Effect of ammonia, darkness, and phenazine methosulfate on whole-cell nitrogenase activity and Fe protein modification in Rhodospirillum rubrum

Kanemoto, R H; Ludden, Paul W.

doi:10.1128/jb.158.2.713-720.1984

Cited by 177 publications

(52 citation statements)

References 40 publications

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“…1F [11]. In Rhodospirillum rubrum oxygen leads to a partial modification of the Fe protein, which is not correlated to the strong inhibitory effect on nitrogenase activity [12]. As reported for Azospirillum brasilense and A. lipoferum only extreme oxygen stress produces a covalent modification of Fe protein [13].…”

Section: Introductionmentioning

confidence: 86%

“…Cells pretreated with ammonia or oxygen were quickly passed through a glass microfiber filter, frozen in liquid nitrogen and extracted by boiling in SDS-sample buffer [19]. Aliquots of cell extracts were separated by SDS-PAGE (11% acrylamide, w/w) and electroblotted on a PVDF filter (Immobilon, Millipore) [12,20]. The filter was incubated in the presence of 3% NBCS in PBS.…”

Section: Immunospecific Western Blotmentioning

confidence: 99%

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Regulation of nitrogenase activity inAnabaena variabilisby modification of the Fe protein

Reich

Böger

1989

FEMS Microbiology Letters

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The nitrogenase components of Anabaena variabilis were isolated to prepare a specific antiserum against the cyanobacterial Fe protein. Thereby, the status of the Fe protein subunits could be analyzed after in‐vivo inactivation of nitrogenase. Firstly, nitrogenase activity was switched off by adding ammonia to intact filaments accompanied with the appearance of a slower migrating subunit of the Fe protein as shown by immunospecific Western blots. Inactivation of nitrogenase was correlated with about equal amounts of modified and non‐modified subunits of the Fe protein. Secondly, a rapid inactivation of nitrogenase was achieved by treatment of cells with oxygen, which was followed by a slow appearance of the modified subunit of the Fe protein.

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Section: Introductionmentioning

confidence: 86%

Section: Immunospecific Western Blotmentioning

confidence: 99%

Regulation of nitrogenase activity inAnabaena variabilisby modification of the Fe protein

Reich

Böger

1989

FEMS Microbiology Letters

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“…Samples were taken for acetylene reduction activity assays and demodification reactions. The modification reaction was arrested by SDS sample buffer, and the pattern of modification was analyzed by 10% low-crosslinker SDS-polyacrylamide gel electrophoresis as described [7].…”

Section: Oxidation Of the All-ferrous Dinitrogenase Reductase From Rmentioning

confidence: 99%

NAD‐, NMN‐, and NADP‐dependent modification of dinitrogenase reductases from Rhodospirillum rubrum and Azotobacter vinelandii

et al. 2005

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Nitrogenase activity in the photosynthetic bacterium Rhodospirillum rubrum is reversibly regulated by ADP-ribosylation of a specific arginine residue of dinitrogenase reductase based on the cellular nitrogen or energy status. In this paper, we have investigated the ability of nicotinamide adenine dinucleotide, NAD (the physiological ADP-ribose donor), and its analogs to support covalent modification of dinitrogenase reductase in vitro. R. rubrum dinitrogenase reductase can be modified by DRAT in the presence of 2 mM NAD, but not with 2 mM nicotinamide mononucleotide (NMN) or nicotinamide adenine dinucleotide phosphate (NADP). We also found that the apo-and the all-ferrous forms of R. rubrum dinitrogenase reductase are not substrates for covalent modification. In contrast, Azotobacter vinelandii dinitrogenase reductase can be modified by the dinitrogenase reductase ADP-ribosyl transferase (DRAT) in vitro in the presence of either 2 mM NAD, NMN or NADP as nucleotide donors. We found that: (1) a simple ribose sugar in the modification site of the A. vinelandii dinitrogenase reductase is sufficient to inactivate the enzyme, (2) phospho-ADP-ribose is the modifying unit in the NADP-modified enzyme, and (3) the NMN-modified enzyme carries two ribosephosphate units in one modification site. This is the first report of NADP-or NMN-dependent modification of a target protein by an ADP-ribosyl transferase.

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“…Immunodetection was performed with a primary antibody raised against purified ''universal'' dinitrogenase reductase provided by Paul Ludden (University of Wisconsin, Madison). This antibody was raised in rabbits against a mixture of purified dinitrogenase reductases from A. vinelandii, Rhodospirillum rubrum, Klebsiella pneumoniae, and Clostridium pasteurianum and has been found effective in a wide range of species [31]. The primary antibody was used at a dilution of 1:25 for a 2-h incubation at room temperature.…”

Section: Immunomicroscopymentioning

confidence: 99%

Endophytic nitrogen fixation in dune grasses (Ammophila arenaria and Elymus mollis) from Oregon

Dalton

Kramer

Azios

et al. 2004

FEMS Microbiology Ecology

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Several tropical grasses harbor symbiotic nitrogen-fixing bacteria within their stem and rhizome tissue that may contribute to the nitrogen nutrition of the host plant. We present evidence here that sand dune grasses (Ammophila arenaria and Elymus mollis) from Oregon also contain nitrogen-fixing bacteria. Surface-sterilized stem and rhizome tissue from these species possess acetylene reduction (nitrogen fixation) activity and large populations (10(5) to 10(6) cfu/g fresh weight) of bacteria. These bacteria were cultured on N-free media and identified by sequencing of 16S rRNA genes or by GC-FAME. Random sequencing of numerous colonies from the initial isolation plates of mixed isolates showed that pseudomonads (Stenotrophomonas and Pseudomonas) were by far the most common microorganism. One isolate -Burkholderia sp. strain Aa1 - reduced acetylene in culture with maximum activity at an O(2) concentration of 2% (v/v) in liquid media or 10% on solid media. PCR screening of all the isolates with nifH and nifD primers was positive only for this species. Immunolocalization studies with antibodies to nitrogenase resulted in labeling within plant cell walls of stems and rhizomes. Evidence for a similar nitrogen-fixing association was also detected in Uniola paniculata (sea oats) and Ammophila brevigulata (American beachgrass). We conclude that these grasses, and probably other dune grasses from temperate climates, contain endophytic, diazotrophic bacteria that may contribute to the phenomenal success of these grasses on nutrient-poor sand.

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Effect of ammonia, darkness, and phenazine methosulfate on whole-cell nitrogenase activity and Fe protein modification in Rhodospirillum rubrum

Cited by 177 publications

References 40 publications

Regulation of nitrogenase activity inAnabaena variabilisby modification of the Fe protein

Regulation of nitrogenase activity inAnabaena variabilisby modification of the Fe protein

NAD‐, NMN‐, and NADP‐dependent modification of dinitrogenase reductases from Rhodospirillum rubrum and Azotobacter vinelandii

Endophytic nitrogen fixation in dune grasses (Ammophila arenaria and Elymus mollis) from Oregon

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