The relationship between nitrogen fixation and the production of HD from D2 by cell-free extracts of soya-bean nodule bacteroids.
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ABSTRACT:
SUBMITTER: Turner GL
PROVIDER: S-EPMC1185134 | biostudies-other | 1969 Nov
REPOSITORIES: biostudies-other
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Project description:The apparent Michaelis constants [K'(N(2)) and K'(C(2)H(2))] and the corresponding apparent maximum velocity values (V') for soya-bean bacteroid nitrogenase increased concomitantly in response to increases in nitrogenase Fe-protein concentration and ATP concentration in cell-free assays and in response to O(2) pressure in intact nodules and bacteroid suspensions. K'(C(2)H(2)) in cell-free assays was also affected by pH and by Na(2)S(2)O(4) concentration. Nitrogenase Fe-protein behaved as a catalytic effector reacting at interacting sites on the nitrogenase Fe-Mo-protein. The results indicated that the Fe-Mo-protein probably bears the catalytic sites for N(2) and C(2)H(2) reduction. It is concluded that reduction of N(2) or C(2)H(2) by this nitrogenase involves a reaction mechanism with a sequence of unknown order. The sequence in which substrate, enzyme, effector, ATP and reductant react determines which of the various rate-constants are involved in the apparent Michaelis constant, whose true kinetic meaning was thus unresolved.
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Project description:The nitrogen fixing ability of common bean (Phaseolus vulgaris L.) in association with rhizobia is often characterized as poor compared to other legumes, and nitrogen fertilizers are commonly used in bean production to achieve high yields, which in general inhibits nitrogen fixation. In addition, plants cannot take up all the nitrogen applied to the soil as a fertilizer leading to runoff and groundwater contamination. The overall objective of this work is to reduce use of nitrogen fertilizer in common bean production. This would be a major advance in profitability for the common bean industry in Canada and would significantly improve the ecological footprint of the crop. In the current work, 22 bean genotypes [including recombinant inbred lines (RILs) from the Mist × Sanilac population and a non-nodulating mutant (R99)] were screened for their capacity to fix atmospheric nitrogen under four nitrogen regimes. The genotypes were evaluated in replicated field trials on N-poor soils over three years for the percent nitrogen derived from atmosphere (%Ndfa), yield, and a number of yield-related traits. Bean genotypes differed for all analyzed traits, and the level of nitrogen significantly affected most of the traits, including %Ndfa and yield in all three years. In contrast, application of rhizobia significantly affected only few traits, and the effect was inconsistent among the years. Nitrogen application reduced symbiotic nitrogen fixation (SNF) to various degrees in different bean genotypes. This variation suggests that SNF in common bean can be improved through breeding and selection for the ability of bean genotypes to fix nitrogen in the presence of reduced fertilizer levels. Moreover, genotypes like RIL_38, RIL_119, and RIL_131, being both high yielding and good nitrogen fixers, have potential for simultaneous improvement of both traits. However, breeding advancement might be slow due to an inconsistent correlation between these traits.
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Project description:To understand the role of Medicago truncatula NODULE INCEPTION (MtNIN) in nodule development, we analyzed the transcriptome of roots ectopically overexpressing NIN, nodules of weak mutant allele nin-16, and nodules of dnf1-2 mutant.
2021-11-09 | GSE178119 | GEO
Project description:The mechanism by which nitrate inhibits nitrogen fixation in soybean (Glycine max L.) is not fully understood. Accumulation of ureide in soybean plant tissues may regulate the nitrogen fixation capacity through a feedback pathway. In this study, unilaterally nodulated dual-root soybeans prepared by grafting were grown in sand culture. They were subjected to the removal of the nodulated side roots, and were given either nitrate supply or no supply to the non-nodulated side roots for 3 days (experiment I). Additionally, they received nitrate supply to the non-nodulated side roots for 1-14 days (experiment II). The results showed that nitrate supply increased the levels of asparagine and ureide in soybean shoots (Experiment I). In Experiment II, nodule dry weight, nodule number, nodule nitrogenase activity, and nodule urate oxidase activity decreased significantly after 3, 7, and 14 days of nitrate supply. Ureide content in the shoots and nodules increased after 1, 3, and 7 days of nitrate supply, but decreased after 14 days of nitrate supply. There was a significant positive correlation between urate oxidase activity and nitrogenase activity. Hence, we deduced that nitrate supply increased the asparagine content in soybean shoots, likely inhibiting ureide degradation, which induced the accumulation of ureide in soybean shoots and nodules, and, in turn, feedback inhibited the nodule nitrogen fixation. In addition, urate oxidase activity can be used to assess the nitrogen fixation capacity of nodules.
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Project description:BackgroundBiological nitrogen fixation (BNF) is an important nitrogen source for legume plants, and highly efficient nitrogen fixation requires sufficient phosphorus (P). However, the mechanism of maintaining nitrogen fixation of the legume nodules under low P concentration remains largely unknown.ResultsA nodule-localized SPX protein, GmSPX8, was discovered by transcriptome and functional analysis of its role in N2 fixation was characterized in soybean nodules. GmSPX8 was preferentially expressed in nodules and its expression was gradually increased during nodule development. And also the expression pattern was investigated using reporter gene β-glucuronidase (GUS) driven by the promoter of GmSPX8. GmSPX8 was greatly induced and the GUS activity was increased by 12.2% under P deficiency. Overexpression of GmSPX8 in transgenic plants resulted in increased nodule number, nodule fresh weight and nitrogenase activity by 15.0%, 16.0%, 42.5%, subsequently leading to increased N and P content by 17.0% and 19.0%, while suppression of GmSPX8 showed significantly impaired nodule development and nitrogen fixation efficiency under low P stress. These data indicated that GmSPX8 conferred nodule development and nitrogen fixation under low P condition. By yeast two-hybrid screening, GmPTF1 was identified as a potential interacting protein of GmSPX8, which was further confirmed by BiFC, Y2H and pull down assay. Transcript accumulation of GmPTF1 and its downstream genes such as GmEXLB1 and EXPB2 were increased in GmSPX8 overexpressed transgenic nodules, and in the presence of GmSPX8, the transcriptional activity of GmPTF1 in yeast cells and tobacco leaves was greatly enhanced.ConclusionsIn summary, these findings contribute novel insights towards the role of GmSPX8 in nodule development and nitrogen fixation partly through interacting with GmPTF1 in soybean under low P condition.
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Project description:Small heat shock proteins (sHSPs) are ubiquitous proteins present in all organisms. The sHSPs are not only upregulated under heat shock as well as other stresses but also are expressed in unstressed cells, indicating quite diverse functions of sHSPs. However, there is little known about the role of sHSPs in nodulation and nitrogen fixation in soybean. In this study, we cloned a candidate protein of sHSP, GmHSP17.1, from proteome of nodule and analyzed its function in soybean nodulation. We found that GmHSP17.1 was a cytosolic protein and preferentially expressed during nodule development. An overexpression of GmHSP17.1 in composite transgenic plants showed increases in nodule number, fresh weight, nodule size, area of infection cells, and nitrogenase activity, and subsequently promoted the content of nitrogen and growth of soybean plants. While GmHSP17.1 RNA interference (RNAi) lines showed significantly impaired nodule development and nitrogen fixation efficiency. Through liquid chromatography-tandem mass spectrometry (LC-MS/MS), GmRIP1 was identified as the first potential target of GmHSP17.1, and was shown to be specifically expressed in soybean nodules. The interaction between GmHSP17.1 and GmRIP1 was further confirmed by yeast-two hybrid (Y2H), bimolecular fluorescence complementation (BiFC) in vivo and pull-down assay in vitro. Furthermore, peroxidase activity was markedly increased in GmHSP17.1 overexpressed nodules and decreased in RNAi lines. As a result, the reactive oxygen species (ROS) content greatly decreased in GmHSP17.1 overexpression lines and increased in suppression lines. Taken together, we conclude that GmHSP17.1 plays an important role in soybean nodulation through interacting with GmRIP1. Our results provide foundation for studying the mechanism of nitrogen fixation and for the genetics improvement of legume plants.
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Project description:Isoflavonoids, which include a variety of secondary metabolites, are derived from the phenylpropanoid pathway and are distributed predominantly in leguminous plants. These compounds play a critical role in plant-environment interactions and are beneficial to human health. Isoflavone synthase (IFS) is a key enzyme in isoflavonoid synthesis and shares a common substrate with flavanone-3-hydroxylase (F3H) and flavone synthase II (FNS II). In this study, CRISPR/Cas9-mediated multiplex gene-editing technology was employed to simultaneously target GmF3H1, GmF3H2 and GmFNSII-1 in soya bean hairy roots and plants. Various mutation types and frequencies were observed in hairy roots. Higher mutation efficiencies were found in the T0 transgenic plants, with a triple gene mutation efficiency of 44.44%, and these results of targeted mutagenesis were stably inherited in the progeny. Metabolomic analysis of T0 triple-mutants leaves revealed significant improvement in isoflavone content. Compared with the wild type, the T3 generation homozygous triple mutants had approximately twice the leaf isoflavone content, and the soya bean mosaic virus (SMV) coat protein content was significantly reduced by one-third after infection with strain SC7, suggesting that increased isoflavone content enhanced the leaf resistance to SMV. The isoflavone content in the seeds of T2 triple mutants was also significantly increased. This study provides not only materials for the improvement of soya bean isoflavone content and resistance to SMV but also a simple system to generate multiplex mutations in soya bean, which may be beneficial for further breeding and metabolic engineering.
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Project description:Rhizobia are soil bacteria that can enter into complex symbiotic relationships with legumes, where rhizobia induce the formation of nodules on the plant root. Inside nodules, rhizobia differentiate into nitrogen-fixing bacteroids that reduce atmospheric nitrogen into ammonia, secreting it to the plant host in exchange for carbon. During the transition from free-living bacteria to bacteroids, rhizobial metabolism undergoes major changes. To investigate the metabolism of bacteroids and contrast it with the free-living state, we quantified the proteome of unlabelled bacteroids relative to 15N-labelled free-living rhizobia. The data were used to build a core metabolic model of pea bacteroids for the strain Rhizobium leguminosarum bv. viciae 3841.
2021-09-09 | PXD019467 | Pride
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