Physiology and Metabolism
Research Article
2 April 2024

Tetrahydrofolate levels influence 2-aminoacrylate stress in Salmonella enterica

ABSTRACT

In Salmonella enterica, the absence of the RidA deaminase results in the accumulation of the reactive enamine 2-aminoacrylate (2AA). The resulting 2AA stress impacts metabolism and prevents growth in some conditions by inactivating a specific target pyridoxal 5′-phosphate (PLP)-dependent enzyme(s). The detrimental effects of 2AA stress can be overcome by changing the sensitivity of a critical target enzyme or modifying flux in one or more nodes in the metabolic network. The catabolic L-alanine racemase DadX is a target of 2AA, which explains the inability of an alr ridA strain to use L-alanine as the sole nitrogen source. Spontaneous mutations that suppressed the growth defect of the alr ridA strain were identified as lesions in folE, which encodes GTP cyclohydrolase and catalyzes the first step of tetrahydrofolate (THF) synthesis. The data here show that THF limitation resulting from a folE lesion, or inhibition of dihydrofolate reductase (FolA) by trimethoprim, decreases the 2AA generated from endogenous serine. The data are consistent with an increased level of threonine, resulting from low folate levels, decreasing 2AA stress.

IMPORTANCE

RidA is an enamine deaminase that has been characterized as preventing the 2-aminoacrylate (2AA) stress. In the absence of RidA, 2AA accumulates and damages various cellular enzymes. Much of the work describing the 2AA stress system has depended on the exogenous addition of serine to increase the production of the enamine stressor. The work herein focuses on understanding the effect of 2AA stress generated from endogenous serine pools. As such, this work describes the consequences of a subtle level of stress that nonetheless compromises growth in at least two conditions. Describing mechanisms that alter the physiological consequences of 2AA stress increases our understanding of endogenous metabolic stress and how the robustness of the metabolic network allows perturbations to be modulated.

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REFERENCES

1.
Jeong H, Tombor B, Albert R, Oltvai ZN, Barabási AL. 2000. The large-scale organization of metabolic networks. Nature 407:651–654.
2.
Albert R, Jeong H, Barabasi AL. 2000. Error and attack tolerance of complex networks. Nature 406:378–382.
3.
Keller MA, Piedrafita G, Ralser M. 2015. The widespread role of non-enzymatic reactions in cellular metabolism. Curr Opin Biotechnol 34:153–161.
4.
de Lorenzo V, Sekowska A, Danchin A. 2015. Chemical reactivity drives spatiotemporal organisation of bacterial metabolism. FEMS Microbiol Rev 39:96–119.
5.
de Crécy-Lagard V, Haas D, Hanson AD. 2018. Newly-discovered enzymes that function in metabolite damage-control. Curr Opin Chem Biol 47:101–108.
6.
Imlay JA. 2013. The molecular mechanisms and physiological consequences of oxidative stress: lessons from a model bacterium. Nat Rev Microbiol 11:443–454.
7.
Semchyshyn HM. 2014. Reactive carbonyl species in vivo: generation and dual biological effects. Sci World J 2014:417842.
8.
Borchert AJ, Ernst DC, Downs DM. 2019. Reactive enamines and imines in vivo: lessons from the RidA paradigm. Trends Biochem Sci 44:849–860.
9.
Brinsmade SR, Paldon T, Escalante-Semerena JC. 2005. Minimal functions and physiological conditions required for growth of Salmonella enterica on ethanolamine in the absence of the metabolosome. J Bacteriol 187:8039–8046.
10.
Schmitz G, Downs DM. 2004. Reduced transaminase B (IlvE) activity caused by the lack of yjgF is dependent on the status of threonine deaminase (IlvA) in Salmonella enterica serovar typhimurium. J Bacteriol 186:803–810.
11.
Ernst DC, Lambrecht JA, Schomer RA, Downs DM. 2014. Endogenous synthesis of 2-aminoacrylate contributes to cysteine sensitivity in Salmonella enterica. J Bacteriol 196:3335–3342.
12.
Ernst DC, Anderson ME, Downs DM. 2016. L-2,3-diaminopropionate generates diverse metabolic stresses in Salmonella enterica. Mol Microbiol 101:210–223.
13.
Lambrecht JA, Schmitz GE, Downs DM. 2013. RidA proteins prevent metabolic damage inflicted by PLP-dependent dehydratases in all domains of life. mBio 4:e00033-13.
14.
Irons JL, Hodge-Hanson K, Downs DM. 2020. Rida proteins protect against metabolic damage by reactive intermediates. Microbiol Mol Biol Rev 84:e00024-20.
15.
Shen W, Borchert AJ, Downs DM. 2022. 2-aminoacrylate stress damages diverse PLP-dependent enzymes in vivo. J Biol Chem 298:101970.
16.
Niehaus TD, Nguyen TND, Gidda SK, ElBadawi-Sidhu M, Lambrecht JA, McCarty DR, Downs DM, Cooper AJL, Fiehn O, Mullen RT, Hanson AD. 2014. Arabidopsis and maize RidA proteins preempt reactive enamine/imine damage to branched-chain amino acid biosynthesis in plastids. Plant Cell 26:3010–3022.
17.
Degani G, Barbiroli A, Regazzoni L, Popolo L, Vanoni MA. 2018. Imine deaminase activity and conformational stability of UK114, the mammalian member of the rid protein family active in amino acid metabolism. Int J Mol Sci 19:945.
18.
Ernst DC, Downs DM. 2018. Mmf1p couples amino acid metabolism to mitochondrial DNA maintenance in Saccharomyces cerevisiae. mBio 9:e00084-18.
19.
Digiovanni S, Visentin C, Degani G, Barbiroli A, Chiara M, Regazzoni L, Di Pisa F, Borchert AJ, Downs DM, Ricagno S, Vanoni MA, Popolo L. 2020. Two novel fish paralogs provide insights into the Rid family of imine deaminases active in pre-empting enamine/imine metabolic damage. Sci Rep 10:10135.
20.
Martínez-Chavarría LC, Sagawa J, Irons J, Hinz AK, Lemon A, Graça T, Downs DM, Vadyvaloo V. 2020. Putative horizontally acquired genes, highly transcribed during Yersinia pestis flea infection, are induced by hyperosmotic stress and function in aromatic amino acid metabolism. J Bacteriol 202:e00733-19.
21.
Irons J, Sacher JC, Szymanski CM, Downs DM. 2019. Cj1388 is a RidA homolog and is required for flagella biosynthesis and/or function in Campylobacter jejuni. Front Microbiol 10:2058.
22.
Irons J, Hodge-Hanson KM, Downs DM. 2018. PA5339, a RidA homolog, is required for full growth in Pseudomonas aeruginosa. J Bacteriol 200:200.
23.
Lambrecht JA, Flynn JM, Downs DM. 2012. Conserved YjgF protein family deaminates reactive enamine/imine intermediates of pyridoxal 5'-phosphate (PLP)-dependent enzyme reactions. J Biol Chem 287:3454–3461.
24.
Flynn JM, Downs DM. 2013. In the absence of RidA, endogenous 2-aminoacrylate inactivates alanine racemases by modifying the pyridoxal 5'-phosphate cofactor. J Bacteriol 195:3603–3609.
25.
Flynn JM, Christopherson MR, Downs DM. 2013. Decreased coenzyme A levels in ridA mutant strains of Salmonella enterica result from inactivated serine hydroxymethyltransferase. Mol Microbiol 89:751–759.
26.
Walsh C. 1982. Suicide substrates: mechanism-based enzyme inactivators. Tetrahedron 38:871–909.
27.
Likos JJ, Ueno H, Feldhaus RW, Metzler DE. 1982. A novel reaction of the coenzyme of glutamate decarboxylase with L-serine O-sulfate. Biochemistry 21:4377–4386.
28.
Ueno H, Likos JJ, Metzler DE. 1982. Chemistry of the inactivation of cytosolic aspartate aminotransferase by serine O-sulfate. Biochemistry 21:4387–4393.
29.
Christopherson MR, Schmitz GE, Downs DM. 2008. YjgF is required for isoleucine biosynthesis when Salmonella enterica is grown on pyruvate medium. J Bacteriol 190:3057–3062.
30.
Ernst DC, Downs DM. 2016. 2-aminoacrylate stress induces a context-dependent glycine requirement in ridA strains of Salmonella enterica. J Bacteriol 198:536–543.
31.
Borchert AJ, Downs DM. 2018. Analyses of variants of the Ser/Thr dehydratase IlvA provide insight into 2-aminoacrylate metabolism in Salmonella enterica. J Biol Chem 293:19240–19249.
32.
Whitaker GH, Ernst DC, Downs DM. 2021. Absence of MMF1 disrupts heme biosynthesis by targeting Hem1pin Saccharomyces cerevisiae. Yeast 38:615–624.
33.
Fulton RL, Downs DM. 2023. Modulators of a robust and efficient metabolism: perspective and insights from the Rid superfamily of proteins. In Poole RK, Kelly DJ (ed), Advanced in microbial physiology. Vol. 83. Elsevier.
34.
Fulton RL, Irons J, Downs DM. 2022. The cysteine desulfurase IscS is a significant target of 2-aminoacrylate damage in Pseudomonas aeruginosa. mBio 13:e0107122.
35.
Ernst DC, Christopherson MR, Downs DM. 2018. Increased activity of cystathionine beta-lyase suppresses 2-aminoacrylate stress in Salmonella enterica. J Bacteriol 200:e00040-18.
36.
Hodge-Hanson KM, Zoino A, Downs DM. 2018. Expression of PLP-independent racemases can reduce 2-aminoacrylate stress in Salmonella enterica. J Bacteriol 200:e00751-17.
37.
Wasserman SA, Walsh CT, Botstein D. 1983. Two alanine racemase genes in Salmonella typhimurium that differ in structure and function. J Bacteriol 153:1439–1450.
38.
Lambert MP, Neuhaus FC. 1972. Factors affecting the level of alanine racemase in Escherichia coli. J Bacteriol 109:1156–1161.
39.
Nar H, Huber R, Auerbach G, Fischer M, Hösl C, Ritz H, Bracher A, Meining W, Eberhardt S, Bacher A. 1995. Active site topology and reaction mechanism of GTP cyclohydrolase I. Proc Natl Acad Sci U S A 92:12120–12125.
40.
Allen S, Zilles JL, Downs DM. 2002. Metabolic flux in both the purine mononucleotide and histidine biosynthetic pathways can influence synthesis of the hydroxymethyl pyrimidine moiety of thiamine in Salmonella enterica. J Bacteriol 184:6130–6137.
41.
Davis RW, Botstein D, Roth JR. 1980. Advanced bacterial genetics. Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y.
42.
Li K, Li T, Yang SS, Wang XD, Gao LX, Wang RQ, Gu J, Zhang XE, Deng JY. 2017. Deletion of nudB causes increased susceptibility to antifolates in Escherichia coli and Salmonella enterica. Antimicrob Agents Chemother 61:e02378-16.
43.
Quinlivan EP, McPartlin J, Weir DG, Scott J. 2000. Mechanism of the antimicrobial drug trimethoprim revisited. FASEB J 14:2519–2524.
44.
Kang L, Shaw AC, Xu D, Xia W, Zhang J, Deng J, Wöldike HF, Liu Y, Su J. 2011. Upregulation of MetC is essential for d-alanine-independent growth of an alr/dadX-deficient Escherichia coli strain. J Bacteriol 193:1098–1106.
45.
Flavin M, Slaughter C. 1964. An intermediate trapped by maleimides in a pyridoxal-phosphate potentiated enzymatic elimination reaction. Biochemistry 3:885–893.
46.
Hillebrand GG, Dye JL, Suelter CH. 1979. Formation of an intermediate and its rate of conversion to pyruvate during the tryptophanase-catalyzed degradation of S-o-nitrophenyl-L-cysteine. Biochemistry 18:1751–1755.
47.
Palmer LD, Dougherty MJ, Downs DM. 2012. Analysis of ThiC variants in the context of the metabolic network of Salmonella enterica. J Bacteriol 194:6088–6095.
48.
Rébora K, Laloo B, Daignan-Fornier B. 2005. Revisiting purine-histidine cross-pathway regulation in Saccharomyces cerevisiae: a central role for a small molecule. Genetics 170:61–70.
49.
Magasanik B, Karibian D. 1960. Purine nucleotide cycles and their metabolic role. J Biol Chem 235:2672–2681.
50.
Yura T. 1956. Evidence of nonidentical alleles in purine requiring mutants of Salmonella typhimurium, Vol. 612, p 63–75. Publications of the Carnegie Institution.
51.
Newell PC, Tucker RG. 1968. Biosynthesis of the pyrimidine moiety of thiamine. A new route of pyrimidine biosynthesis involving purine intermediates. Biochem J 106:279–287.
52.
Bazurto JV, Downs DM. 2014. Amino-4-imidazolecarboxamide ribotide directly inhibits coenzyme A biosynthesis in Salmonella enterica. J Bacteriol 196:772–779.
53.
Pimkin M, Markham GD. 2008. The CBS subdomain of inosine 5'-monophosphate dehydrogenase regulates purine nucleotide turnover. Mol Microbiol 68:342–359.
54.
Rohlman CE, Matthews RG. 1990. Role of purine biosynthetic intermediates in response to folate stress in Escherichia coli. J Bacteriol 172:7200–7210.
55.
Nierlich DP, Magasanik B. 1965. Regulation of purine ribonucleotide synthesis by end product inhibition. The effect of adenine and guanine ribonucleotides on the 5'- phosphoribosyl-pyrophosphate amidotransferase of Aerobacter aeroge. J Biol Chem 240:358–365.
56.
Martin RG. 1963. The first enzyme in histidine biosynthesis: the nature of feedback inhibition. J Biol Chem 238:257–268.
57.
Shedlovsky AE, Magasanik B. 1962. A defect in histidine biosynthesis causing an adenine deficiency. J Biol Chem 237:3725–3730.
58.
Christopherson MR, Lambrecht JA, Downs D, Downs DM. 2012. Suppressor analyses identify threonine as a modulator of ridA mutant phenotypes in Salmonella enterica. PLoS One 7:e43082.
59.
Kwon YK, Higgins MB, Rabinowitz JD. 2010. Antifolate-induced depletion of intracellular glycine and purines inhibits thymineless death in E. coli. ACS Chem Biol 5:787–795.
60.
Borchert AJ, Walejko JM, Guennec AL, Ernst DC, Edison AS, Downs DM. 2019. Integrated metabolomics and transcriptomics suggest the global metabolic response to 2-aminoacrylate stress in Salmonella enterica. Metabolites 10:12.
61.
Downs DM, Bazurto JV, Gupta A, Fonseca LL, Voit EO. 2018. The three-legged stool of understanding metabolism: integrating metabolomics with biochemical genetics and computational modeling. AIMS Microbiol 4:289–303.
62.
Koenigsknecht MJ, Downs DM. 2010. Thiamine biosynthesis can be used to dissect metabolic integration. Trends Microbiol 18:240–247.
63.
Bazurto JV, Heitman NJ, Downs DM. 2015. Aminoimidazole carboxamide ribotide exerts opposing effects on thiamine synthesis in Salmonella enterica. J Bacteriol 197:2821–2830.
64.
Vogel HJ, Bonner DM. 1956. Acetylornithinase of Escherichia coli: partial purification and some properties. J Biol Chem 218:97–106.
65.
Balch WE, Wolfe RS. 1976. New approach to the cultivation of methanogenic bacteria: 2-mercaptoethanesulfonic acid (HS-CoM)-dependent growth of Methanobacterium ruminantium in a pressurized atmosphere. Appl Environ Microbiol 32:781–791.
66.
Schmieger H. 1972. Phage P22-mutants with increased or decreased transduction abilities. Mol Gen Genet 119:75–88.
67.
Chan RK, Botstein D, Watanabe T, Ogata Y. 1972. Specialized transduction of tetracycline resistance by phage P22 in Salmonella typhimurium: II. Properties of a high-frequency-transducing lysate. Virology 50:883–898.
68.
Downs DM, Petersen L. 1994. apbA, a new genetic locus involved in thiamine biosynthesis in Salmonella typhimurium. J Bacteriol 176:4858–4864.
69.
Datsenko KA, Wanner BL. 2000. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A 97:6640–6645.
70.
Way JC, Davis MA, Morisato D, Roberts DE, Kleckner N. 1984. New Tn10 derivatives for transposon mutagenesis and for construction of lacZ operon fusions by transposition. Gene 32:369–379.
71.
Guzman LM, Belin D, Carson MJ, Beckwith J. 1995. Tight regulation, modulation, and high-level expression by vectors containing the arabinose PBAD promoter. J Bacteriol 177:4121–4130.
72.
Olson B, Markwell J. 2007. Assays for determination of protein concentration. Curr Protoc Protein Sci Chapter 3:Unit.
73.
Compton SJ, Jones CG. 1985. Mechanism of dye response and interference in the Bradford protein assay. Anal Biochem 151:369–374.

Information & Contributors

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Published In

cover image Journal of Bacteriology
Journal of Bacteriology
Volume 206Number 418 April 2024
eLocator: e00042-24
Editor: George O'Toole, Geisel School of Medicine at Dartmouth, Hanover, New Hampshire, USA
PubMed: 38563759

History

Received: 6 February 2024
Accepted: 12 March 2024
Published online: 2 April 2024

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Keywords

  1. 2-aminoacrylate stress
  2. folate biosynthesis
  3. alanine racemase
  4. metabolic integration
  5. RidA

Data Availability

All relevant data are included in the content of this article.

Contributors

Authors

Department of Microbiology, University of Georgia, Athens, Georgia, USA
Author Contributions: Conceptualization, Formal analysis, Investigation, Methodology, Writing – original draft, and Writing – review and editing.
Department of Microbiology, University of Georgia, Athens, Georgia, USA
Author Contributions: Conceptualization, Formal analysis, Funding acquisition, Methodology, Resources, Supervision, and Writing – review and editing.

Editor

George O'Toole
Editor
Geisel School of Medicine at Dartmouth, Hanover, New Hampshire, USA

Notes

The authors declare no conflict of interest.

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