• Applied Biological Chemistry
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• v.48 no.3
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• pp.189-200
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• 2005

Biological nitrogen fixation is an important process for academic and industrial aspects. This review will briefly compare industrial and biological nitrogen fixation and cover the characteristics of biological nitrogen fixation studied in Azotobacter vinelandii. Various organisms can carry out biological nitrogen fixation and recently the researches on the reaction mechanism were concentrated on the free-living microorganism, A. vinelandii. Nitrogen fixation, which transforms atmospheric $N_2$ into ammonia, is chemically a reduction reaction requiring electron donation. Nitrogenase, the biological nitrgen fixer, accepts electrons from biological electron donors, and transfers them to the active site, FeMo-cofactor, through $Fe_4S_4$ cluster in Fe protein and P-cluster in MoFe protein. The electron transport and the proton transport are very important processes in the nitrogenase catalysis to understand its reaction mechanism, and the interactions between FeMo-cofactor and nitrogen molecule are at the center of biological nitrogen fixation mechanism. Spectroscopic studies including protein X-ray crystallography, EPR and $M{\ddot{o}}ssbauer$, biochemical approaches including substrate and inhibitor interactions as well as site-directed mutation study, and chemical approach to synthesize the FeMo-cofactor model compounds were used for biological nitrogen fixation study. Recent research results from these area were presented, and finally, a new nitrogenase reaction mechanism will be proposed based on the various research results.

• Korean Journal of Microbiology
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• v.30 no.5
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• pp.391-396
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• 1992

Anabaena variabilis A TCC 29413. a photoautotrophic and nitrogen fixing cyanobacteria. was investigated on the environmental factors regulating the growth and nitrogen lixation activity. A good growth of cyanobacteria] cells was observed due to nitrogen t1xation by the heterocyst differentiation in nitrogen free Allen and Arnon (]/8) medium. The nitrogenase activity was appeared to be in proportion to the cell growth lor 6 days then drastically decreased in the later growth period when the nitraTe was accumulated to high level in the culture to cause the inhibition. The optima] conditions lilr the cell growth and nitrogenase activity of A. varillbili.l were anaerobic. IO.OO0 lux. $30^{\circ}C$ and pH 8 with the nitrogen Cree minimal medium. The activity was significantly inhihited by the low concentrations of ammonium and nitrate. but was stimulated b) the ]ow Ieve] of phosphate and carbonate sources. The treatments of several toxic heavy metals showed strong inhibition of the cell growth and nitrogenase activity by O.3~10 ppm in the order of $Hg^{2+}$ > $Cd^{2+}$ > $Co^{2+}$ > $Zn^{2+}$ > $Ph^{2+}$, and the concentrations for 50% inhibition of the maximum activity were 0.41. 0.47. 0.5 L 0.66 and 8.1 ppm. respectively. The addition of carbohydrates (0.5~ 1.0%) in the dark condition stimulated the growth and activity in the order of sucrose > fructose > glucose.

• Journal of Microbiology and Biotechnology
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• v.16 no.1
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• pp.52-56
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• 2006

The effect of bensulfuron-methyl on the nontargeted cyanobacteria was greater on A. variabillis than N. commune. Both A. variabillis and N. commune were initially able to utilize low concentrations of the herbicide, bensulfuron-methyl, whereas higher concentrations of bensulfuron-methyl or the hydrolyzed products of the herbicide were found to be toxic. Growth and photosynthesis inhibitions of over $50\%$ were observed, when 8 to 10 ppm of the herbicide was applied. Nitrogenase activities of the cyanobacteria were decreased by $94-98\%$ in A. variabillis and by $85-86\%$ in N. commune after 24 h of incubation with 10 ppm and 20 ppm of bensulfuron-methyl. Nitrogenase activities were also inhibited by the addition of ammonium salts as low as 0.05 mM. Furthermore, the toxic effect of the herbicide was the highest at pH 4-6, showing approximately $42-60\%$ toxicity, whereas much lower toxicity $(9-28\%)$ was observed at higher pH of 7-10, due to base-catalyzed hydrolysis of bensulfuron-methyl.

• Environmental Engineering Research
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• v.20 no.1
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• pp.41-50
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• 2015

The cyanobacterium Anabaena doliolum Ind1 isolated from a coal mining site was tested for removal of cadmium at optimum pH 7.0 and temperature $25^{\circ}C$. The organism recorded high percentage of metal removal (92-69%) within seven days of exposure to 0.5-2.0 ppm cadmium. Biosorption onto the cell surface was the primary mode of metal removal. Fourier transform infrared spectroscopy (FTIR) established hydroxyl, amides, carboxyl, sulphate and carbonyl groups to be the major functional groups on the cell surface involved in cadmium binding. Cellular ultrastructure and a range of vital physiological processes (i.e., photosynthetic pigments, respiration, photosynthesis, heterocyst frequency and nitrogenase activity) remained unaffected upon 0.5 ppm treatment; higher concentrations of cadmium exerted visible adverse effects. Amongst the five photosynthetic pigments tested, phycocyanin was the most targeted pigment (inhibition was 15-89%). Both respiration and photosynthetic activities were inhibited by cadmium with more severe effect seen on respiration. 2.0 ppm cadmium exposure also had drastic negative effect on nitrogenase activity (87% decreased).

• Journal of Environmental Science International
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• v.21 no.9
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• pp.1139-1147
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• 2012

To find out the growth conditions for the maximum activity of nitrogenase which catalyzes nitrogen fixation in Rhodobacter sphaeroides, the promoter activities of nifA and nifH were analyzed and the results indicated that expression of both nifA and nifH was increased in response to deprivation of both O2 concentration and nitrogen source. The nifA mutant was constructed by deleting the gene to investigate the effect of NifA, the transcriptional regulator, on the nifH and nifA expression in R. sphaeroides. Analysis of expression of nif genes using the nifA::lacZ and nifH::lacZ fusions in the nifA mutant revealed that NifA acts as a positive activator for nifH and an autoregulator in its own expression. The promoter activities of nifA and nifH in the prrA mutant grown under anaerobic and ${NH_4}^+$-free conditions were derepressed, comparing with those of the wild-type grown under the same conditions, indicating that the prrA product acts as a positive regulator in expression of nifA and nifH.

• Korean Journal of Microbiology
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• v.29 no.6
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• pp.408-415
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• 1991

TOL plsmid size of Flavobacterium odoratum SUB53 was estimated as 83 Md and the optimum concentration of m-toluate degradation by TOL plasmid was 5 mM. $H_{2}$ production by Rhodopseudomonas sphaeroides KCTC1425 was largely dependent on nitrogenase activity and showed the highest at 30 mM malate with 7 mM glutamate as nitrogen source. Nitrogenase activities were inhibited by 0.3 mM $NH_{4}^{+}$ions, to be appeared the decrease of $H_{2}$ production. Conjugation of TOL plasmids from F. odoratum SUB53 and Pseudomonas putida mt-2 to R. sphaeroides showed the optimum at the exponential stage of recipient cells in presence of helper plasmid pRK2013. According to the investigation of catechol-1,2-oxygenase (C-1, 2-O) and catechol-2,3-oxygenase (C-2,3-O) activities of R. sphaeroides C1 (TOL SUB53) and C2 (TOL mt-2), the gene for C-2,3-O is located on TOL plasmid and gene for C-1, 2-O on the chromosome of R. sphaeroides. m-Toluate was biodegraded by TOL plasmid in R. sphaeroides C1 and C2, presumably to be produced $H_{2}$ gas from the secondary metabolites of m-toluate.e.

• Korean Journal of Microbiology
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• v.27 no.1
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• pp.21-26
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• 1989

The chromosomal DNA of Klebsiella penumoniae KNF1 was partially digested with HindIII. pBR322 was completely digested with same enzyme with additional alkaline phosphatase treatment. Both DNA samples were ligated and transformed into E. coli KO60. Single coloneis of the transformed cells were isolated on N0free agar media. These colonies were ampicillin-resistant and tetracycline-sensitive. When the plasmids in transformants were cured, nitrogen-fixing activities were lost. Therefore, these transformants harbored recombinant plasmid including nif genes of K. pneumoniae. Nitrogenase activity of transformant was higher than or the same as that of K. pneumoniae KNF1.

• Mycobiology
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• v.32 no.4
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• pp.173-178
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• 2004

Fifteen Rhizobium spp. from nodules of 6 common pulses collected from 6 districts of Assam were studied for their infectivity, intrinsic antibiotic resistance, nitrogenase activity and effect of dual inoculation with two native Arbuscular Mycorrhizal Fungi viz. Glomus mosseae(GM) and Gigaspora gilmarie(GG). Out of the 15 isolates 9 were found nodulation positive and 6 of them(AR1, BR8, BR12, AR10, UR10 & GR21) were subjected to intrinsic antibiotic sensitivity test of which AR1 showed resistance against all the 9 test antibiotics. Isolates AR1 and GR21 showed the highest(4.25 mole, $gm^{-1}hour^{-1}$) and the lowest(1.05 mole, $gm^{-1}hour^{-1}$) nitrogenase activity respectively. In Most Probable Number count, the maximum Rhizobium population $5.8{\times}10^5$, was found in both Blackgram and Greengram variety of pulses. The maximum dry weight of nodules(3.14 g), dry weight of shoot(10.08 g), nitrogen content(7.68 mg, $plant^{-1}$), chlorophyll content(1.89 mg, $g^{-1}$), phosphorus content of shoot(6.17 mg, $g^{-1}$) and yield(535.67 kg, $Ha^{-1}$) were found when AR1 dually inoculated with GM in Blackgram.

• Journal of Plant Biology
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• v.37 no.3
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• pp.387-394
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• 1994

Effects of environmental factors of light, temperature, nitrogen sources and water stress were analyzed quantitatively on the nodule formation and nitrogen fixation activity of autumn olive plant (Elaeagnu$ umbellala Thunb.) during the seedling growth. Seedlings showed the maximum nitrogenase activity of $72.5\;\mu\textrm{M}\;C_2H_4{\cdot}g\;fr\;wt\;nodule^{-1}{\cdot}h^{-1}$ in the early nodulation stage. The relative growth rate and T/R ratio changed from $1.60%{\cdot}d^{-1}$ and 1.12 in the earlier stage to $3.75%{\cdot}d^{-1}$ and 2.31 in the later stage, respectively. light conditions of 20-25, 1015 and 4-6% resulted in decreases of 41, 54 and 71% of the nitrogenase activity, respectively. Nodules incubated in 15, 20, 25 and $30^{\circ}C$ showed the activities of 5.4, 24.7, 51.6 and $58.5\;\mu\textrm{M}\;C_2H_4{\cdot}g\;fr\;wt\;nodule^{-1}{\cdot}h^{-1}$ respectively. Pretreatment with low temperature ($15^{\circ}C$) followed incubation at $30^{\circ}C$ attained higher nitrogenase activity ($66.5\;\mu\textrm{M}\;C_2H_4{\cdot}g\;fr\;wt\;nodule^{-1}{\cdot}h^{-1}$) than that with higher temperature ($35^{\circ}C$). The oxygen pressure above 16 kPa is necessary for saturation of the nodule activity, but the activity was inhibited severely by physical impact such as the exision or isolation of nodules from the root. The relative activities of early nodules grown in pH 5.5, 6.5 and 8.0 were 89, 100 and 40% and those grown in 1 and 3 mM of $NO_3\;and\;NH_4$ were 6, 1 and 68, 50%, respectively. Watering levels of 20, 50 and 100 mL during the seedling growth resulted in 35, 120 and 8 mg of nodule formation and 33.6, 58.4 and $8.4\;\mu\textrm{M}\;C_2H_4{\cdot}g\;fr\;wt\;nodule^{-1}{\cdot}h^{-1}$ of the nitrogenase activity, respectively. Water stress with 86% decrease of soil water content caused temporary wilting point of leaf and a complete disappearance of nitrogenase activity of nodules, though the water content and transpiration rate in plant were reduced to 90 and 53%, respectively.tively.

• Korean Journal of Soil Science and Fertilizer
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• v.27 no.2
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• pp.136-146
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• 1994

To research the effect of chemotaxis of Rhizobia toward the root exudate on nitrogen fixing ability in soybean Rhizobia symbiosis system. Root exudate from seedlings of Glycine max. L was collected aseptic conditions. B. japonicum KCTC 2422 induced the formation of symbiotic nitrogen fixing nodules on the root of soybean plant and possessed motility and chemotaxis toward the 2mM proline. LPN-100 mutant was $Nod^-$, $Che^+$, and LPN-101 was $Che^-$, $Nod^+$ strains. Physiological properties of mutants were similar to parent strain. The crude root exudate was tested for its chemotactic ability using the capillary tube method. Chemotactic responses of RCR 3407 toward crude root exudate were 2.2, 2.6, 2.9, those of KCTC 2422 were 2.3, 2.9, 3.0, respectively. The crude root exudate was fractionated into neutral, cationic and anionic fractions. Chemotactic responses of KCTC 2422 was least with anionic fraction, most with neutral and intermediate with cationic fraction. B. japonicum KCTC 2422 was attracted by carbohydrates, amino acids and carboxylic acid. Carbohydrates and amino acids were good chemoattractants and carboxylic acids were intermediate chemoattractants. The peak concentration was $10^{-3}M$ for ribose, glucose, glutamine, aspartic acid and carboxylic acids, with exception of xylose, arabinose, tryptophan, which elicited maximum responses at $10^{-4}M$. The formation of nodules and nitrogenase activity of soybean inoculated with KCTC 2422 was determined in 7days after inoculation, and those of LPN-101 was detected in 15days after inoculation, but LPN-100 didn't form of nodules in soybean plants.