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PubMed1981

Enzymes of Purine Biosynthesis and Catabolism in Glycine max: I. COMPARISON OF ACTIVITIES WITH N(2) FIXATION AND COMPOSITION OF XYLEM EXUDATE DURING NODULE DEVELOPMENT.

During the period examined from 12 to 63 days after planting, the ureides, allantoin and allantoic acid, were the predominant nitrogenous solutes in the xylem exudate of soybeans (Glycine max [L.]) growing solely on symbiotically fixed nitrogen, accounting for approximately 60% and greater than 95% of the total nitrogen in the xylem exudate before and after the onset of active nitrogen fixation, respectively. For plants between 18 and 49 days of age, the apparent rate of ureide export estimated from concentrations of ureides in xylem exudate collected over a period of one hour was closely related to the rate of nitrogen fixation estimated from measurements of C(2)H(2) reduction by nodulated root systems. After this time, the apparent rate of ureide export per plant continued to increase, reaching a maximum value at day 63 of 12 micromoles per plant per hour, even though the rate of C(2)H(2) reduction per plant declined approximately four-fold. The most probable pathway for the biosynthesis of ureides involves the catabolism of purines. The levels of phosphoribosylpyrophosphate (PRPP) synthetase, which catalyzes the formation of the PRPP required for purine synthesis, increased in parallel with the rates of nitrogen fixation (C(2)H(2)) from day 18 reaching a maximum value of 13.9 micromoles per plant per hour at day 49, and then both activities declined rapidly. During the period of active nitrogen fixation the ratio of PRPP synthesis estimated from measurements of PRPP synthetase activity in cell-free extracts to the apparent rate of ureide export was between 1 and 2. The activities of the enzymes of purine catabolism, xanthine dehydrogenase, uricase, and allantoinase, increased in parallel with the increases in nodule mass and the export of ureides with maximum activities of 13, 119, and 79 micromoles per plant per hour, corresponding with apparent rates of ureide export in the range of 9.5 to 11.9 micromoles per plant per hour. These results demonstrate that there is a close association between nitrogen fixation, PRPP synthetase activity, and ureide export in soybeans and support the proposal that recently-fixed nitrogen is utilized in the de novo synthesis of purines which are subsequently catabolized to produce the ureides.

Journal Article↗
PubMed1976

Pathway of nitrogen metabolism after fixation of 13N-labeled nitrogen gas by the cyanobacterium, Anabaena cylindrica.

Methods have been developed for identifying the pathway of assimilation of N2-derived nitrogen. The products of fixation of 13N-labeled nitrogen gas ([13N]N2), and the distribution of 13N within glutamine, were determined after short periods of labeling (approximately 1 to 120 s) and also in pulse-chase experiments. Ammonia, the amide nitrogen of glutamine, and the alpha-amino nitrogen of glutamate, in that order, were the first observed products of fixation of N2 by the cyanobacterium (blue-green alga), Anabaena cylindrica. This sequence of the formation of nitrogenous products was confirmed by the use of inhibitors. The presence of 1 mM methionine sulfoximine permitted continued formation of 13NH3, while virtually preventing 13N-labeling of amino acids. In the presence of 1 mM azaserine, glutamine was labeled, but not other amino acids. Our observations demonstrate unequivocally that N2-derived nitrogen fixed by this organism is metabolized initially by the glutamine synthetase/glutamate synthase pathway.

Cyanobacteria↗
PubMed1995

Attenuation of Symbiotic Effectiveness by Rhizobium meliloti SAF22 Related to the Presence of a Cryptic Plasmid.

Several wild-type strains of Rhizobium meliloti isolated from alfalfa nodules exhibited different plasmid profiles, yet did not differ in growth rate in yeast-mannitol medium, utilization of 43 different carbon sources, intrinsic resistance to 14 antibiotics, or detection of 16 enzyme activities. In contrast, three measures of effectiveness in symbiotic nitrogen fixation with alfalfa (shoot length, dry weight, and nitrogen content) indicated that R. meliloti SAF22, whose plasmid profile differs from those of the other strains tested, is significantly less effective than other wild-type strains in symbiotic nitrogen fixation. Light microscopy of nodules infected with strain SAF22 showed an abnormal center of nitrogen fixation zone III, with bacteria occupying a smaller portion of the infected host cells and vacuoles occupying a significantly larger portion of adjacent uninfected host cells. In contrast, the effective nodules infected with other wild types or plasmid pRmSAF22c-cured segregants of SAF22 did not display this cytological abnormality.

Journal Article↗
PubMed2004

Regional inventory of soil surface nitrogen balances in Indian agriculture (2000-2001).

Nitrogen regulates several ecological and biogeochemical processes and excess reactive nitrogen in the environment can lead to pollution problems, including the deterioration of air quality, disruption of forest processes, acidification of lakes and streams, and degradation of coastal waters. Much of the excess nitrogen inputs are related to food and energy production. An important step to understanding the sources of nitrogen and ultimately defining solutions to excess nitrogen is to describe the geographic distribution of agricultural nitrogen contributions from different regions. In this study, soil surface nitrogen loads were quantified for different states of India for the period 2000-2001. Nearly 35.4 Tg of nitrogen has been estimated as inputs from different sources, with output nitrogen from harvested crops of about 21.20 Tg. The soil surface nitrogen balance, estimated as inputs minus outputs, is found to be about 14.4 Tg surplus from the agricultural land of India. Livestock manure constituted a major percentage of total inputs (44.06%), followed by inorganic fertilizer (32.48%), atmospheric deposition (11.86%) and nitrogen fixation (11.58%). Nitrogen balance varied from deficit to surplus for different states. The highest nitrogen surplus was found in Uttar Pradesh (2.50 Tg) followed by Madhya Pradesh (1.83 Tg), Andhra Pradesh (1.79 Tg), etc. A negative nitrogen balance was found in Orissa (-0.01 Tg), Andaman Nicobar Islands (-0.32 Tg) and for some of the northeastern states. Major fertilizer consumption states were found to be Tamilnadu (204 kg/ha), Haryana (132 kg/ha), Punjab (148 kg/ha), followed by others. Similarly, nitrogen inputs from total livestock excretions were found to be high for Kerala (616 kg/ha), Jammu and Kashmir (389 kg/ha), Tamil Nadu (338 kg/ha), etc. The average nitrogen surplus of about 54 kg/ha observed for the agricultural land of the entire country of India is comparatively higher than the average surplus of about 31 kg/ha reported for European countries. These results, obtained from nutrient mass balance calculations, will be useful to formulate nutrient management plans relating to fertilizer usage, livestock management and for adopting some best management strategies at a state level in India.

Agriculture↗
PubMed2003

Signal transduction by the global regulator RegB is mediated by a redox-active cysteine.

All living organisms alter their physiology in response to changes in oxygen tension. The photosynthetic bacterium uses the RegB-RegA signal transduction cascade to control a wide variety of oxygen-responding processes such as respiration, photosynthesis, carbon fixation and nitrogen fixation. We demonstrate that a highly conserved cysteine has a role in controlling the activity of the sensor kinase, RegB. In vitro studies indicate that exposure of RegB to oxidizing conditions results in the formation of an intermolecular disulfide bond and that disulfide bond formation is metal-dependent, with the metal fulfilling a structural role. Formation of a disulfide bond in vitro is also shown to convert the kinase from an active dimer into an inactive tetramer state. Mutational analysis indicates that a cysteine residue flanked by cationic amino acids is involved in redox sensing in vitro and in vivo. These residues appear to constitute a novel 'redox-box' that is present in sensor kinases from diverse species of bacteria.

Amino Acid Sequence↗
PubMed1980

Genetic analysis of a pleiotropic mutant of Klebsiella pneumoniae affected in nitrogen metabolism.

Genetic and reversion analyses of a thermosensitive pleiotropic mutant strain of Klebsiella pneumoniae with defects in nitrogen fixation and nitrogen metabolism have shown that the pleiotropie behaviour of mutant is due to a single mutation in a gene designated nim. This gene is contransducible with trp at a frequency of about 30% (using bacteriophage P1) and with cys at a frequency of about 14%. The gene order is cys, trp, nim. The defect in the nim mutant is complemented by the E. coli F' element, F'148.

Chromosome Mapping↗
PubMed1987

Gabaculine Inhibition of Chlorophyll Biosynthesis and Nodulation in Phaseolus lunatus L.

Gabaculine (3-amino-2,3-dihydrobenzoic acid) was an inhibitor of in vivo chlorophyll biosynthesis in lima bean (Phaseolus lunatus L. cv Henderson). When applied to roots of 9-day-old plants, 10 micromolar gabaculine was sufficient to terminate biosynthesis of new chlorophyll. The trifoliolate leaves which emerged after gabaculine treatment were yellow. Gabaculine-treated plants had slightly lower dry weights; yet, overall plant size showed very little change. Chlorophyll fluorescence induction kinetics and CO(2) exchange measurements were used to monitor both immediate and long-term effects of gabaculine on photosynthesis. A lowered rate of the decline from the maximum level of fluorescence was observed after 10 hours for nitrate-supplemented plants, and all treated plants showed a slightly increased level of original fluorescence after 6 days. No change was observed in the rate of photosynthesis by unifoliolate leaves. The trifoliolate leaves, though not able to photosynthesize, were able to continue respiration. This suggested that heme biosynthesis for mitochondrial cytochromes was not abolished. In untreated lima bean, root nodules were induced by Rhizobium sp. 127E15. Following gabaculine treatment, root nodules formed, but were largely ineffective in nitrogen fixation. Nodule dry weight, nitrogen fixation activity, and leghemoglobin content were decreased by gabaculine.

Journal Article↗
PubMed1992

Diversity among Rhizobia Effective with Robinia pseudoacacia L.

The diversity of rhizobia that form symbioses with roots of black locust (Robinia pseudoacacia L.), an economically important leguminous tree species, was examined by inoculating seedling root zones with samples of soil collected from the United States, Canada, and China. Bacteria were isolated from nodules, subcultured, and verified to be rhizobia. The 186 isolates varied significantly in their resistance to antibiotics and NaCl, their growth on different carbohydrates, and their effect on the pH of culture media. Most isolates showed intermediate antibiotic resistance, the capacity to use numerous carbohydrates, and a neutral to acid pH response. Isolates had greater similarity within sampling locations than among sampling locations. The isolates were grouped by using numerical taxonomy techniques, and representative strains of 37 groups were selected. The mean generation times of these isolates ranged from 3 to 9 h, and the protein profile of each of the 37 isolates was unique. Nitrogen fixation, total nitrogen accumulation, and plant growth varied significantly among black locust seedlings inoculated with the representative isolates. We conclude that great variation exists among Rhizobium spp. that nodulate black locust, and selection of strains for efficiency of the symbiotic association appears possible.

Journal Article↗
PubMed1987

Comparative organization of nitrogen fixation-specific genes from Azotobacter vinelandii and Klebsiella pneumoniae: DNA sequence of the nifUSV genes.

In the facultative anaerobe Klebsiella pneumoniae 17 nitrogen fixation-specific genes (nif genes) have been identified. Homologs to 12 of these genes have now been isolated from the aerobic diazotroph Azotobacter vinelandii. Comparative studies have indicated that these diverse microorganisms share striking similarities in the genetic organization of their nif genes and in the primary structure of their individual nif gene products. In this study the complete nucleotide sequence of the nifUSV gene clusters from both K. pneumoniae and A. vinelandii were determined. These genes are identically organized on their respective genomes, and the individual genes and their products exhibit a high degree of interspecies sequence homology.

Aerobiosis↗
PubMed2003

Isolation and symbiotic characterization of transposon Tn5-induced arginine auxotrophs of Sinorhizobium meliloti.

Seventeen arginine auxotrophic mutants of Sinorhizobium meliloti Rmd201 were isolated by random transposon Tn5 mutagenesis using Tn5 delivery vector pGS9. Based on intermediate feeding studies, these mutants were designated as argA/argB/argC/argD/argE (ornithine auxotrophs), argF/argI, argG and argH mutants. The ornithine auxotrophs induced ineffective nodules whereas all other arginine auxotrophs induced fully effective nodules on alfalfa plants. In comparison to the parental strain induced nodule, only a few nodule cells infected with rhizobia were seen in the nitrogen fixation zone of the nodule induced by the ornithine auxotroph. TEM studies showed that the bacteroids in the nitrogen fixation zone of ornithine auxotroph induced nodule were mostly spherical or oval unlike the elongated bacteroids in the nitrogen fixation zone of the parental strain induced nodule. These results indicate that ornithine or an intermediate of ornithine biosynthesis, or a chemical factor derived from one of these compounds is required for the normal development of nitrogen fixation zone and transformation of rhizobial bacteria into bacteroids during symbiosis of S. meliloti with alfalfa plants.

Arginine↗