Identification of Pseudomonas proteins coordinately induced by acidic amino acids and their amides: a two-dimensional electrophoresis study

Sonawane, Avinash; Klöppner, Ute; Hövel, Sven; Völker, Uwe; Röhm, Klaus‐Heinrich

doi:10.1099/mic.0.26454-0

Cited by 24 publications

(19 citation statements)

References 32 publications

(23 reference statements)

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“…We have shown previously that the aat operon and the glutaminase-encoding ansB gene of P. putida KT2440 are upregulated by the AauRS two-component system (Sonawane et al, 2003b). Here we provide evidence indicating that at least four additional genes are under the control of AauR, i.e.…”

supporting

confidence: 55%

Characterization of a Pseudomonas putida ABC transporter (AatJMQP) required for acidic amino acid uptake: biochemical properties and regulation by the Aau two-component system

Singh

Röhm

2008

Microbiology

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supporting

confidence: 55%

Characterization of a Pseudomonas putida ABC transporter (AatJMQP) required for acidic amino acid uptake: biochemical properties and regulation by the Aau two-component system

Singh

Röhm

2008

Microbiology

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“…An aspartate ammonia-lyase has been described in several other gram-negative organisms including Klebsiella aerogenes (47), Salmonella enterica serovar Typhimurium (27), Pseudomonas species (43), Eikenella corrodens (39), and Haemophilus influenzae (41), and the H. influenzae aspartase was shown to possess the additional function of binding plasminogen (41).…”

mentioning

confidence: 99%

Enzymes Involved in Anaerobic Respiration Appear To Play a Role inActinobacillus pleuropneumoniaeVirulence

et al. 2005

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Actinobacillus pleuropneumoniae, the etiological agent of porcine pleuropneumonia, is able to survive on respiratory epithelia, in tonsils, and in the anaerobic environment of encapsulated sequesters. It was previously demonstrated that a deletion of the anaerobic dimethyl sulfoxide reductase gene (dmsA) results in attenuation in acute disease (N. Baltes, S. Kyaw, I. Hennig-Pauka, and G. F. Gerlach, Infect. Immun. 71:6784-6792, 2003). In the present study, using two-dimensional polyacrylamide gel electrophoresis and quadrupole time-of-flight mass spectrometry, we identified an aspartate ammonia-lyase (AspA) which is upregulated upon induction with bronchoalveolar lavage fluid (BALF). This enzyme is involved in the production of fumarate, an alternative electron acceptor under anaerobic conditions. The coding gene (aspA) was cloned and shown to be present in all A. pleuropneumoniae serotype reference strains. The transcriptional start point was identified downstream of a putative FNR binding motif, and BALF-dependent activation of aspA was confirmed by construction of an isogenic A. pleuropneumoniae mutant carrying a chromosomal aspA::luxAB transcriptional fusion. Two aspA deletion mutants, A. pleuropneumoniae ⌬aspA and A. pleuropneumoniae ⌬aspA⌬dmsA, were constructed, both showing reduced growth under anaerobic conditions in vitro. Pigs challenged with either of the two mutants in an aerosol infection model showed a lower lung lesion score than that of the A. pleuropneumoniae wildtype (wt) controls. Pigs challenged with A. pleuropneumoniae ⌬aspA⌬dmsA had a significantly lower clinical score, and this mutant was rarely reisolated from unaltered lung tissue; in contrast, A. pleuropneumoniae ⌬aspA and the A. pleuropneumoniae wt were consistently reisolated in high numbers. These results suggest that enzymes involved in anaerobic respiration are necessary for the pathogen's ability to persist on respiratory tract epithelium and play an important role in A. pleuropneumoniae pathogenesis.

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“…The proteomics data presented here, together with the results of reporter gene fusions, indicate that aauR acts as a 54 -dependent transcriptional activator of at least two genes or operons, i.e., ansB, encoding PGA, and PP1071 to PP1068, which encodes a glutamate transporter of the ABC type. As shown before, the expression of these two genes was also diminished in an rpoN-deficient mutant of strain KT2440 (17), indicating that the alternate sigma factor 54 is required for their expression, as well. Additional evidence for this assumption is provided by the finding that the ansB (PP2543) promoter contains the Ϫ12/Ϫ24 sequence motif characteristic of 54 -responsive genes (5).…”

Section: Discussionmentioning

confidence: 77%

“…As mentioned above, both proteins are strongly induced by glutamate and repressed by glucose (17). Spot R1 was identified as the protein product of PP1389 (annotated as carboxyphosphonoenolpyruvate phosphonomutase), while spot R2 corresponds to a conserved protein of unknown function.…”

Section: Resultsmentioning

confidence: 99%

“…It metabolizes a wide range of carbon and nitrogen sources, including many amino acids. During growth of P. putida KT2440 on glutamate as a sole source of carbon and nitrogen, a number of proteins are upregulated, including a periplasmic glutaminase/asparaginase (PGA) and two components of an ATP-binding cassette (ABC)-type transporter, i.e., its ATP-binding subunit (encoded by PP1068) and the cognate periplasmic solute-binding protein (encoded by PP1071) (17). A BLAST search in the Comprehensive Microbial Resource database (http://cmr.tigr.org /tigr-scripts/CMR/CmrHomePage.cgi) indicated that the ABC transporter encoded by PP1071 to PP1068 is most probably involved in the uptake of glutamate and/or aspartate.…”

mentioning

confidence: 99%

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The AauR-AauS Two-Component System Regulates Uptake and Metabolism of Acidic Amino Acids in Pseudomonas putida

Sonawane

Singh

Röhm

2006

Appl Environ Microbiol

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Pseudomonas putida KT2440 metabolizes a wide range of carbon and nitrogen sources, including many amino acids. In this study, a 54 -dependent two-component system that controls the uptake and metabolism of acidic amino acids was identified. The system (designated aau, for acidic amino acid utilization) involves a sensor histidine kinase, AauS, encoded by PP1067, and a response regulator, AauR, encoded by PP1066. aauR and aauS deletion mutants were unable to efficiently utilize aspartate (Asp), glutamate (Glu), and glutamine (Gln) as sole sources of carbon and nitrogen. Growth of the mutants was partially restored when the abovementioned amino acids were supplemented with glucose or succinate as an additional carbon source. Uptake of Gln, Asp, and asparagine (Asn) by the aauR mutant was moderately reduced, while Glu uptake was severely impaired. In the absence of glucose, the aauR mutant even secreted Glu into the medium. Furthermore, disruption of aauR affected the activities of several key enzymes of Glu and Asp metabolism, leading to the intracellular accumulation of Glu and greatly reduced survival times under conditions of nitrogen starvation. By a proteomics approach, four major proteins were identified that are downregulated during growth of the aauR mutant on Glu. Two of these were identified as periplasmic glutaminase/asparaginase and the solutebinding protein of a Glu/Asp transporter. Transcriptional analysis of lacZ fusions containing the putative promoter regions of these genes confirmed that their expression is indeed affected by the aau system. Three further periplasmic solute-binding proteins were strongly expressed during growth of the aauR deletion mutant on Glu but downregulated during cultivation on glucose/NH 4 ؉ . These systems may be involved in amino acid efflux.

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Identification of Pseudomonas proteins coordinately induced by acidic amino acids and their amides: a two-dimensional electrophoresis study

Cited by 24 publications

References 32 publications

Characterization of a Pseudomonas putida ABC transporter (AatJMQP) required for acidic amino acid uptake: biochemical properties and regulation by the Aau two-component system

Characterization of a Pseudomonas putida ABC transporter (AatJMQP) required for acidic amino acid uptake: biochemical properties and regulation by the Aau two-component system

Enzymes Involved in Anaerobic Respiration Appear To Play a Role inActinobacillus pleuropneumoniaeVirulence

The AauR-AauS Two-Component System Regulates Uptake and Metabolism of Acidic Amino Acids in Pseudomonas putida

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