditions of nitrogen excess. As a consequence, PII polypeptide synthesis is increased under conditions of nitrogen fixation, which strongly suggests that PII plays an important role under these conditions. Null mutant strains of glnB, ntrB-ntrC, nifA, and point mutant strains in glnA were analyzed. NtrB and NtrC are not involved in the regulation of glnBA expression, in contrast to PII and glutamine synthetase. Glutamine synthetase probably acts by modulating the intracellular N status, and PII acts by modifying the properties of an unidentified regulator which might be a functional homolog of NtrC. In addition, a Nif- null mutant strain of glnB was characterized further. A Nif+ phenotype was restored to the strain by nifA from Klebsiella pneumoniae but not by nifA from A. brasilense. This mutant strain is not impaired in NifA synthesis, which is relatively independent of the growth conditions in A. brasilense. It is therefore most likely that PII is required for NifA activation under conditions of nitrogen fixation. Deletion mapping and site-directed mutagenesis showed glnAp was located within a 45-bp DNA fragment upstream of the mRNA start site, dissimiar to previously described consensus sites for sigma factors.

Ammonia↗
PubMedDate not supplied

[Effect of lyophilization on the nitrogenase activity of Azotobacter vinelandii].

The activity of nitrogenase in the cells of Azobacter vinelandii grown from lyophilized and non-lyophilized cultures depends on the donor of hydrogen and the concentration of oxygen in the gaseous phase. The lyophilized cells are more sensitive to oxygen (O2 optimum for nitrogen fixation is ca. 1 percent) than the non-lyophilized cells (ca. 5 percent). The determination of acetylene reduction in the course of the culture growth has shown that nitrogen fixation in the lyophilized cells takes place after a lag-period (about six hours) at a rate lower than that of the non-lyophilized cells. The results obtained suggest that lyophilization increases the sensitivity of the cells to oxygen and decreases their nitrogenase activity which is however restored after a while.

Aerobiosis↗
PubMed2003

Azoarcus sp. strain BH72 as a model for nitrogen-fixing grass endophytes.

The availability of nitrogen often limits plant growth in terrestrial ecosystems. The only biological reaction counterbalancing the loss of N from soils or ecosystems is biological nitrogen fixation, the enzymatic reduction of N2 to ammonia. Some gramineous crops such as certain Brazilian sugar cane cultivars or Kallar grass can derive a substantial part of the plant nitrogen from biological nitrogen fixation. Our research on grass-associated diazotrophs focuses on endophytic bacteria, microorganisms that multiply and spread inside plants without causing damage of the host plants or conferring an ecological threat to the plant. This review summarizes the current knowledge on the diazotrophic endophyte Azoarcus sp. BH72, which is capable of colonizing the interior of rice roots, one of the globally most important crops.

Azoarcus↗
PubMed1988

Characteristics of a nitrogen-fixing methanotroph, Methylocystis T-1.

A methane-oxidizing bacterium capable of nitrogen fixation was isolated from soil taken from an area which leaked methane gas. Strain T-1 was a catalase and oxidase-positive, gram-negative straight rod-shaped strictly aerobic bacterium which formed lipid cysts and type II intracytoplasmic membranes. The organism was a microaerophilic nitrogen-fixing methanotroph. Strain T-1 is considered to be classified into Methylocystis. The organism evolved hydrogen gas when grown in the nitrogen-free medium of atmospheric oxygen concentrations of 1.5% or more. Below this level, however, hydrogen gas was not evolved. In addition to methanol, formaldehyde and formate, ethanol, acetate and hydrogen gas served as oxidizable substrates for the acetylene reduction test. H2-stimulated nitrogenase activity was limited in a very narrow range of oxygen concentration and not detected at 2% O2. With acetate as the substrate, however, about an 80% of the maximum acetylene reduction activity was detected at 2% O2. These results suggest that strain T-1 is capable of recycling the hydrogen gas evolved during nitrogen fixation under low partial pressures of O2.

Hydrogen↗
PubMed1990

The nifH promoter region of Rhizobium leguminosarum: nucleotide sequence and promoter elements controlling activation by NifA protein.

The nucleotide (nt) sequence of the Rhizobium leguminosarum nifH promoter region contains a consensus promoter, a consensus upstream activator sequence (UAS), a pseudo (psi) promoter and a psi UAS. We mapped the transcription start point for the consensus promoter sequence by primer extension. This promoter differs from the consensus in one of the four supposedly invariant nt and can be activated by the Klebsiella pneumoniae nifA product in Escherichia coli. Under these conditions the psi promoter and psi UAS do not function. A low-copy-number plasmid construct containing the psi UAS as well as the consensus UAS delayed the onset of symbiotic nitrogen fixation in nodules induced on Pisum sativum. Studies of high-copy-number nifH promoter constructs showed that partial deletion of the consensus UAS does not alter the ability to inhibit nitrogen fixation by titration of NifA suggesting that NifA can also complex with RNA polymerase containing the alternative sigma-factor RpoN.

Bacterial Proteins↗
PubMed2000

Mutagenesis and functional characterization of the glnB, glnA, and nifA genes from the photosynthetic bacterium Rhodospirillum rubrum.

Nitrogen fixation is tightly regulated in Rhodospirillum rubrum at two different levels: transcriptional regulation of nif expression and posttranslational regulation of dinitrogenase reductase by reversible ADP-ribosylation catalyzed by the DRAT-DRAG (dinitrogenase reductase ADP-ribosyltransferase-dinitrogenase reductase-activating glycohydrolase) system. We report here the characterization of glnB, glnA, and nifA mutants and studies of their relationship to the regulation of nitrogen fixation. Two mutants which affect glnB (structural gene for P(II)) were constructed. While P(II)-Y51F showed a lower nitrogenase activity than that of wild type, a P(II) deletion mutant showed very little nif expression. This effect of P(II) on nif expression is apparently the result of a requirement of P(II) for NifA activation, whose activity is regulated by NH(4)(+) in R. rubrum. The modification of glutamine synthetase (GS) in these glnB mutants appears to be similar to that seen in wild type, suggesting that a paralog of P(II) might exist in R. rubrum and regulate the modification of GS. P(II) also appears to be involved in the regulation of DRAT activity, since an altered response to NH(4)(+) was found in a mutant expressing P(II)-Y51F. The adenylylation of GS plays no significant role in nif expression or the ADP-ribosylation of dinitrogenase reductase, since a mutant expressing GS-Y398F showed normal nitrogenase activity and normal modification of dinitrogenase reductase in response to NH(4)(+) and darkness treatments.

Amino Acid Sequence↗
PubMed2000

Carbon and nitrogen metabolism in Rhizobium.

One of the paradigms of symbiotic nitrogen fixation has been that bacteroids reduce N2 to ammonium and secrete it without assimilation into amino acids. This has recently been challenged by work with soybeans showing that only alanine is excreted in 15N2 labelling experiments. Work with peas shows that the bacteroid nitrogen secretion products during in vitro experiments depend on the experimental conditions. There is a mixed secretion of both ammonium and alanine depending critically on the concentration of bacteroids and ammonium concentration. The pathway of alanine synthesis has been shown to be via alanine dehydrogenase, and mutation of this enzyme indicates that in planta there is likely to be mixed secretion of ammonium and alanine. Alanine synthesis directly links carbon catabolism and nitrogen assimilation in the bacteroid. There is now overwhelming evidence that the principal carbon sources of bacteroids are the C4-dicarboxylic acids. This is based on labelling and bacteroid respiration data, and mutation of both the dicarboxylic acid transport system (dct) and malic enzyme. L-malate is at a key bifurcation point in bacteroid metabolism, being oxidized to oxaloacetate and oxidatively decarboxylated to pyruvate. Pyruvate can be aminated to alanine or converted to acetyl-CoA where it either enters the TCA cycle by condensation with oxaloacetate or forms polyhydroxybutyrate (PHB). Thus regulation of carbon and nitrogen metabolism are strongly connected. Efficient catabolism of C4-dicarboxylates requires the balanced input and removal of intermediates from the TCA cycle. The TCA cycle in bacteroids may be limited by the redox state of NADH/NAD+ at the 2-ketoglutarate dehydrogenase complex, and a number of pathways may be involved in bypassing this block. These pathways include PHB synthesis, glutamate synthesis, glycogen synthesis, GABA shunt and glutamine cycling. Their operation may be critical in maintaining the optimum redox poise and carbon balance of the TCA cycle. They can also be considered to be overflow pathways since they act to remove or add electrons and carbon into the TCA cycle. Optimum operation of the TCA cycle has a major impact on nitrogen fixation.

Alanine↗
PubMed2006

Watering, fertilization, and slurry inoculation promote recovery of biological crust function in degraded soils.

Biological soil crusts are very sensitive to human-induced disturbances and are in a degraded state in many areas throughout their range. Given their importance in the functioning of arid and semiarid ecosystems, restoring these crusts may contribute to the recovery of ecosystem functionality in degraded areas. We conducted a factorial microcosm experiment to evaluate the effects of inoculation type (discrete fragments vs slurry), fertilization (control vs addition of composted sewage sludge), and watering frequency (two vs five times per week) on the cyanobacterial composition, nitrogen fixation, chlorophyll content, and net CO2 exchange rate of biological soil crusts inoculated on a semiarid degraded soil from SE Spain. Six months after the inoculation, the highest rates of nitrogen fixation and chlorophyll a content were found when the biological crusts were inoculated as slurry, composted sewage sludge was added, and the microcosms were watered five times per week. Net CO2 exchange rate increased when biological crusts were inoculated as slurry and the microcosms were watered five times per week. Denaturing gradient gel electrophoresis fingerprints and phylogenetic analyses indicated that most of the cyanobacterial species already present in the inoculated crust had the capability to spread and colonize the surface of the surrounding soil. These analyses showed that cyanobacterial communities were less diverse when the microcosms were watered five times per week, and that watering frequency (followed in importance by the addition of composted sewage sludge and inoculation type) was the treatment that most strongly influenced their composition. Our results suggest that the inoculation of biological soil crusts in the form of slurry combined with the addition of composted sewage sludge could be a suitable technique to accelerate the recovery of the composition and functioning of biological soil crusts in drylands.

Acetylene↗
PubMed1979

Phenotypic reversion of nitrogenase in pleiotropic mutants of Rhizobium meliloti.

In two out of three pleiotropic mutants of Rhizobium meliloti, defective in nitrate reductase induced by amino acid utilization in vegetative bacteria and in symbiotic nitrogen fixation, nitrogenase activity could be restored completely by purines and partially by the amino acids L-glutamate, L-aspartate, L-glutamine, and L-asparagine. The compounds restoring effectiveness in nitrogen fixation did not restore nitrate reductase activity in vegetative bacteria. The restoration of effectiveness supports our earlier conclusion that the mutation is not in the structural gene for a suggested common subunit of nitrogenase and nitrate reductase.

Amino Acids↗
PubMedDate not supplied

[Synthesis and breakdown of poly-beta-hydroxybutyric acid in Rhizobium lupini].

The effect of various substrates on synthesis and decomposition of poly-beta-hydroxybutyric acid (PHBA) was studied in cell suspensions of the effective strain of Rhizobium lupini and in suspensions of the bacteroids of this strain which were isolated from lupine nodules at different growth stages of the plants. Glucose and beta-hydroxybutyrate were found to be the best substrates for synthesis of PHBA in all variants. The content of PHBA in the presence of these substrates increased in suspensions by 2.0 to 2.5 times during ten hours of incubation. In the presence of succinate, PHBA was actively synthesized only by the bacteroids isolated during the stage of active nitrogen fixation (flowering of the plants). In the absence of exogenous substrates, PHBA was decomposed, especially if ammonium ions were present in suspensions. No synthesis of PHBA was registered in the presence of ammonium and glucose, and the rate of PHBA decomposition was high in this case during the stage of active nitrogen fixation.

Ammonia↗
PubMed1999

Studies on the heme and H2-uptake reaction from Azotobacter vinelandii bacterial ferritin.

Bacterial ferritin of Azotobacter vinelandii (AvBF) is directly able to pick electrons up for iron release from or transfer them for storage to a platinum electrode in the absence of mediator or other reducer. The ferritin containing the structure of heme-Co2+ in part shows weakened activity to electrode and decreases the rate of iron release greatly. A reversible reduction process of the ferritin is observed by the spectral change regularly ranging from 310 to 260 nm under mixed gases containing 98% H2 and 2% to O2. The activity of nitrogen fixation from the whole cell of A. vinelandii increases greatly by H2 reduction with potentials ranging from -397 to -425 mV vs. NHE, indicating two important roles of H2-uptake reaction of the ferritin in increasing activity of nitrogen fixation and in supplying iron to synthesize nitrogenase.

Azotobacter vinelandii↗
PubMed2005

Effective symbiosis between Rhizobium etli and Phaseolus vulgaris requires the alarmone ppGpp.

The symbiotic interaction between Rhizobium etli and Phaseolus vulgaris, the common bean plant, ultimately results in the formation of nitrogen-fixing nodules. Many aspects of the intermediate and late stages of this interaction are still poorly understood. The R. etli relA gene was identified through a genome-wide screening for R. etli symbiotic mutants. RelA has a pivotal role in cellular physiology, as it catalyzes the synthesis of (p)ppGpp, which mediates the stringent response in bacteria. The synthesis of ppGpp was abolished in an R. etli relA mutant strain under conditions of amino acid starvation. Plants nodulated by an R. etli relA mutant had a strongly reduced nitrogen fixation activity (75% reduction). Also, at the microscopic level, bacteroid morphology was altered, with the size of relA mutant bacteroids being increased compared to that of wild-type bacteroids. The expression of the sigma(N)-dependent nitrogen fixation genes rpoN2 and iscN was considerably reduced in the relA mutant. In addition, the expression of the relA gene was negatively regulated by RpoN2, the symbiosis-specific sigma(N) copy of R. etli. Therefore, an autoregulatory loop controlling the expression of relA and rpoN2 seems operative in bacteroids. The production of long- and short-chain acyl-homoserine-lactones by the cinIR and raiIR systems was decreased in an R. etli relA mutant. Our results suggest that relA may play an important role in the regulation of gene expression in R. etli bacteroids and in the adaptation of bacteroid physiology.

Adaptation, Physiological↗
PubMed2003

Inhibition of cell division blocks the synthesis of the second nitrogenase (Nif2) in the cyanobacterium Anabaena variabilis.

Anabaena variabilis ATCC 29413 belongs to the cyanobacteria that use a specific cell type, heterocysts, for fixation of atmospheric nitrogen under aerobic conditions. Nitrogen fixation under anaerobic conditions is catalyzed by a Mo-dependent nitrogenase (Nif2) that is expressed in the vegetative cells. We demonstrate here using immunolocalization/light microscopy (LM) that the synthesis of NifH2 is mainly initiated in dividing vegetative cells along the trichomes. Blocking cell division by cephalexin abolished nitrogenase synthesis under anaerobic conditions.

Acetylene↗