scholarly journals Artificial activation of nif gene expression in nodule bacteria Ex Planta

2019 ◽  
Vol 17 (2) ◽  
pp. 35-42
Author(s):  
Andrey K. Baymiev ◽  
Roman S. Gumenko ◽  
Anastasiya A. Vladimirova ◽  
Ekaterina S. Akimova ◽  
Zilya R. Vershinina ◽  
...  
Keyword(s):  
Nitrogen Fixation ◽  
Nitrogenase Activity ◽  
Regulatory Protein ◽  
Inducible Promoter ◽  
Nitrogen Fixing ◽  
Nodule Bacteria ◽  
Free Living ◽  
Living State ◽  
Artificial Activation ◽  
Nifa Gene

Background. Rhizobia are the most effective nitrogen-fixing organisms that can fix nitrogen only in symbiosis with leguminous plants. The general transcriptional activator of nitrogen fixation genes in diazotrophic bacteria is NifA. In this work, the possibility of modifying the regulation of nitrogen fixation in the nodule bacteria Mesorhizobium, Ensifer and Rhizobium was studied by introducing an additional copy of the nifA gene into the bacterial genomes during the regulation of induced bacterial promoters. Materials and methods. A series of expression genetic constructs with NifA genes of nodule bacteria strains under the control of an inducible promoter Pm were created. The resulting constructs were transformed into strains of nodule bacteria. The obtained recombinant strains were investigated for the appearance of their nitrogen-fixing activity in the free-living state. Results. It was shown that the expression of nifA in recombinant cells of all three genera of bacteria leads to the appearance of insignificant nitrogenase activity. At the same time, the level of nitrogenase activity does not have a correlation with the level of expression of the introduced nifA gene, which, most likely, is a consequence of the multilevel regulation of nitrogen fixation. Conclusion. The possibility of artificial activation of nitrogenase activity in nodule bacteria in the free-living state by introducing the NifA regulatory protein gene into bacteria was shown.

scholarly journals nifDK Clusters Located on the Chromosome and Megaplasmid of Bradyrhizobium sp. Strain DOA9 Contribute Differently to Nitrogenase Activity During Symbiosis and Free-Living Growth

2016 ◽  
Vol 29 (10) ◽  
pp. 767-773 ◽  
Author(s):  
Jenjira Wongdee ◽  
Pongpan Songwattana ◽  
Nico Nouwen ◽  
Rujirek Noisangiam ◽  
Joel Fardoux ◽  
...  
Keyword(s):  
Enzyme Activity ◽  
Nitrogen Fixation ◽  
Nitrogenase Activity ◽  
Transcriptional Analysis ◽  
Main Role ◽  
Free Living ◽  
Living State ◽  
Bradyrhizobium Sp ◽  
Reporter Strains ◽  
Free Living Conditions

Bradyrhizobium sp. strain DOA9 contains two copies of the nifDK genes, nifDKc, located on the chromosome, and nifDKp, located on a symbiotic megaplasmid. Unlike most rhizobia, this bacterium displays nitrogenase activity under both free-living and symbiotic conditions. Transcriptional analysis using gusA reporter strains showed that both nifDK operons were highly expressed under symbiosis, whereas nifDKc was the most abundantly expressed under free-living conditions. During free-living growth, the nifDKp mutation did not affect nitrogenase activity, whereas nitrogenase activity was drastically reduced with the nifDKc mutant. This led us to suppose that nifDKc is the main contributor of nitrogenase activity in the free-living state. In contrast, during symbiosis, no effect of the nifDKc mutation was observed and the nitrogen-fixation efficiency of plants inoculated with the nifDKp mutant was reduced. This suggests that nifDKp plays the main role in nitrogenase enzyme activity during symbiosis. Together, these data suggest that Bradyrhizobium sp. strain DOA9 contains two functional copies of nifDK genes that are regulated differently and that, depending on their lifestyle, contribute differently to nitrogenase activity.

scholarly journals Azorhizobium caulinodans PIIand GlnK Proteins Control Nitrogen Fixation and Ammonia Assimilation

1999 ◽  
Vol 181 (8) ◽  
pp. 2655-2658 ◽  
Author(s):  
Nathalie Michel-Reydellet ◽  
P. Alexandre Kaminski
Keyword(s):  
Gene Expression ◽  
Nitrogen Fixation ◽  
Nitrogenase Activity ◽  
Ammonia Assimilation ◽  
Ammonium Excretion ◽  
Azorhizobium Caulinodans ◽  
Wild Type ◽  
Free Living ◽  
Living State ◽  
High Level

ABSTRACT We herein report that Azorhizobium caulinodansPII and GlnK are not necessary for glutamine synthetase (GS) adenylylation whereas both proteins are required for complete GS deadenylylation. The disruption of both glnB andglnK resulted in a high level of GS adenylylation under the condition of nitrogen fixation, leading to ammonium excretion in the free-living state. PII and GlnK also controllednif gene expression because NifA activated nifHtranscription and nitrogenase activity was derepressed in glnB glnK double mutants, but not in wild-type bacteria, grown in the presence of ammonia.

Nitrogen fixation

1976 ◽  
Vol 274 (934) ◽  
pp. 341-358 ◽  
Keyword(s):  
Nitrogen Fixation ◽  
Metabolic Regulation ◽  
Higher Plants ◽  
Focal Point ◽  
Nitrogenase Activity ◽  
Nitrogen Fixing ◽  
Blue Green Algae ◽  
International Biological Programme ◽  
The 1960S ◽  
Biological Programme

The International Biological Programme served as a focal point for studies on biological nitrogen fixation during the 1960s. The introduction of the acetylene reduction technique for measuring nitrogenase activity in the field led to estimates becoming available of the contribution of lichens, blue-green algae, nodulated non-legumes and bacterial-grass associations, as well as of legumes. Other studies carried out on the physiology and biochemistry of the process led to the eventual purification and characterization of the nitrogenase enzyme. These studies, collectively, provided the springboard for current work, so essential in view of the present energy crisis, on how to increase the use and efficiency of nitrogen-fixing plants, on the metabolic regulation of the nitrogenase enzyme and on the genetics of the nitrogen-fixing process, both in higher plants and in free-living micro-organisms.

Research on Improvement of Nitrogenase Activity of Free-Living Nitrogen-Fixing Bacteria Using DBD Plasma

2013 ◽  
Vol 671-674 ◽  
pp. 2674-2678 ◽  
Author(s):  
Yan Yun Zhu ◽  
Xiao Li Zhu ◽  
Fang She Yang
Keyword(s):  
Rhizosphere Soil ◽  
Barrier Discharge ◽  
Nitrogenase Activity ◽  
Nitrogen Fixing ◽  
Bacterial Strains ◽  
Dbd Plasma ◽  
Nitrogen Fixing Bacteria ◽  
Free Living ◽  
Reduction Assay ◽  
Rrna Sequencing

Nitrogen-fixing bacteria were screened from the rhizosphere soil of plants in Shaanxi in China. 36 free-living nitrogen-fixing bacterial strains were isolated and their nitrogenase activity were determined by acetylene reduction assay (ARA), two strains named FLNB03 and FLNB09 with higher nitrogenase activity were isolated and identified by 16S rRNA sequencing. The datum showed that FLNB03 was similar to Acinetobacter and their similarity reached 99%, FLNB09 was similar to Agrobacterium sp. and their similarity reached 99%. Then both of them were treated using Dielectric Barrier Discharge (DBD) plasma for mutation and their mutants called FLNB03-2 and FLNB09-3 were obtained. The nitrogenase activity of FLNB03-2 was 0.61±0.10 nmol•107cfu-1•h-1, and that of FLNB09-3 was 0.40±0.05 nmol•107cfu-1•h-1, their nitrogenase activity increased by 22.00% and 14.29% than their original bacteria respectively. FLNB03-2 and FLNB09-3 might be used as microbial fertilizer.

Rhizobium meliloti exopolysaccharides: genetic analyses and symbiotic importance

1991 ◽  
Vol 19 (3) ◽  
pp. 636-644 ◽  
Author(s):  
T. Lynne Reuber ◽  
Jason Reed ◽  
Jane Glazebrook ◽  
M. Alexandra Glucksmann ◽  
Dianne Ahmann ◽  
...  
Keyword(s):  
Rhizobium Meliloti ◽  
Nodule Development ◽  
Nitrogen Fixing ◽  
Free Living ◽  
Genetic Analyses ◽  
Living State ◽  
Meliloti Strain

Summary Genetic experiments have indicated that succinoglycan (EPS I), the acidic Calcofluor-binding exopolysaccharide, of the nitrogen-fixing bacterium Rhizobium meliloti strain Rm1021 is required for nodule invasion and possibly for later events in nodule development on alfalfa and other hosts. Fourteen exo loci on the second megaplasmid have been identified that are required for, or affect, the synthesis of EPS I. Mutations in certain of these loci completely abolish the production of EPS I and result in mutants that form empty Fix- nodules. We have identified two loci, exoR and exoS, that are involved in the regulation of EPS I synthesis in the free-living state. Certain exo mutations which completely abolish EPS I production are lethal in an exoR95 or exoS96 background. Histochemical analyses of the expression of exo genes during nodulation using exo :: TnphoA fusions have indicated that the exo genes are expressed most strongly in the invasion zone. In addition, we have discovered that R. meliloti has a latent capacity to synthesize a second exopolysaccharide (EPS II) that can substitute for the role(s) of EPS I in nodulation of alfalfa but not of other hosts. Possible roles for exopolysaccharides in symbiosis are discussed.

NON-SYMBIOTIC NITROGEN FIXATION IN SOME QUEBEC SOILS

1965 ◽  
Vol 11 (1) ◽  
pp. 29-38 ◽  
Author(s):  
P-C. Chang ◽  
R. Knowles
Keyword(s):  
Nitrogen Fixation ◽  
Symbiotic Nitrogen Fixation ◽  
Soil Types ◽  
Anaerobic Incubation ◽  
Nitrogen Fixing ◽  
Free Living ◽  
Aerobic Conditions ◽  
Bacterial Counts ◽  
Facultatively Anaerobic ◽  
Nitrogen Fixers

The occurrence of free-living nitrogen fixers, the potential for nitrogen fixation, and the correlation between the nitrogen-fixing capacities of the soils and bacterial counts were studied using representative Quebec soils.Clostridium occurred more frequently than did Azotobacter. Studies with N15showed that nitrogen fixation was more frequent under anaerobic than under aerobic conditions in all the soil types studied in their unamended state. The addition of glucose stimulated nitrogen fixation. During anaerobic incubation, nitrogen fixation was found to be correlated significantly with the increase in numbers of both total aerobes and Clostridia. The results suggested that facultatively anaerobic nitrogen fixers, and aerobic nitrogen fixers other than Azotobacter, were present.

Genetics of plant-microbe nitrogen-fixing symbiosis

1985 ◽  
Vol 85 (3-4) ◽  
pp. 239-252
Author(s):  
G. C. Machray ◽  
W. D. P. Stewart
Keyword(s):  
Nitrogen Fixation ◽  
Genetic Analysis ◽  
Symbiotic Nitrogen Fixation ◽  
Model System ◽  
Present Knowledge ◽  
Nitrogen Fixing ◽  
Free Living ◽  
Genetic Level ◽  
And Control ◽  
Control Of Expression

SynopsisA wide variety of plant-microbe nitrogen-fixing symbioses which include cyanobacteria as the nitrogenfixing partner exist. While some information has been gathered on the biochemical changes in the cyanobacterium upon entering into symbiosis, very little is known about the accompanying changes at the genetic level. Much of our present knowledge of the organisation and control of expression of nitrogenfixation (nif) genes is derived from studies of the free-living diazotroph Klebsiella pneumoniae. This organism thus provides a model system and source of experimental material for the genetic analysis of symbiotic nitrogen fixation. We describe the use of cloned K. pneumoniae genes for nitrogen fixation and its regulation in the genetic analysis' of nitrogen fixation in cyanobacteria which can enter into symbiosis with plants. These studies reveal some dissimilarities in the organisation of nif genes and raise questions as to the genetic control of nitrogen fixation in symbiosis.

scholarly journals Poly-β-Hydroxybutyrate Turnover inAzorhizobium caulinodans Is Required for Growth and AffectsnifA Expression

1998 ◽  
Vol 180 (19) ◽  
pp. 5070-5076 ◽  
Author(s):  
Karine Mandon ◽  
Nathalie Michel-Reydellet ◽  
Sergio Encarnación ◽  
P. Alexandre Kaminski ◽  
Alfonso Leija ◽  
...  
Keyword(s):  
Nitrogenase Activity ◽  
Constitutive Expression ◽  
Reducing Power ◽  
Azorhizobium Caulinodans ◽  
Free Living ◽  
Nucleotide Pools ◽  
Living State ◽  
Mutant Strains ◽  
Growth Properties ◽  
Phb Synthase

ABSTRACT Azorhizobium caulinodans is able to fix nitrogen in the free-living state and in symbiosis with the tropical legumeSesbania rostrata. The bacteria accumulate poly-β-hydroxybutyrate (PHB) under both conditions. The structural gene for PHB synthase, phbC, was inactivated by insertion of an interposon. The mutant strains obtained were devoid of PHB, impaired in their growth properties, totally devoid of nitrogenase activity ex planta (Nif−), and affected in nucleotide pools and induced Fix− nodules devoid of bacteria. The Nif− phenotype was the consequence of the lack ofnifA transcription. Nitrogenase activity was partially restored to a phbC mutant by constitutive expression of thenifA gene. However, this constitutive nifAexpression had no effect on the nucleotide content or on growth of thephbC mutant. It is suggested that PHB is required for maintaining the reducing power of the cell and therefore the bacterial growth. These observations also suggest a new control ofnifA expression to adapt nitrogen fixation to the availability of carbon and reducing equivalents.

scholarly journals Anabaena sp. Strain PCC 7120 Gene devH Is Required for Synthesis of the Heterocyst Glycolipid Layer

2005 ◽  
Vol 187 (7) ◽  
pp. 2326-2331 ◽  
Author(s):  
Martha E. Ramírez ◽  
Pratibha B. Hebbar ◽  
Ruanbao Zhou ◽  
C. Peter Wolk ◽  
Stephanie E. Curtis
Keyword(s):  
Nitrogen Fixation ◽  
Nitrogenase Activity ◽  
Regulatory Protein ◽  
Protein A ◽  
Filamentous Cyanobacterium ◽  
Fixed Nitrogen ◽  
Ultrastructural Studies ◽  
Anabaena Sp ◽  
Pcc 7120 ◽  
Mutant Forms

ABSTRACT In response to deprivation for fixed nitrogen, the filamentous cyanobacterium Anabaena sp. strain PCC 7120 provides a microoxic intracellular environment for nitrogen fixation through the differentiation of semiregularly spaced vegetative cells into specialized cells called heterocysts. The devH gene is induced during heterocyst development and encodes a product with characteristics of a trans-acting regulatory protein. A devH mutant forms morphologically distinguishable heterocysts but is Fox−, incapable of nitrogen fixation in the presence of oxygen. We demonstrate that rearrangements of nitrogen fixation genes take place normally in the devH mutant and that it is Fix+, i.e., has nitrogenase activity under anoxic conditions. The Fox− phenotype was shown by ultrastructural studies to be associated with the absence of the glycolipid layer of the heterocyst envelope. The expression of glycolipid biosynthetic genes in the mutant is greatly reduced, and heterocyst glycolipids are undetectable.

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