Dataset Information

Expression of Pyridoxal 5'-Phosphate-Independent Racemases Can Reduce 2-Aminoacrylate Stress in Salmonella enterica.

ABSTRACT: The RidA protein (PF01042) from Salmonella enterica is a deaminase that quenches 2-aminoacrylate (2AA) and other reactive metabolites. In the absence of RidA, 2AA accumulates, damages cellular enzymes, and compromises the metabolic network. In vitro, RidA homologs from all domains of life deaminate 2AA, and RidA proteins from plants, bacteria, yeast, and humans complement the mutant phenotype of a ridA mutant strain of S. enterica In the present study, a methanogenic archaeon, Methanococcus maripaludis S2, was used to probe alternative mechanisms to restore metabolic balance. M. maripaludis MMP0739, which is annotated as an aspartate/glutamate racemase, complemented a ridA mutant strain and reduced the intracellular 2AA burden. The aspartate/glutamate racemase YgeA from Escherichia coli or S. enterica, when provided in trans, similarly restored wild-type growth to a ridA mutant. These results uncovered a new mechanism to ameliorate metabolic stress, and they suggest that direct quenching by RidA is not the only strategy to quench 2AA.IMPORTANCE 2-Aminoacrylate is an endogenously generated reactive metabolite that can damage cellular enzymes if not directly quenched by the conserved deaminase RidA. This study used an archaeon to identify a RidA-independent mechanism to prevent metabolic stress caused by 2AA. The data suggest that a gene product annotated as an aspartate/glutamate racemase (MMP0739) produces a metabolite that can quench 2AA, expanding our understanding of strategies available to quench reactive metabolites.

SUBMITTER: Hodge-Hanson KM

PROVIDER: S-EPMC5892117 | biostudies-literature | 2018 May

REPOSITORIES: biostudies-literature

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Publications

Expression of Pyridoxal 5'-Phosphate-Independent Racemases Can Reduce 2-Aminoacrylate Stress in Salmonella enterica.

Hodge-Hanson Kelsey M KM Zoino Allison A Downs Diana M DM

Journal of bacteriology 20180409 9

The RidA protein (PF01042) from Salmonella enterica is a deaminase that quenches 2-aminoacrylate (2AA) and other reactive metabolites. In the absence of RidA, 2AA accumulates, damages cellular enzymes, and compromises the metabolic network. In vitro, RidA homologs from all domains of life deaminate 2AA, and RidA proteins from plants, bacteria, yeast, and humans complement the mutant phenotype of a ridA mutant strain of S. enterica In the present study, a methanogenic ...[more]

PMID: 29440254

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Project description:Inorganic orthophosphate (Pi, PO43-) is the preferred phosphorus (P) source of most living cells. In enteric bacterial species such as Salmonella enterica serovar Typhimurium (Salmonella), removal of Pi from the growth medium activates a dedicated two-component signal transduction system which is composed of the membrane-bound sensor kinase PhoR and its cognate transcriptional regulator PhoB. Activated PhoB promotes transcription of genes that are involved in adaptations to low Pi conditions, such as high-affinity Pi transporters and genes required for the assimilation of alternative P sources, allowing bacteria to adapt to low Pi environments. Here, we characterize the PhoB-dependent and independent transcriptional changes elicited by Salmonella in response to P starvation. We identify a set of organic molecules that can serve as the sole P source and pinpoint genes that are involved in their assimilation. Intriguingly, a number of these genes are not under PhoB/PhoR control, indicating that in the absence of environmental Pi, Salmonella may fulfill its biosynthetic demands for P without activating a transcriptional response to Pi starvation. IMPORTANCE Phosphorus (P) is the fifth most abundant element in living cells. This element is acquired mainly as inorganic phosphate (Pi, PO43-). In enteric bacteria, P starvation activates a two-component signal transduction system which is composed of the membrane sensor protein PhoR and its cognate transcription regulator PhoB. PhoB, in turn, promotes the transcription of genes that help maintain Pi homeostasis. Here, we characterize the P starvation response of the bacterium Salmonella enterica. We determine the PhoB-dependent and independent transcriptional changes promoted by P starvation and identify proteins enabling the utilization of a range of organic substrates as sole P sources. We show that transcription and activity of a subset of these proteins are independent of PhoB and Pi availability. These results establish that Salmonella enterica can maintain Pi homeostasis and repress PhoB/PhoR activation even when cells are grown in medium lacking Pi.

Project description:The reactive intermediate deaminase RidA (EC 3.5.99.10) is conserved across all domains of life and deaminates reactive enamine species. When Salmonella enterica ridA mutants are grown in minimal medium, 2-aminoacrylate (2AA) accumulates, damages several pyridoxal 5'-phosphate (PLP)-dependent enzymes, and elicits an observable growth defect. Genetic studies suggested that damage to serine hydroxymethyltransferase (GlyA), and the resultant depletion of 5,10-methelenetetrahydrofolate (5,10-mTHF), was responsible for the observed growth defect. However, the downstream metabolic consequence from GlyA damage by 2AA remains relatively unexplored. This study sought to use untargeted proton nuclear magnetic resonance (1H NMR) metabolomics to determine whether the metabolic state of an S. enterica ridA mutant was accurately reflected by characterizing growth phenotypes. The data supported the conclusion that metabolic changes in a ridA mutant were due to the IlvA-dependent generation of 2AA, and that the majority of these changes were a consequence of damage to GlyA. While many of the metabolic differences for a ridA mutant could be explained, changes in some metabolites were not easily modeled, suggesting that additional levels of metabolic complexity remain to be unraveled.IMPORTANCE The accumulation of the reactive enamine intermediate 2-aminoacrylate (2AA) elicits global metabolic stress in many prokaryotes and eukaryotes by simultaneously damaging multiple pyridoxal 5'-phosphate (PLP)-dependent enzymes. This work employed 1H NMR to expand our understanding of the consequence(s) of 2AA stress on metabolite pools and effectively identify the metabolic changes stemming from one damaged target: GlyA. This study shows that nutrient supplementation during 1H NMR metabolomics experiments can disentangle complex metabolic outcomes stemming from a general metabolic stress. Metabolomics shows great potential to complement classical reductionist approaches to cost-effectively accelerate the rate of progress in expanding our global understanding of metabolic network structure and physiology. To that end, this work demonstrates the utility in implementing nutrient supplementation and genetic perturbation into metabolomics workflows as a means to connect metabolic outputs to physiological phenomena and establish causal relationships.

Dataset Information

Expression of Pyridoxal 5'-Phosphate-Independent Racemases Can Reduce 2-Aminoacrylate Stress in Salmonella enterica.

Publications

Expression of Pyridoxal 5'-Phosphate-Independent Racemases Can Reduce 2-Aminoacrylate Stress in Salmonella enterica.

Similar Datasets

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