Nitrogenase Enzyme Research Articles

ИНДИКАЦИЯ ФРАГМЕНТОВ ГЕНОВ ФЕРМЕНТОВ У БАКТЕРИЙ ВИДА BACILLUS MEGATERIUM

The soil microbial component plays a critical role in maintaining the soil functionality, performing several vital functions such as soil formation, decomposition of dead and decayed organic matter, circulation of macro and micro elements and transformation of toxic chemicals into non-toxic forms. Various Bacillus species, including B. megaterium, are known to fix atmospheric nitrogen. To ensure nitrogen fixation and diversity analysis of diazotrophs, the most widely used gene marker is nifH, which is responsible for synthesis of proteins of the nitrogenase complex. Various soil microorganisms, including B. megaterium, are endowed with the ability to solubilize phosphorus inaccessible form to accessible one and / or mineralize organic phosphorus to accessible phosphorus form. Phosphatases and phytases are two groups of enzymes that catalyze transformation of organic phosphates to inorganic ones. It was found that Bacillus megaterium exhibits acidic and alkaline phosphatase activity and produces phytases. The aim of this work was to determine by RT-PCR method the presence of genes responsible for enzyme synthesis of phytase, nitrogenase, and alkaline phosphatase in strains of previously isolated bacteria of Bacillus megaterium species. To specify the presence of genes encoding synthesis of the desired enzymes in Bacillus megaterium, an in-silico analysis of the annotated genomes of this bacterial species, presented in the NCBI information database, was carried out. Further on, the selection of primers for screening the target parts was made. According to the results of the study, 7 out of 16 isolated B. megaterium strains contained all three required DNA parts responsible for synthesis of phytase, nitrogenase, and alkaline phosphatase enzymes.

Looking for Options to Sustainably Fixate Nitrogen. Are Molecular Metal Oxides Catalysts a Viable Avenue?

Fast and reliable industrial production of ammonia (NH3) is fundamentally sustaining modern society. Since the early 20th Century, NH3 has been synthesized via the Haber–Bosch process, running at conditions of around 350–500°C and 100–200 times atmospheric pressure (15–20 MPa). Industrial ammonia production is currently the most energy-demanding chemical process worldwide and contributes up to 3% to the global carbon dioxide emissions. Therefore, the development of more energy-efficient pathways for ammonia production is an attractive proposition. Over the past 20 years, scientists have imagined the possibility of developing a milder synthesis of ammonia by mimicking the nitrogenase enzyme, which fixes nitrogen from the air at ambient temperatures and pressures to feed leguminous plants. To do this, we propose the use of highly reconfigurable molecular metal oxides or polyoxometalates (POMs). Our proposal is an informed design of the polyoxometalate after exploring the catabolic pathways that cyanobacteria use to fix N2 in nature, which are a different route than the one followed by the Haber–Bosch process. Meanwhile, the industrial process is a “brute force” system towards breaking the triple bond N-N, needing high pressure and high temperature to increase the rate of reaction, nature first links the protons to the N2 to later easier breaking of the triple bond at environmental temperature and pressure. Computational chemistry data on the stability of different polyoxometalates will guide us to decide the best design for a catalyst. Testing different functionalized molecular metal oxides as ammonia catalysts laboratory conditions will allow for a sustainable reactor design of small-scale production.

The Chloroplast of Chlamydomonas reinhardtii as a Testbed for Engineering Nitrogen Fixation into Plants.

Eukaryotic organisms such as plants are unable to utilise nitrogen gas (N2) directly as a source of this essential element and are dependent either on its biological conversion to ammonium by diazotrophic prokaryotes, or its supply as chemically synthesised nitrate fertiliser. The idea of genetically engineering crops with the capacity to fix N2 by introduction of the bacterial nitrogenase enzyme has long been discussed. However, the expression of an active nitrogenase must overcome several major challenges: the coordinated expression of multiple genes to assemble an enzyme complex containing several different metal cluster co-factors; the supply of sufficient ATP and reductant to the enzyme; the enzyme’s sensitivity to oxygen; and the intracellular accumulation of ammonium. The chloroplast of plant cells represents an attractive location for nitrogenase expression, but engineering the organelle’s genome is not yet feasible in most crop species. However, the unicellular green alga Chlamydomonas reinhardtii represents a simple model for photosynthetic eukaryotes with a genetically tractable chloroplast. In this review, we discuss the main advantages, and limitations, of this microalga as a testbed for producing such a complex multi-subunit enzyme. Furthermore, we suggest that a minimal set of six transgenes are necessary for chloroplast-localised synthesis of an ‘Fe-only’ nitrogenase, and from this set we demonstrate the stable expression and accumulation of the homocitrate synthase, NifV, under aerobic conditions. Arguably, further studies in C. reinhardtii aimed at testing expression and function of the full gene set would provide the groundwork for a concerted future effort to create nitrogen-fixing crops.

Examining the Effects of the Nitrogen Environment on Growth and N2-Fixation of Endophytic Herbaspirillum seropedicae in Maize Seedlings by Applying 11C Radiotracing.

Herbaspirillum seropedicae, as an endophyte and prolific root colonizer of numerous cereal crops, occupies an important ecological niche in agriculture because of its ability to promote plant growth and potentially improve crop yield. More importantly, there exists the untapped potential to harness its ability, as a diazotroph, to fix atmospheric N2 as an alternative nitrogen resource to synthetic fertilizers. While mechanisms for plant growth promotion remain controversial, especially in cereal crops, one irrefutable fact is these microorganisms rely heavily on plant-borne carbon as their main energy source in support of their own growth and biological functions. Biological nitrogen fixation (BNF), a microbial function that is reliant on nitrogenase enzyme activity, is extremely sensitive to the localized nitrogen environment of the microorganism. However, whether internal root colonization can serve to shield the microorganisms and de-sensitize nitrogenase activity to changes in the soil nitrogen status remains unanswered. We used RAM10, a GFP-reporting strain of H. seropedicae, and administered radioactive 11CO2 tracer to intact 3-week-old maize leaves and followed 11C-photosynthates to sites within intact roots where actively fluorescing microbial colonies assimilated the tracer. We examined the influence of administering either 1 mM or 10 mM nitrate during plant growth on microbial demands for plant-borne 11C. Nitrogenase activity was also examined under the same growth conditions using the acetylene reduction assay. We found that plant growth under low nitrate resulted in higher nitrogenase activity as well as higher microbial demands for plant-borne carbon than plant growth under high nitrate. However, carbon availability was significantly diminished under low nitrate growth due to reduced host CO2 fixation and reduced allocation of carbon resources to the roots. This response of the host caused significant inhibition of microbial growth. In summary, internal root colonization did little to shield these endophytic microorganisms from the nitrogen environment.

Computational study of ammonia generation by iron(III) and iron(IV) complexes supported by trigonal bipyramidal iron

AbstractHigh‐valent species such as terminal iron nitrides (FeN) are carried out for many organic and inorganic transformations. Simultaneously, they provide significant insight into the reactivity of various metalloenzyme as they involved in the reaction of nitrogenase enzyme. Various biomimetic model complexes were reported to understand nitrogenase enzyme's reactivity. In this framework, Peters et al. have published the facile formation of intermediate species [(TPB)FeIII/IVN]0/1+ from the complex [(TPB)FeN2] (here, TPB = tris(o‐diisopropylphosphinophenyl) borane. However, all species were thoroughly synthesized and characterized. But the mechanism is still elusive in terms of reactivity and the roles of intermediate species. In this work, we have tried to explore the mechanism of ammonia generation from these high‐valent [(TPB)FeIII/IVN]0/1+ species employing the experimental conditions. Our computed results shows a very small energy barrier of 6.7 kJ/mol for the first transition state of protonation by the [(TPB)FeIIIN] species (path1) in the NH bond activation of path1, however comparatively large energy barrier was reported for path2. From this reaction mechanism, it is established that species [(TPB)FeIIIN] is more reactive than [(TPB)FeIVN]+. The reactivity difference between these two species is mainly due to the nature of FeN bond, its basicity and electron delocalization during the NH bond activation. Comprehensive electronic structure investigation of the transition state reveals that the substrate will follow the low energy σ‐type pathway and the electron from the NH bond electron will go in the σz2 orbital. However, the high‐energy π‐type pathway, where the NH bond electron will go in the π*xz orbital. The spectroscopic parameters (Absorption, and Mössbauer) computed for some species, are compared to experimental observation to get belief on the computed data.

Alleviation of Salt Stress in Wheat Seedlings via Multifunctional Bacillus aryabhattai PM34: An In-Vitro Study

Plant growth-promoting rhizobacteria play a substantial role in plant growth and development under biotic and abiotic stress conditions. However, understanding about the functional role of rhizobacterial strains for wheat growth under salt stress remains largely unknown. Here we investigated the antagonistic bacterial strain Bacillus aryabhattai PM34 inhabiting ACC deaminase and exopolysaccharide producing ability to ameliorate salinity stress in wheat seedlings under in vitro conditions. The strain PM34 was isolated from the potato rhizosphere and screened for different PGP traits comprising nitrogen fixation, potassium, zinc solubilization, indole acetic acid, siderophore, and ammonia production, along with various extracellular enzyme activities. The strain PM34 showed significant tolerance towards both abiotic stresses including salt stress (NaCl 2 M), heavy metal (nickel, 100 ppm, and cadmium, 300 ppm), heat stress (60 °C), and biotic stress through mycelial inhibition of Rhizoctonia solani (43%) and Fusarium solani (41%). The PCR detection of ituC, nifH, and acds genes coding for iturin, nitrogenase, and ACC deaminase enzyme indicated the potential of strain PM34 for plant growth promotion and stress tolerance. In the in vitro experiment, NaCl (2 M) decreased the wheat growth while the inoculation of strain PM34 enhanced the germination% (48%), root length (76%), shoot length (75%), fresh biomass (79%), and dry biomass (87%) over to un-inoculated control under 2M NaCl level. The results of experiments depicted the ability of antagonistic bacterial strain Bacillus aryabhattai PM34 to augment salt stress tolerance when inoculated to wheat plants under saline environment.

Metabolomics and Dual RNA-Sequencing on Root Nodules Revealed New Cellular Functions Controlled by Paraburkholderia phymatum NifA.

Paraburkholderia phymatum STM815 is a nitrogen-fixing endosymbiont that nodulate the agriculturally important Phaseolus vulgaris and several other host plants. We previously showed that the nodules induced by a STM815 mutant of the gene encoding the master regulator of nitrogen fixation NifA showed no nitrogenase activity (Fix−) and increased in number compared to P. vulgaris plants infected with the wild-type strain. To further investigate the role of NifA during symbiosis, nodules from P. phymatum wild-type and nifA mutants were collected and analyzed by metabolomics and dual RNA-Sequencing, allowing us to investigate both host and symbiont transcriptome. Using this approach, several metabolites’ changes could be assigned to bacterial or plant responses. While the amount of the C4-dicarboxylic acid succinate and of several amino acids was lower in Fix− nodules, the level of indole-acetamide (IAM) and brassinosteroids increased. Transcriptome analysis identified P. phymatum genes involved in transport of C4-dicarboxylic acids, carbon metabolism, auxin metabolism and stress response to be differentially expressed in absence of NifA. Furthermore, P. vulgaris genes involved in autoregulation of nodulation (AON) are repressed in nodules in absence of NifA potentially explaining the hypernodulation phenotype of the nifA mutant. These results and additional validation experiments suggest that P. phymatum STM815 NifA is not only important to control expression of nitrogenase and related enzymes but is also involved in regulating its own auxin production and stress response. Finally, our data indicate that P. vulgaris does sanction the nifA nodules by depleting the local carbon allocation rather than by mounting a strong systemic immune response to the Fix− rhizobia.

Symbiotic nitrogen fixation in the reproductive structures of a basidiomycete fungus

Nitrogen (N) fixation is a driving force for the formation of symbiotic associations between N2-fixing bacteria and eukaryotes.1 Limited examples of these associations are known in fungi, and none with sexual structures of non-lichenized species.2-6 The basidiomycete Guyanagaster necrorhizus is a sequestrate fungus endemic to the Guiana Shield.7 Like the root rot-causing species in its sister genera Armillaria and Desarmillaria, G.necrorhizus sporocarps fruit from roots of decaying trees (Figures 1A-1C),8 and genome sequencing is consistent with observations that G.necrorhizus is a white-rotting decomposer. This species also represents the first documentation of an arthropod-dispersed sequestrate fungus. Numerous species of distantly related wood-feeding termites, which scavenge for N-rich food, feed on the mature spore-bearing tissue, or gleba, of G.necrorhizus. During feeding, mature spores adhere to termites for subsequent dispersal.9 Using chemical assays, isotope analysis, and high-throughput sequencing, we show that the sporocarps harbor actively N2-fixing Enterobacteriaceae species and that the N content within fungal tissue increases with maturation. Untargeted proteomic profiling suggests that ATP generation in the gleba is accomplished via fermentation. The use of fermentation-an anaerobic process-indicates that the sporocarp environment is anoxic, likely an adaptation to protect the oxygen-sensitive nitrogenase enzyme. Sporocarps also have a thick outer covering, possibly to limit oxygen diffusion. The enriched N content within mature sporocarps may offer a dietary inducement for termites in exchange for spore dispersal. These results show that the flexible metabolic capacity of fungi may facilitate N2-fixing associations, as well as higher-level organismal associations.

Introduction of H2-Uptake Hydrogenase Genes Into Rhizobial Strains Improves Symbiotic Nitrogen Fixation in Vicia sativa and Lotus corniculatus Forage Legumes

Biological nitrogen fixation by the Rhizobium-legume symbiosis allows the conversion of atmospheric nitrogen into ammonia within root nodules mediated by the nitrogenase enzyme. Nitrogenase activity results in the evolution of hydrogen as a result of a side reaction intrinsic to the activity of this enzyme. Some rhizobia, and also other nitrogen fixers, induce a NiFe uptake hydrogenase (Hup) to recycle hydrogen produced by nitrogenase, thus improving the efficiency of the nitrogen fixation process. In this work we report the generation and symbiotic behavior of hydrogenase-positive Rhizobium leguminosarum and Mesorhizobium loti strains effective in vetch (Vicia sativa) and birsfoot trefoil (Lotus corniculatus) forage crops, respectively. The ability of hydrogen recycling was transferred to these strains through the incorporation of hup minitransposon TnHB100, thus leading to full recycling of hydrogen in nodules. Inoculation of Vicia and Lotus plants with these engineered strains led to significant increases in the levels of nitrogen incorporated into the host legumes. The level of improvement of symbiotic performance was dependent on the recipient strain and also on the legume host. These results indicate that hydrogen recycling has the potential to improve symbiotic nitrogen fixation in forage plants.

Nitrogen Fixation and Diazotrophs – A Review

Nitrogen fixation involves formation of ammonium from N2, which needs a high input of energy. Biological nitrogen fixation utilizes the enzyme nitrogenase and ATP to fix nitrogen. Nitrogenase contains a Fe-protein and a Mo-Fe-protein and other metal cofactors. Soil diazotrophs possess the function of fixing atmospheric N2 into biologically available ammonium in ecosystems. In Aechaea, nitrogen fixation has been reported in some methanogens such as Methanobacteriales, Methanococcales, and Methanosarcinales. Community structure and diversity of diazotrophic are correlated with soil pH. All known organisms which involve in nitrogen-fixing which are called diazatrophs are prokaryotes, and both bacterial and archaeal domains are responsible for that. Diazotrophs are categorized into two main groups namely: root-nodule bacteria and plant growth-promoting rhizobacteria. Diazotrophs include free living bacteria, such as Azospirillum, Cupriavidus, and some sulfate reducing bacteria, and symbiotic diazotrophs such Rhizobium and Frankia. Two important parameters which may affect diazotroph communities are temperature and soil moisture in different seasons. To have sustainable agriculture, replacing expensive chemical nitrogen fertilizers with environmentally friendly ways is the most accepted practice.

Positive cooperativity during Azotobacter vinelandii nitrogenase-catalyzed acetylene reduction

The MoFe protein component of the nitrogenase enzyme complex is the substrate reducing site and contains two sets of symmetrically arrayed metallo centers called the P (Fe8S7) and the FeMoco (MoFe7S9-C-homocitrate) centers. The ATP-binding Fe protein is the specific reductant for the MoFe protein. Both symmetrical halves of the MoFe protein are thought to function independently during nitrogenase catalysis. Forming [AlF4]− transition-state complexes between the MoFe protein and the Fe protein of Azotobacter vinelandii ranging from 0 to 2 Fe protein/MoFe protein produced a series of complexes whose specific activity decreases with increase in bound Fe protein/MoFe protein ratio. Reduction of 2H+ to H2 was inhibited in a linear manner with an x-intercept at 2.0 with increasing Fe protein binding, whereas acetylene reduction to ethylene decreased more rapidly with an x-intercept near 1.5. H+ reduction is a distinct process occurring independently at each half of the MoFe protein but acetylene reduction decreases more rapidly than H+ reduction with increasing Fe protein/MoFe protein ratio, suggesting that a response is transmitted between the two αβ halves of the MoFe protein for acetylene reduction as Fe protein is bound. A mechanistic model is derived to investigate this behavior. The model predicts that each site functions independently for 2H+ reduction to H2. For acetylene reduction, the model predicts positive (synchronous) not negative cooperativity arising from acetylene binding to both sites before substrate reduction occurs. When this model is applied to inhibition by Cp2 and modified Av2 protein (L127∆) that form strong, non-dissociable complexes, positive cooperativity is absent and each site acts independently. The results suggest a new paradigm for the catalytic function of the MoFe protein during nitrogenase catalysis.

THE EFFECT OF RHIZOBIA AND NANOPARTICLES ON THE ANATOMICAL GROWTH OF ROOT NODULES OF TWO BEAN PLANT VARIETIES PHASEOLUS VULGARIES L

An experiment was conducted during the spring season of 2018-2019 in the experimental fields of the Faculty of Agriculture / University of Al-Qadisiyah, aiming to study the response of two varieties of the Phaseolus Vulgaris L to the bio-fertilization of RizobiumPhaseolisp with two elements (iron and molybdenum) and their interactions on growth and formation root nodules of plant , as well as some of the anatomical features of theit. The experiment was designed in a Completely Randomized Design (CRD) with three replications in a regulation of three factors (2 × 4 × 2) including bio-fertilizer (inoculated and non-inoculated), nanoscale elements (80 iron, 10 molybdenum, 80 iron + 10 molybdenum). mg. L-1 and (Pole and Bush) varieties. Differences between means were determined by using Duncan’s Multiple Range Test at 0.01 level of confidence. The seeds were treated with bio-fertilizer before planting, the addition of nanoscale elements was carried out after a month of cultivation through soil application method. Measurements were taken at the end of the growing season, that is, at the harvest stage. The result showed that inoculation with rhizobia and nanoparticles had asignificant effect in increasing the number of root nodules and the activity of nitrogenase enzyme, Regarding the anatomical characteristics of the root nodules ,the use of bio-fertilizer and the addition of nano-elements led to asignificant increase in the size of the root nodules (diameter of it),while it negatively affected the thickness of the epidermal and cortical layers.

Transition Metal Chalcogenides as a Versatile and Tunable Platform for Catalytic CO2 and N2 Electroreduction.

Group VI transition metal chalcogenides are the subject of increasing research interest for various electrochemical applications such as low-temperature water electrolysis, batteries, and supercapacitors due to their high activity, chemical stability, and the strong correlation between structure and electrochemical properties. Particularly appealing is their utilization as electrocatalysts for the synthesis of energy vectors and value-added chemicals such as C-based chemicals from the CO2 reduction reaction (CO2R) or ammonia from the nitrogen fixation reaction (NRR). This review discusses the role of structural and electronic properties of transition metal chalcogenides in enhancing selectivity and activity toward these two key reduction reactions. First, we discuss the morphological and electronic structure of these compounds, outlining design strategies to control and fine-tune them. Then, we discuss the role of the active sites and the strategies developed to enhance the activity of transition metal chalcogenide-based catalysts in the framework of CO2R and NRR against the parasitic hydrogen evolution reaction (HER); leveraging on the design rules applied for HER applications, we discuss their future perspective for the applications in CO2R and NRR. For these two reactions, we comprehensively review recent progress in unveiling reaction mechanisms at different sites and the most effective strategies for fabricating catalysts that, by exploiting the structural and electronic peculiarities of transition metal chalcogenides, can outperform many metallic compounds. Transition metal chalcogenides outperform state-of-the-art catalysts for CO2 to CO reduction in ionic liquids due to the favorable CO2 adsorption on the metal edge sites, whereas the basal sites, due to their conformation, represent an appealing design space for reduction of CO2 to complex carbon products. For the NRR instead, the resemblance of transition metal chalcogenides to the active centers of nitrogenase enzymes represents a powerful nature-mimicking approach for the design of catalysts with enhanced performance, although strategies to hinder the HER must be integrated in the catalytic architecture.

Atomically Dispersed Iron Metal Site in a Porphyrin-Based Metal-Organic Framework for Photocatalytic Nitrogen Fixation.

The rational design of photocatalysts for efficient nitrogen (N2) fixation at ambient conditions is important for revolutionizing ammonia production and quite challenging because the great difficulty lies in the adsorption and activation of the inert N2. Inspired by a biological molecule, chlorophyll, featuring a porphyrin structure as the photosensitizer and enzyme nitrogenase featuring an iron (Fe) atom as a favorable binding site for N2via π-backbonding, here we developed a porphyrin-based metal-organic framework (PMOF) with Fe as the active center as an artificial photocatalyst for N2 reduction reaction (NRR) under ambient conditions. The PMOF features aluminum (Al) as metal node imparting high stability and Fe incorporated and atomically dispersed by residing at each porphyrin ring promoting the adsorption and the activation of N2, termed Al-PMOF(Fe). Compared with the pristine Al-PMOF, Al-PMOF(Fe) exhibits a substantial enhancement in NH3 yield (635 μg g-1cat.) and production rate (127 μg h-1 g-1cat.) of 82% and 50%, respectively, on par with the best-performing MOF-based NRR catalysts. Three cycles of photocatalytic NRR experimental results corroborate a stable photocatalytic activity of Al-PMOF(Fe). The combined experimental and theoretical results reveal that the Fe-N site in Al-PMOF(Fe) is the active photocatalytic center that can mitigate the difficulty of the rate-determining step in photocatalytic NRR. The possible reaction pathways of NRR on Al-PMOF(Fe) were established. Our study of porphyrin-based MOF for the photocatalytic NRR will provide insight into the rational design of catalysts for artificial photosynthesis.

Rapid Response of Nitrogen Cycling Gene Transcription to Labile Carbon Amendments in a Soil Microbial Community.

Episodic inputs of labile carbon (C) to soil can rapidly stimulate nitrogen (N) immobilization by soil microorganisms. However, the transcriptional patterns that underlie this process remain unclear. In order to better understand the regulation of N cycling in soil microbial communities, we conducted a 48-h laboratory incubation with agricultural soil where we stimulated the uptake of inorganic N by amending the soil with glucose. We analyzed the metagenome and metatranscriptome of the microbial communities at four time points that corresponded with changes in N availability. The relative abundances of genes remained largely unchanged throughout the incubation. In contrast, glucose addition rapidly increased the transcription of genes encoding ammonium and nitrate transporters, enzymes responsible for N assimilation into biomass, and genes associated with the N regulatory network. This upregulation coincided with an increase in transcripts associated with glucose breakdown and oxoglutarate production, demonstrating a connection between C and N metabolism. When concentrations of ammonium were low, we observed a transient upregulation of genes associated with the nitrogen-fixing enzyme nitrogenase. Transcripts for nitrification and denitrification were downregulated throughout the incubation, suggesting that dissimilatory transformations of N may be suppressed in response to labile C inputs in these soils. These results demonstrate that soil microbial communities can respond rapidly to changes in C availability by drastically altering the transcription of N cycling genes.IMPORTANCE A large portion of activity in soil microbial communities occurs in short time frames in response to an increase in C availability, affecting the biogeochemical cycling of nitrogen. These changes are of particular importance as nitrogen represents both a limiting nutrient for terrestrial plants as well as a potential pollutant. However, we lack a full understanding of the short-term effects of labile carbon inputs on the metabolism of microbes living in soil. Here, we found that soil microbial communities responded to labile carbon addition by rapidly transcribing genes encoding proteins and enzymes responsible for inorganic nitrogen acquisition, including nitrogen fixation. This work demonstrates that soil microbial communities respond within hours to carbon inputs through altered gene expression. These insights are essential for an improved understanding of the microbial processes governing soil organic matter production, decomposition, and nutrient cycling in natural and agricultural ecosystems.

Effect of Potassium and Iron Levels on Growth and Yield of Kharif Rice Bean (Vigna umbellata L.)

Background: Rice bean (Vigna umbellata L.) has recently been notified as a promising pulse crop. It is grown for green manure, green fodder and pulses. Potassium plays a major role in increasing the legume yield and yield components, besides provides tolerance to stress such as high-low temperature and drought. Similarly, iron is important for chlorophyll synthesis and is a key component of the nitrogenase enzyme, which is important for nitrogen fixation.Methods: A field experiment was conducted during kharif season of 2020 at Crop Research Farm, Department of Agronomy, Sam Higginbottom University of Agriculture, Technology and Sciences, Prayagraj (U.P). India. Study the ‘‘Effect of potassium and iron levels on growth and yield of kharif Rice bean (Vigna umbellata L.)”. The present experiment was laid out in randomized block design (RBD).Result: The results revealed that the application of 20 kg K2O/ha + 15 kg Fe/ha recorded maximum plant height (111.17 cm), branches per plant (23.40), nodules per plant at 60 DAS (35.67), plant dry weight (41.33 g/plant), pods per plant (26.20), seeds per pod (7.93), seed yield (1.67 t/ha), stover yield (3.95 t/ha), harvest index (29.69%), net return (Rs 81,155.6/ha) and B: C ratio (2.43). It can be concluded that 20 kg K2O /ha + 15 kg Fe/ha was found more productive as well as economic for Rice bean.

Mechanical coupling in the nitrogenase complex.

The enzyme nitrogenase reduces dinitrogen to ammonia utilizing electrons, protons, and energy obtained from the hydrolysis of ATP. Mo-dependent nitrogenase is a symmetric dimer, with each half comprising an ATP-dependent reductase, termed the Fe Protein, and a catalytic protein, known as the MoFe protein, which hosts the electron transfer P-cluster and the active-site metal cofactor (FeMo-co). A series of synchronized events for the electron transfer have been characterized experimentally, in which electron delivery is coupled to nucleotide hydrolysis and regulated by an intricate allosteric network. We report a graph theory analysis of the mechanical coupling in the nitrogenase complex as a key step to understanding the dynamics of allosteric regulation of nitrogen reduction. This analysis shows that regions near the active sites undergo large-scale, large-amplitude correlated motions that enable communications within each half and between the two halves of the complex. Computational predictions of mechanically regions were validated against an analysis of the solution phase dynamics of the nitrogenase complex via hydrogen-deuterium exchange. These regions include the P-loops and the switch regions in the Fe proteins, the loop containing the residue β-188Ser adjacent to the P-cluster in the MoFe protein, and the residues near the protein-protein interface. In particular, it is found that: (i) within each Fe protein, the switch regions I and II are coupled to the [4Fe-4S] cluster; (ii) within each half of the complex, the switch regions I and II are coupled to the loop containing β-188Ser; (iii) between the two halves of the complex, the regions near the nucleotide binding pockets of the two Fe proteins (in particular the P-loops, located over 130 Å apart) are also mechanically coupled. Notably, we found that residues next to the P-cluster (in particular the loop containing β-188Ser) are important for communication between the two halves.

Trace metal and nitrogen concentrations differentially affect bloom forming cyanobacteria of the genus Dolichospermum

The role of nitrogen and phosphorus in promoting cyanobacterial blooms has been researched for many decades. In contrast, much less is known about the effect of trace metals on regulating cyanobacterial bloom formation. This is particularity relevant for nitrogen-fixing species, as some trace metals act as co-factors in nitrogenase and nitrite reductase enzymes. In the present study, we assessed the response of two nitrogen-fixing species, Dolichospermum lemmermannii and Dolichospermum planctonicum, to copper (Cu), cobalt (Co), molybdenum (Mo), iron (Fe) and nitrogen (N) using a culture-based approach. Response surface modelling showed growth rates in D. planctonicum were positively influenced by increased Co and Fe (p < 0.05), whereas Fe (p < 0.001) and Mo (p < 0.05) increased growth rates in D. lemmermannii. Heterocyst frequency in D. planctonicum was associated with nitrogen (negative response, p < 0.001) and all trace metals tested (positive; p < 0.001 or < 0.05) except Mo. Nitrogen had a strong negative effect (p < 0.001), and Fe a positive (p < 0.01) impact on heterocyst frequency in D. lemmermannii. These results indicate that in addition to regulating nitrogen and phosphorus inputs, management efforts should consider minimising trace metals from entering waterbodies.

Measuring nitrogen fixation by the acetylene reduction assay (ARA): is 3 the magic ratio?

Despite some well-documented drawbacks, the acetylene reduction assay (ARA) remains one of the most widespread methods for measuring biological nitrogen (N2) fixation (BNF) in symbiotic and free-living niches due to its low cost, simplicity, and high throughput potential. Because ARA measures a proxy reaction (the reduction of acetylene to ethylene by the nitrogenase enzyme), a conversion ratio (‘R ratio’) is required to estimate equivalent fixation of N2. Based on the biochemistry of the reactions, the theoretical ratio is usually taken to be 3:1. However, 15N2 calibrations often generate ratios that deviate considerably from this value. We synthesized calibrated R ratios for terrestrial BNF studies, asking whether values converge on the theoretical ratio and vary across N-fixing niches. From 253 mean values (n = 2,072 samples), we find that some niches (legumes, soil, litter) do center on 3:1, while others fall significantly above (wood, lichen) or below (biocrusts). Moss in particular shows a bimodal distribution that may indicate contributions from alternative nitrogenases. However, almost all niches have very wide distributions (up to 2 orders of magnitude); applying ratio values spanning even the 25th -75th percentile cause BNF rates to vary by a factor of 1.5–2.5, and up to > 8. Despite this, only a minority of studies (~ 30% of 345) perform calibrations, and this proportion has not increased over time. We conclude that high variability precludes the use of theoretical values to obtain accurate BNF estimates via ARA, and that historical data should be considered with appropriate caution. Values should be calibrated directly when the goal is to generate accurate rates or cross-condition comparisons.

ОЦЕНКА ЭФФЕКТИВНОСТИ ИСПОЛЬЗОВАНИЯ РИЗОТОРФИНАИ МИНЕРАЛЬНЫХ УДОБРЕНИЙДЛЯ ПОВЫШЕНИЯ УРОЖАЙНОСТИ КОЗЛЯТНИКА ВОСТОЧНОГО

The study of the effect of Rizotorfin inoculant, molyb-denum and mineral fertilizers with nitrogen rates of 30 and 60 kg ha and without nitrogen against the background of phosphorus and potassium P60K60on the development of fodder galegaplants on the first and second growing sea-sons in the steppe zone of the Altai Region showed high responsiveness of fodder galegato improved nutritional conditions. Inoculation and fertilizer application increased the survival and winter hardiness of plants, increased herbage growth, development and yield. Greater survival and winter hardiness were achieved by inoculation with Rizotorfin inoculantand, against its background, mineral fertilizer and molybdenum application. This is determined by the complex effect of bacteria in the inoculant on plants that consists in providing them with a large amount of ni-trogen, growth-promoting hormones and biofungicides. Molybdenum contributes to the activation of the nitrogen-ase enzyme system of bacteria involved in fixing atmos-pheric nitrogen; therefore, its effectiveness increases when used together with Rizotorfin. The herbage of fodder galegaplants fully begins to form only on the second grow-ing season therefore mowing is inappropriate on the year of sowing. The yield of fodder galegaplants may be in-creased by mowing the herbage several times per season. In the Altai Region, two mowings are possible. Inoculation and mineral fertilizers increased thegrass yields on both years of the study. But Rizotorfin inoculantincreased fod-der galegayields to a greater extent. The yield gains through inoculation and mineral fertilizers on the first grow-ing season amounted to 11.1-44.4%, and on the second growingseason -6.8-27.4%. The efficiency of inoculation increased with the combined use of Rizotorfin and mineral fertilizers and ensured a significant gain as compared to the control. The greatest effect was obtained through in-oculation with pure Rizotorfin and combined use of Rizot-orfin with molybdenum and the mineral fertilizer in a dose of N60P60K60.