Structural homologues PII and Pz of Azospirillum brasilense provide intracellular signalling for selective regulation of various nitrogen‐dependent functions

Zamaroczy, Miklos de

doi:10.1046/j.1365-2958.1998.00938.x

Cited by 77 publications

(107 citation statements)

References 67 publications

(102 reference statements)

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“…Similarly, NtrC-mediated activation of glnK has been described for other bacteria, including E. coli, K. pneumoniae and Azospirillum brasilense (Atkinson & Ninfa, 1998;de Zamaroczy, 1998;He et al, 1998;Jack et al, 1999;van Heeswijk et al, 1996). In addition to NtrC-mediated glnK activation under N-limiting conditions, low but significant glnK expression occurred in an ntrC mutant background in both the presence and absence of ammonium, whereas no b-galactosidase activity was detectable in the strains containing the vector plasmid pML5 carrying the promoterless lacZ gene (data not shown).…”

Section: Resultsmentioning

confidence: 60%

Role of GlnB and GlnK in ammonium control of both nitrogenase systems in the phototrophic bacterium Rhodobacter capsulatus

et al. 2003

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In most bacteria, nitrogen metabolism is tightly regulated and P II proteins play a pivotal role in the regulatory processes. Rhodobacter capsulatus possesses two genes (glnB and glnK ) encoding P II -like proteins. The glnB gene forms part of a glnB-glnA operon and the glnK gene is located immediately upstream of amtB, encoding a (methyl-) ammonium transporter. Expression of glnK is activated by NtrC under nitrogen-limiting conditions. The synthesis and activity of the molybdenum and iron nitrogenases of R. capsulatus are regulated by ammonium on at least three levels, including the transcriptional activation of nifA1, nifA2 and anfA by NtrC, the regulation of NifA and AnfA activity by two different NtrC-independent mechanisms, and the post-translational control of the activity of both nitrogenases by reversible ADP-ribosylation of NifH and AnfH as well as by ADP-ribosylation independent switch-off. Mutational analysis revealed that both P II -like proteins are involved in the ammonium regulation of the two nitrogenase systems. A mutation in glnB results in the constitutive expression of nifA and anfA. In addition, the post-translational ammonium inhibition of NifA activity is completely abolished in a glnB-glnK double mutant. However, AnfA activity was still suppressed by ammonium in the glnB-glnK double mutant. Furthermore, the P II -like proteins are involved in ammonium control of nitrogenase activity via ADP-ribosylation and the switch-off response. Remarkably, in the glnB-glnK double mutant, all three levels of the ammonium regulation of the molybdenum (but not of the alternative) nitrogenase are completely circumvented, resulting in the synthesis of active molybdenum nitrogenase even in the presence of high concentrations of ammonium.

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Section: Resultsmentioning

confidence: 60%

Role of GlnB and GlnK in ammonium control of both nitrogenase systems in the phototrophic bacterium Rhodobacter capsulatus

et al. 2003

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“…In vitro, GlnZ only interacts with DraG and GlnB only interacts with DraT at detectable levels (6,20). Similarly, GlnB, but not GlnZ, activates the nif gene transcriptional regulator NifA (21,26) and GlnB is more efficient than GlnZ in the regulation of the NtrB/NtrC two component system (19,21). The only known common protein target for these paralogues is the ammonium transport protein AmtB (23).…”

Section: Resultsmentioning

confidence: 99%

“…Nitrogenase regulation by the DraT and DraG enzymes has been most fully described in A. brasilense and R. rubrum (21,27). R. rubrum encodes three P II paralogues, namely GlnK, GlnB, and GlnJ (28).…”

Section: Comparison Of Glnz-drag Complex With the Putative Glnj-dragmentioning

confidence: 99%

“…Several lines of evidences showed that both the DraT and DraG activities are regulated by interaction with P II proteins (6,19,20) of which A. brasilense encodes two paralogues, namely GlnB and GlnZ (21). DraT interacts with GlnB while DraG interacts with GlnZ, in vitro these two protein complexes are stabilized by ADP and disrupted in the presence of MgATP and 2-OG (6).…”

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confidence: 99%

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Crystal structure of the GlnZ-DraG complex reveals a different form of P_II-target interaction

Rajendran

Gerhardt

Bjelić

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

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Nitrogen metabolism in bacteria and archaea is regulated by a ubiquitous class of proteins belonging to the P II family. P II proteins act as sensors of cellular nitrogen, carbon, and energy levels, and they control the activities of a wide range of target proteins by protein-protein interaction. The sensing mechanism relies on conformational changes induced by the binding of small molecules to P II and also by P II posttranslational modifications. In the diazotrophic bacterium Azospirillum brasilense, high levels of extracellular ammonium inactivate the nitrogenase regulatory enzyme DraG by relocalizing it from the cytoplasm to the cell membrane. Membrane localization of DraG occurs through the formation of a ternary complex in which the P II protein GlnZ interacts simultaneously with DraG and the ammonia channel AmtB. Here we describe the crystal structure of the GlnZ-DraG complex at 2.1 Å resolution, and confirm the physiological relevance of the structural data by site-directed mutagenesis. In contrast to other known P II complexes, the majority of contacts with the target protein do not involve the T-loop region of P II . Hence this structure identifies a different mode of P II interaction with a target protein and demonstrates the potential for P II proteins to interact simultaneously with two different targets. A structural model of the AmtB-GlnZDraG ternary complex is presented. The results explain how the intracellular levels of ATP, ADP, and 2-oxoglutarate regulate the interaction between these three proteins and how DraG discriminates GlnZ from its close paralogue GlnB.N itrogen is an indispensable element for life. In bacteria and plants, many aspects of the regulation of nitrogen metabolism are controlled by a class of ubiquitous proteins called P II signal transduction proteins (1-4). P II proteins are present in all domains of life and are one of the most widely distributed signal transduction proteins in nature: some prokaryotes encode multiple P II homologues (2, 3). P II proteins act as a central processing unit receiving signals of carbon, energy, and nitrogen levels, integrating and transforming them into different P II conformational states. The multitude of P II conformations control their ability to interact with, and thus regulate the activity of, a variety of target proteins including transporters, enzymes, and transcriptional regulators (4). P II proteins form compact barrel-shaped homotrimeric structures, in which each monomer is 12-14 kDa containing three functionally important loops: the T-, B-, and C-loops (4). The 20-residue long T-loop is structurally very flexible and participates in protein interactions in all the structures of P II -target complexes solved to date. The T-loop also contains the site of posttranslational modification if it is needed. In Proteobacteria, Y51 in the T-loop is subjected to reversible uridylylation in response to the cellular glutamine levels (5). The effector molecules ATP, ADP, and 2-oxoglutarate (2-OG) influence the conformational states of P II pr...

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“…In A. brasilense, P II and P Z proteins are involved differently in nitrogen-dependent regulation of various physiological functions (25). The P II amino acid sequence is about 60% identical to that of P Z and could explain the detection of two hybridization signals in A. brasilense and A. amazonense Southern blots ( Figure 1E, lane 1S and lane 2).…”

Section: Interspecies Conservation Of Nif/gln Genesmentioning

confidence: 95%

Partial characterization of nif genes from the bacterium Azospirillum amazonense

Potrich

Passaglia

Schrank

2001

Braz J Med Biol Res

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Azospirillum amazonense revealed genomic organization patterns of the nitrogen fixation genes similar to those of the distantly related species A. brasilense. Our work suggests that A. brasilense nifHDK, nifENX, fixABC operons and nifA and glnB genes may be structurally homologous to the counterpart genes of A. amazonense. This is the first analysis revealing homology between A. brasilense nif genes and the A. amazonense genome. Sequence analysis of PCR amplification products revealed similarities between the amino acid sequences of the highly conserved nifD and glnB genes of A. amazonense and related genes of A. brasilense and other bacteria. However, the A. amazonense non-coding regions (the upstream activator sequence region and the region between the nifH and nifD genes) differed from related regions of A. brasilense even in nitrogenase structural genes which are highly conserved among diazotrophic bacteria. The feasibility of the 16S ribosomal RNA gene-based PCR system for specific detection of A. amazonense was shown. Our results indicate that the PCR primers for 16S rDNA defined in this article are highly specific to A. amazonense and can distinguish this species from A. brasilense.

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Structural homologues P_II and P_z of Azospirillum brasilense provide intracellular signalling for selective regulation of various nitrogen‐dependent functions

Cited by 77 publications

References 67 publications

Role of GlnB and GlnK in ammonium control of both nitrogenase systems in the phototrophic bacterium Rhodobacter capsulatus

Role of GlnB and GlnK in ammonium control of both nitrogenase systems in the phototrophic bacterium Rhodobacter capsulatus

Crystal structure of the GlnZ-DraG complex reveals a different form of P_II-target interaction

Partial characterization of nif genes from the bacterium Azospirillum amazonense

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Structural homologues PII and Pz of Azospirillum brasilense provide intracellular signalling for selective regulation of various nitrogen‐dependent functions

Cited by 77 publications

References 67 publications

Role of GlnB and GlnK in ammonium control of both nitrogenase systems in the phototrophic bacterium Rhodobacter capsulatus

Role of GlnB and GlnK in ammonium control of both nitrogenase systems in the phototrophic bacterium Rhodobacter capsulatus

Crystal structure of the GlnZ-DraG complex reveals a different form of PII-target interaction

Partial characterization of nif genes from the bacterium Azospirillum amazonense

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Structural homologues P_II and P_z of Azospirillum brasilense provide intracellular signalling for selective regulation of various nitrogen‐dependent functions

Crystal structure of the GlnZ-DraG complex reveals a different form of P_II-target interaction