Respiratory Protection of Nitrogenase Activity in Azotobacter vinelandii—Roles of the Terminal Oxidases

1997 ◽  
Vol 17 (3) ◽  
pp. 303-317 ◽  
Author(s):  
Robert K. Poole ◽  
Susan Hill
Keyword(s):  
Gene Expression ◽  
Nitrogen Fixation ◽  
Oxygen Reduction ◽  
Crucial Role ◽  
Azotobacter Vinelandii ◽  
Nitrogenase Activity ◽  
Terminal Oxidase ◽  
Respiratory Protection ◽  
Terminal Oxidases ◽  
Cytochrome Bd

Nitrogen fixation by aerobic prokaryotes appears paradoxical: the nitrogen-fixing enzymes—nitrogenases—are notoriously oxygen-labile, yet many bacteria fix nitrogen aerobically. This review summarises the evidence that cytochrome bd, a terminal oxidase unrelated to the mitochondrial and many other bacterial oxidases, plays a crucial role in aerotolerant nitrogen fixation in Azotobacter vinelandii and other bacteria by rapidly consuming oxygen during uncoupled respiration. We review the pertinent properties of this oxidase, particularly its complement of redox centres, the catalytic cycle of oxygen reduction, the affinity of the oxidase for oxygen, and the regulation of cytochrome bd gene expression. The roles of other oxidases and other mechanisms for limiting damage to nitrogenase are assessed.

para-Hydroxybenzoate supported nitrogen fixation in Azotobacter vinelandii strain OP (13705)

1988 ◽  
Vol 34 (11) ◽  
pp. 1271-1275 ◽  
Author(s):  
Jay B. Peterson ◽  
Lynn S. Peterson
Keyword(s):  
Nitrogen Fixation ◽  
Cytoplasmic Membrane ◽  
Azotobacter Vinelandii ◽  
Nitrogenase Activity ◽  
Molecular Nitrogen ◽  
Carbon Sources ◽  
Gel Filtration ◽  
Cell Extracts ◽  
Reduction Assay ◽  
Membrane Bound

Azotobacter vinelandii cells grew with molecular nitrogen and p-hydroxybenzoate as the sole added nitrogen and carbon sources. Nitrogenase activity in p-hydroxybenzoate grown cells was demonstrated with the acetylene reduction assay. Cell extracts contained the enzymes p-hydroxybenzoate hydroxylase (EC 1.14.13.2) and protocatechuate 3,4-dioxygenase (EC 1.13.1.3); oxygenases associated with p-hydroxybenzoate metabolism. These enzymes separated from respiration particles with gel filtration chromatography, indicating that they are soluble and not membrane bound. This evidence indicates that oxygen enters to the inner face of the cytoplasmic membrane during nitrogen fixation.

The Terminal Oxidase in the Marine Bacterium Pseudomonas nautica 617

1997 ◽  
Vol 17 (3) ◽  
pp. 343-346 ◽  
Author(s):  
Helen Simpson ◽  
Michel Denis ◽  
Francesco Malatesta
Keyword(s):  
Hydrogen Peroxide ◽  
Carbon Monoxide ◽  
Oxygen Reduction ◽  
Epr Spectroscopy ◽  
Marine Bacterium ◽  
Large Family ◽  
Terminal Oxidase ◽  
Quinol Oxidase ◽  
Terminal Oxidases ◽  
Production Of Hydrogen

The molecular properties of a novel membrane quinol oxidase from the marine bacterium Pseudomonas nautica 617 are presented. The protein contains 2b hemes/mole which may be distinguished by EPR spectroscopy but not by optical spectroscopy and electrochemistry. Respiration, though being cyanide insensitive, is not inhibited by carbon monoxide and oxygen reduction is carried out only half-way with production of hydrogen peroxide. The terminal oxidase represents, therefore, a unique example in the large family of terminal oxidases known up to date.

scholarly journals mRNA Extraction and Reverse Transcription-PCR Protocol for Detection of nifH Gene Expression by Azotobacter vinelandii in Soil

2003 ◽  
Vol 69 (4) ◽  
pp. 1928-1935 ◽  
Author(s):  
Helmut Bürgmann ◽  
Franco Widmer ◽  
William V. Sigler ◽  
Josef Zeyer
Keyword(s):  
Gene Expression ◽  
Nitrogen Fixation ◽  
Reverse Transcription ◽  
Liquid Culture ◽  
Azotobacter Vinelandii ◽  
Nifh Gene ◽  
Rna Extraction ◽  
Soil Microbial Communities ◽  
Nitrogen Fixing ◽  
Reverse Transcription Pcr

ABSTRACT The study of free-living nitrogen-fixing organisms in bulk soil is hampered by the great diversity of soil microbial communities and the difficulty of relating nitrogen fixation activities to individual members of the diazotroph populations. We developed a molecular method that allows analysis of nifH mRNA expression in soil in parallel with determinations of nitrogen-fixing activity and bacterial growth. In this study, Azotobacter vinelandii growing in sterile soil and liquid culture served as a model system for nifH expression, in which sucrose served as the carbon source and provided nitrogen-limited conditions, while amendments of NH4NO3 were used to suppress nitrogen fixation. Soil RNA extraction was performed with a new optimized direct extraction protocol that yielded nondegraded total RNA. The RNA extracts were of high purity, free of DNA contamination, and allowed highly sensitive and specific detection of nifH mRNA by a reverse transcription-PCR. The level of nifH gene expression was estimated by PCR amplification of reverse-transcribed nifH mRNA fragments with A. vinelandii-specific nifH primers. This new approach revealed that nifH gene expression was positively correlated with bulk nitrogen fixation activity in soil (r 2 = 0.72) and in liquid culture (r 2 = 0.84) and therefore is a powerful tool for studying specific regulation of gene expression directly in the soil environment.

scholarly journals Contrasted Reactivity to Oxygen Tensions inFrankiasp. Strain CcI3 throughout Nitrogen Fixation and Assimilation

2014 ◽  
Vol 2014 ◽  
pp. 1-8 ◽  
Author(s):  
Faten Ghodhbane-Gtari ◽  
Karima Hezbri ◽  
Amir Ktari ◽  
Imed Sbissi ◽  
Nicholas Beauchemin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nitrogen Fixation ◽  
Mrna Level ◽  
Mrna Levels ◽  
Respiratory Protection ◽  
Oxygen Species ◽  
Nitrogen Fixing ◽  
Structural Genes ◽  
Oxygen Tensions ◽  
Strain Cci3

Reconciling the irreconcilable is a primary struggle in aerobic nitrogen-fixing bacteria. Although nitrogenase is oxygen and reactive oxygen species-labile, oxygen tension is required to sustain respiration. In the nitrogen-fixingFrankia, various strategies have been developed through evolution to control the respiration and nitrogen-fixation balance. Here, we assessed the effect of different oxygen tensions onFrankiasp. strain CcI3 growth, vesicle production, and gene expression under different oxygen tensions. Both biomass and vesicle production were correlated with elevated oxygen levels under both nitrogen-replete and nitrogen-deficient conditions. The mRNA levels for the nitrogenase structural genes (nifHDK) were high under hypoxic and hyperoxic conditions compared to oxic conditions. The mRNA level for the hopanoid biosynthesis genes (sqhC andhpnC) was also elevated under hyperoxic conditions suggesting an increase in the vesicle envelope. Under nitrogen-deficient conditions, thehup2 mRNA levels increased with hyperoxic environment, whilehup1 mRNA levels remained relatively constant. Taken together, these results indicate thatFrankiaprotects nitrogenase by the use of multiple mechanisms including the vesicle-hopanoid barrier and increased respiratory protection.

scholarly journals Terminal Oxidases of Bacillus subtilisStrain 168: One Quinol Oxidase, Cytochromeaa3 or Cytochrome bd, Is Required for Aerobic Growth

2000 ◽  
Vol 182 (23) ◽  
pp. 6557-6564 ◽  
Author(s):  
Lena Winstedt ◽  
Claes von Wachenfeldt
Keyword(s):  
Bacillus Subtilis ◽  
Sequence Analysis ◽  
Terminal Oxidase ◽  
Rich Medium ◽  
Aerobic Growth ◽  
Quinol Oxidase ◽  
Terminal Oxidases ◽  
Cytochrome Bd ◽  
Heme Copper Oxidases ◽  
Quinol Oxidases

ABSTRACT The gram-positive endospore-forming bacterium Bacillus subtilis has, under aerobic conditions, a branched respiratory system comprising one quinol oxidase branch and one cytochrome oxidase branch. The system terminates in one of four alternative terminal oxidases. Cytochrome caa 3 is a cytochromec oxidase, whereas cytochrome bd and cytochromeaa 3 are quinol oxidases. A fourth terminal oxidase, YthAB, is a putative quinol oxidase predicted from DNA sequence analysis. None of the terminal oxidases are, by themselves, essential for growth. However, one quinol oxidase (cytochromeaa 3 or cytochrome bd) is required for aerobic growth of B. subtilis strain 168. Data indicating that cytochrome aa 3 is the major oxidase used by exponentially growing cells in minimal and rich medium are presented. We show that one of the two heme-copper oxidases, cytochrome caa 3 or cytochromeaa 3, is required for efficient sporulation ofB. subtilis strain 168 and that deletion of YthAB in a strain lacking cytochrome aa 3 makes the strain sporulation deficient.

scholarly journals Metabolic model of nitrogen-fixing obligate aerobe Azotobacter vinelandii demonstrates adaptation to oxygen concentration and metal availability

2021 ◽  
Author(s):  
Alexander B Alleman ◽  
Florence Mus ◽  
John W Peters
Keyword(s):  
Nitrogen Fixation ◽  
Biological Nitrogen Fixation ◽  
Azotobacter Vinelandii ◽  
Metabolic Model ◽  
Metal Availability ◽  
Respiratory Protection ◽  
Nitrogen Fixing ◽  
A Genome ◽  
Biological Nitrogen ◽  
Genome Scale

There is considerable interest in promoting biological nitrogen fixation as a mechanism to reduce the inputs of nitrogenous fertilizers in agriculture, a problem of agronomic, economic, and environmental importance. For the potential impact of biological nitrogen fixation in agriculture to be realized, there are considerable fundamental knowledge gaps that need to be addressed. Biological nitrogen fixation or the reduction of N2 to NH3 is catalyzed by nitrogenase which requires a large amount of energy in the form of ATP and low potential electrons. Nitrogen-fixing organisms that respire aerobically have an advantage in meeting the energy demands of biological nitrogen fixation but face challenges of protecting nitrogenase from inactivation in the presence of oxygen. Here, we have constructed a genome-scale metabolic model of the aerobic metabolism of nitrogen-fixing bacteria Azotobacter vinelandii, which uses a complex electron transport system, termed respiratory protection, to consume oxygen at a high rate keeping intracellular conditions microaerobic. Our model accurately determines growth rate under high oxygen and high substrate concentration conditions, demonstrating the large flux of energy directed to respiratory protection. While respiratory protection mechanisms compensate the energy balance in high oxygen conditions, it does not account for all substrate intake, leading to increased maintenance rates. We have also shown how A. vinelandii can adapt under different oxygen concentrations and metal availability by rearranging flux through the electron transport system. Accurately determining the energy balance in a genome-scale metabolic model is required for future engineering approaches.

scholarly journals The Klebsiella pneumoniae cytochrome bd' terminal oxidase complex and its role in microaerobic nitrogen fixation

Microbiology ◽  
1997 ◽  
Vol 143 (8) ◽  
pp. 2673-2683 ◽  
Author(s):  
N. S. Juty ◽  
F. Moshiri ◽  
M. Merrick ◽  
C. Anthony ◽  
S. Hill
Keyword(s):  
Nitrogen Fixation ◽  
Klebsiella Pneumoniae ◽  
Terminal Oxidase ◽  
Cytochrome Bd

scholarly journals cDNA Macroarray Analysis of Gene Expression in Ineffective Nodules Induced on the Lotus japonicus sen1 Mutant

2004 ◽  
Vol 17 (11) ◽  
pp. 1223-1233 ◽  
Author(s):  
Norio Suganuma ◽  
Atsuko Yamamoto ◽  
Ai Itou ◽  
Tsuneo Hakoyama ◽  
Mari Banba ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nitrogen Fixation ◽  
Nitrogenase Activity ◽  
Cysteine Proteinase ◽  
Lotus Japonicus ◽  
Vascular Bundles ◽  
Cdna Macroarray ◽  
Plant Genes ◽  
Macroarray Analysis ◽  
Mutant Forms

The Lotus japonicus sen1 mutant forms ineffective nodules in which development is arrested at the stage of bacterial differentiation into nitrogen-fixing bacteroids. Here, we used cDNA macroarray systems to compare gene expression in ineffective nodules induced on the sen1 mutant with gene expression in wild-type nodules, in order to identify the host plant genes that are involved in nitrogen fixation. Macroarray analysis coupled with Northern blot analysis revealed that the expression of 18 genes was significantly enhanced in ineffective sen1 nodules, whereas the expression of 30 genes was repressed. Many of the enhanced genes encoded hydrolase enzymes, such as cysteine proteinase and asparaginase, that might function in the early senescence of sen1 nodules. By contrast, the repressed genes encoded nodulins, enzymes that are involved in carbon and nitrogen metabolism, membrane transporters, enzymes involved in phytohormone metabolism and secondary metabolism, and regulatory proteins. These proteins might have a role in the establishment of nitrogen fixation. In addition, we discovered two novel genes that encoded glutamate-rich proteins and were localized in the vascular bundles of the nodules. The expression of these genes was repressed in the ineffective nodules, which had lower levels of nitrogenase activity.

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