Experimental evidence for plasmid-bornenor-nirgenes inSinorhizobium melilotiJJ1c10

2004 ◽  
Vol 50 (9) ◽  
pp. 657-667 ◽  
Author(s):  
Yiu-Kwok Chan ◽  
Wayne A McCormick
Keyword(s):  
Nitric Oxide ◽  
Nitrous Oxide ◽  
Sinorhizobium Meliloti ◽  
Nitrite Reduction ◽  
Nucleotide Sequence Analysis ◽  
Gene Products ◽  
Oxide Reduction ◽  
Accessory Gene ◽  
Genes Encoding ◽  
Denitrification Genes

In denitrification, nir and nor genes are respectively required for the sequential dissimilatory reduction of nitrite and nitric oxide to form nitrous oxide. Their location on the pSymA megaplasmid of Sinorhizobium meliloti was confirmed by Southern hybridization of its clones with specific structural gene probes for nirK and norCB. A 20-kb region of pSymA containing the nor-nir genes was delineated by nucleotide sequence analysis. These genes were linked to the nap genes encoding periplasmic proteins involved in nitrate reduction. The nor-nir-nap segment is situated within 30 kb downstream from the nos genes encoding nitrous oxide reduction, with a fix cluster intervening between nir and nos. Most of these predicted nor-nir and accessory gene products are highly homologous with those of related proteobacterial denitrifiers. Functional tests of Tn5 mutants confirmed the requirement of the nirV product and 1 unidentified protein for nitrite reduction as well as the norB-D products and another unidentified protein for nitric oxide reduction. Overall comparative analysis of the derived amino acid sequences of the S. meliloti gene products suggested a close relationship between this symbiotic N2fixer and the free-living non-N2-fixing denitrifier Pseudomonas G-179, despite differences in their genetic organization. This relationship may be due to lateral gene transfer of denitrification genes from a common donor followed by rearrangement and recombination of these genes.Key words: denitrification genes, nitric oxide reductase, nitrite reductase, Rhizobiaceae, Sinorhizobium meliloti.

The complete denitrification pathway of the symbiotic, nitrogen-fixing bacterium Bradyrhizobium japonicum

2005 ◽  
Vol 33 (1) ◽  
pp. 141-144 ◽  
Author(s):  
E.J. Bedmar ◽  
E.F. Robles ◽  
M.J. Delgado
Keyword(s):  
Nitric Oxide ◽  
Nitrous Oxide ◽  
Bradyrhizobium Japonicum ◽  
Mutational Analysis ◽  
Control Level ◽  
Gene Clusters ◽  
Alternative Form ◽  
Nitrate Respiration ◽  
Regulatory Cascade ◽  
Denitrification Genes

Denitrification is an alternative form of respiration in which bacteria sequentially reduce nitrate or nitrite to nitrogen gas by the intermediates nitric oxide and nitrous oxide when oxygen concentrations are limiting. In Bradyrhizobium japonicum, the N2-fixing microsymbiont of soya beans, denitrification depends on the napEDABC, nirK, norCBQD, and nosRZDFYLX gene clusters encoding nitrate-, nitrite-, nitric oxide- and nitrous oxide-reductase respectively. Mutational analysis of the B. japonicum nap genes has demonstrated that the periplasmic nitrate reductase is the only enzyme responsible for nitrate respiration in this bacterium. Regulatory studies using transcriptional lacZ fusions to the nirK, norCBQD and nosRZDFYLX promoter region indicated that microaerobic induction of these promoters is dependent on the fixLJ and fixK2 genes whose products form the FixLJ–FixK2 regulatory cascade. Besides FixK2, another protein, nitrite and nitric oxide respiratory regulator, has been shown to be required for N-oxide regulation of the B. japonicum nirK and norCBQD genes. Thus nitrite and nitric oxide respiratory regulator adds to the FixLJ–FixK2 cascade an additional control level which integrates the N-oxide signal that is critical for maximal induction of the B. japonicum denitrification genes. However, the identity of the signalling molecule and the sensing mechanism remains unknown.

scholarly journals Phylogenomics reveals dynamic evolution of fungal nitric oxide reductases and their relationship to secondary metabolism

2018 ◽  
Author(s):  
Steven A. Higgins ◽  
Christopher W. Schadt ◽  
Patrick B. Matheny ◽  
Frank E. Löffler
Keyword(s):  
Nitric Oxide ◽  
Secondary Metabolism ◽  
Genomic Analysis ◽  
Primary Role ◽  
Ancestral State ◽  
The Novel ◽  
Gene Duplications ◽  
Genes Encoding ◽  
Denitrification Genes ◽  
Genomic Regions

AbstractFungi expressing P450nor, an unconventional nitric oxide (NO) reducing cytochrome P450, are thought to be significant contributors to soil nitrous oxide (N2O) emissions. However, fungal contributions to N2O emissions remain uncertain due to inconsistencies in measurements of N2O formation by fungi. Much of the N2O emitted from antibiotic-amended soil microcosms is attributed to fungal activity, yet fungal isolates examined in pure culture are poor N2O producers. To assist in reconciling these conflicting observations and produce a benchmark genomic analysis of fungal denitrifiers, genes underlying fungal denitrification were examined in >700 fungal genomes. Of 167p450nor–containing genomes identified, 0, 30, and 48 also harbored the denitrification genesnarG,napAornirK, respectively. Compared tonapAandnirK,p450norwas twice as abundant and exhibited two to five-fold more gene duplications, losses, and transfers, indicating a disconnect betweenp450norpresence and denitrification potential. Furthermore, co-occurrence ofp450norwith genes encoding NO-detoxifying flavohemoglobins (Spearman r = 0.87,p= 1.6e−10) confounds hypotheses regarding P450nor’s primary role in NO detoxification. Instead, ancestral state reconstruction united P450nor with actinobacterial cytochrome P450s (CYP105) involved in secondary metabolism (SM) and 19 (11 %)p450nor-containing genomic regions were predicted to be SM clusters. Another 40 (24 %) genomes harbored genes nearbyp450norpredicted to encode hallmark SM functions, providing additional contextual evidence linkingp450norto SM. These findings underscore the potential physiological implications of widespreadp450norgene transfer, support the novel affiliation ofp450norwith fungal SM, and challenge the hypothesis ofp450nor’s primary role in denitrification.

scholarly journals Nitric Oxide Metabolism in Neisseria meningitidis

2002 ◽  
Vol 184 (11) ◽  
pp. 2987-2993 ◽  
Author(s):  
Muna F. Anjum ◽  
Tânia M. Stevanin ◽  
Robert C. Read ◽  
James W. B. Moir
Keyword(s):  
Nitric Oxide ◽  
Nitrous Oxide ◽  
Neisseria Meningitidis ◽  
Gene Product ◽  
Meningococcal Disease ◽  
Nitrite Reductase ◽  
Nitrosative Stress ◽  
Anaerobic Growth ◽  
Gene Products ◽  
Nitric Oxide Metabolism

ABSTRACT Neisseria meningitidis, the causative agent of meningococcal disease in humans, is likely to be exposed to nitrosative stress during natural colonization and disease. The genome of N. meningitidis includes the genes aniA and norB, predicted to encode nitrite reductase and nitric oxide (NO) reductase, respectively. These gene products should allow the bacterium to denitrify nitrite to nitrous oxide. We show that N. meningitidis can support growth microaerobically by the denitrification of nitrite via NO and that norB is required for anaerobic growth with nitrite. NorB and, to a lesser extent, the cycP gene product cytochrome c′ are able to counteract toxicity due to exogenously added NO. Expression of these genes by N. meningitidis during colonization and disease may confer protection against exogenous or endogenous nitrosative stress.

Transcriptional regulation of nitric oxide reduction in Ralstonia eutropha H16

2005 ◽  
Vol 33 (1) ◽  
pp. 193-194 ◽  
Author(s):  
A. Büsch ◽  
K. Strube ◽  
B. Friedrich ◽  
R. Cramm
Keyword(s):  
Nitric Oxide ◽  
Transcriptional Activation ◽  
Ralstonia Eutropha ◽  
Cysteine Residue ◽  
Oxide Reduction ◽  
Signal Molecule ◽  
Terminal Domain ◽  
Genes Encoding ◽  
Nitric Oxide Reduction ◽  
Ralstonia Eutropha H16

Nitric oxide reduction in Ralstonia eutropha H16 is catalysed by the quinol-dependent NO reductase NorB. norB and the adjacent norA form an operon that is controlled by the σ54-dependent transcriptional activator NorR in response to NO. A NorR derivative containing MalE in place of the N-terminal domain binds to a 73 bp region upstream of norA that includes three copies of the putative upstream activator sequence GGT-(N7)-ACC. Mutations altering individual bases of this sequence resulted in an 80–90% decrease in transcriptional activation by wild-type NorR. Similar motifs are present in several proteobacteria upstream of genes encoding proteins of NO metabolism. The N-terminal domain of NorR contains a GAF module and is hypothesized to interact with a signal molecule. A NorR derivative lacking this domain activates the norAB promoter constitutively. Amino acid exchanges within the GAF module identified a cysteine residue that is essential for promoter activation by NorR. Signal sensing by NorR is negatively modulated by the iron-containing protein NorA.

scholarly journals Expression of Nitrite and Nitric Oxide Reductases in Free-Living and Plant-Associated Agrobacterium tumefaciens C58 Cells

2005 ◽  
Vol 71 (8) ◽  
pp. 4427-4436 ◽  
Author(s):  
Seung-Hun Baek ◽  
James P. Shapleigh
Keyword(s):  
Nitric Oxide ◽  
Agrobacterium Tumefaciens ◽  
Nitrogen Atmosphere ◽  
Low Oxygen ◽  
Free Living ◽  
Nitric Oxide Synthase Inhibitor ◽  
Genes Encoding ◽  
Agrobacterium Tumefaciens C58 ◽  
Synthase Inhibitor ◽  
Denitrification Genes

ABSTRACT A number of the bacteria that form associations with plants are denitrifiers. To learn more about how the association with plants affects expression of denitrification genes, the regulation of nitrite and nitric oxide reductases was investigated in Agrobacterium tumefaciens. Analysis of free-living cells revealed that expression of the genes encoding nitrite and nitric oxide reductases, nirK and nor, respectively, requires low-oxygen conditions, nitric oxide, and the transcriptional regulator NnrR. Expression of nor was monitored in plant-associated bacteria using nor-gfp fusion expression. In root association experiments, only a small percentage of the attached cells were fluorescent, even when they were incubated under a nitrogen atmosphere. Inactivation of nirK had no significant effect on the ability of A. tumefaciens to bind to plant roots regardless of the oxygen tension, but it did decrease the occurrence of root-associated fluorescent cells. When wild-type cells containing the gfp fusion were infiltrated into leaves, most cells eventually became fluorescent. The same result was obtained when a nirK mutant was used, suggesting that nitric oxide activated nor expression in the endophytic bacteria. Addition of a nitric oxide synthase inhibitor to block nitric oxide generation by the plant prevented gfp expression in infiltrated nitrite reductase mutants, demonstrating that plant-derived nitric oxide can activate nor expression in infiltrated cells.

Denitrification in Sinorhizobium meliloti

2011 ◽  
Vol 39 (6) ◽  
pp. 1886-1889 ◽  
Author(s):  
María J. Torres ◽  
María I. Rubia ◽  
Eulogio J. Bedmar ◽  
María J. Delgado
Keyword(s):  
Dna Sequences ◽  
Bradyrhizobium Japonicum ◽  
Sinorhizobium Meliloti ◽  
Current Knowledge ◽  
Gene Products ◽  
Free Living ◽  
Symbiotic Plasmid ◽  
Meliloti Strain ◽  
Complete Set ◽  
Denitrification Genes

Denitrification is the complete reduction of nitrate or nitrite to N2, via the intermediates nitric oxide (NO) and nitrous oxide (N2O), and is coupled to energy conservation and growth under O2-limiting conditions. In Bradyrhizobium japonicum, this process occurs through the action of the napEDABC, nirK, norCBQD and nosRZDFYLX gene products. DNA sequences showing homology with nap, nirK, nor and nos genes have been found in the genome of the symbiotic plasmid pSymA of Sinorhizobium meliloti strain 1021. Whole-genome transcriptomic analyses have demonstrated that S. meliloti denitrification genes are induced under micro-oxic conditions. Furthermore, S. meliloti has also been shown to possess denitrifying activities in both free-living and symbiotic forms. Despite possessing and expressing the complete set of denitrification genes, S. meliloti is considered a partial denitrifier since it does not grow under anaerobic conditions with nitrate or nitrite as terminal electron acceptors. In the present paper, we show that, under micro-oxic conditions, S. meliloti is able to grow by using nitrate or nitrite as respiratory substrates, which indicates that, in contrast with anaerobic denitrifiers, O2 is necessary for denitrification by S. meliloti. Current knowledge of the regulation of S. meliloti denitrification genes is also included.

scholarly journals The nitric oxide response in plant-associated endosymbiotic bacteria

2011 ◽  
Vol 39 (6) ◽  
pp. 1880-1885 ◽  
Author(s):  
Juan J. Cabrera ◽  
Cristina Sánchez ◽  
Andrew J. Gates ◽  
Eulogio J. Bedmar ◽  
Socorro Mesa ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Sinorhizobium Meliloti ◽  
Current Knowledge ◽  
Bacterial Cells ◽  
Cellular Targets ◽  
Endosymbiotic Bacteria ◽  
Signalling Molecule ◽  
Genes Encoding ◽  
Rhizobial Species ◽  
Legume Symbiosis

Nitric oxide (NO) is a gaseous signalling molecule which becomes very toxic due to its ability to react with multiple cellular targets in biological systems. Bacterial cells protect against NO through the expression of enzymes that detoxify this molecule by oxidizing it to nitrate or reducing it to nitrous oxide or ammonia. These enzymes are haemoglobins, c-type nitric oxide reductase, flavorubredoxins and the cytochrome c respiratory nitrite reductase. Expression of the genes encoding these enzymes is controlled by NO-sensitive regulatory proteins. The production of NO in rhizobia–legume symbiosis has been demonstrated recently. In functioning nodules, NO acts as a potent inhibitor of nitrogenase enzymes. These observations have led to the question of how rhizobia overcome the toxicity of NO. Several studies on the NO response have been undertaken in two non-dentrifying rhizobial species, Sinorhizobium meliloti and Rhizobium etli, and in a denitrifying species, Bradyrhizobium japonicum. In the present mini-review, current knowledge of the NO response in those legume-associated endosymbiotic bacteria is summarized.

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