Pterin and Folate Salvage. Plants and <i>Escherichia coli</i> Lack Capacity to Reduce Oxidized Pterins

Noiriel, Alexandre; Naponelli, Valeria; Gregory, Jesse F.; Hanson, Andrew D.

doi:10.1104/pp.106.093633

Cited by 24 publications

(34 citation statements)

References 37 publications

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“…Supporting the latter observation, we found that an E. coli GCHY-I mutant (which is unable to make pterins) can use the dihydro but not the oxidized forms of neopterin, monapterin, or 6-hydroxymethylpterin to support folate synthesis [60]. Futhermore, expression of a typical folM -like gene ( Xylella fastidiosa PD0677, Figure 5D, Table 2) from a plasmid did not enable this mutant to use oxidized pterins, indicating that – like FolM – the PD0677 gene product does not act on oxidized pterins (Figure 8).…”

Section: Resultsmentioning

confidence: 84%

“…Futhermore, expression of a typical folM -like gene ( Xylella fastidiosa PD0677, Figure 5D, Table 2) from a plasmid did not enable this mutant to use oxidized pterins, indicating that – like FolM – the PD0677 gene product does not act on oxidized pterins (Figure 8). In control experiments in which Leishmania PTR1 was expressed from a plasmid, the mutant was able to use oxidized pterins, confirming that it is oxidized pterin reduction (and not uptake) that is lacking in E. coli (Figure 8) [60]. …”

Section: Resultsmentioning

confidence: 91%

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Comparative genomics of bacterial and plant folate synthesis and salvage: predictions and validations

et al. 2007

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BackgroundFolate synthesis and salvage pathways are relatively well known from classical biochemistry and genetics but they have not been subjected to comparative genomic analysis. The availability of genome sequences from hundreds of diverse bacteria, and from Arabidopsis thaliana, enabled such an analysis using the SEED database and its tools. This study reports the results of the analysis and integrates them with new and existing experimental data.ResultsBased on sequence similarity and the clustering, fusion, and phylogenetic distribution of genes, several functional predictions emerged from this analysis. For bacteria, these included the existence of novel GTP cyclohydrolase I and folylpolyglutamate synthase gene families, and of a trifunctional p-aminobenzoate synthesis gene. For plants and bacteria, the predictions comprised the identities of a 'missing' folate synthesis gene (folQ) and of a folate transporter, and the absence from plants of a folate salvage enzyme. Genetic and biochemical tests bore out these predictions.ConclusionFor bacteria, these results demonstrate that much can be learnt from comparative genomics, even for well-explored primary metabolic pathways. For plants, the findings particularly illustrate the potential for rapid functional assignment of unknown genes that have prokaryotic homologs, by analyzing which genes are associated with the latter. More generally, our data indicate how combined genomic analysis of both plants and prokaryotes can be more powerful than isolated examination of either group alone.

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Section: Resultsmentioning

confidence: 84%

Section: Resultsmentioning

confidence: 91%

Comparative genomics of bacterial and plant folate synthesis and salvage: predictions and validations

et al. 2007

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“…The hypothesis described above was tested first in E. coli, because E. coli has H 4 -MPt as a prominent intracellular pterin (21) and secretes large amounts of monapterin in some form (40). The reduction state of the secreted form is uncertain, because H 4 -MPt and H 2 -MPt can spontaneously oxidize to MPt and are expected to do so once released to the medium (29,34). A folX deletant was constructed, and its intra-and extracellular pterin and folate profiles were compared to those of the parent wild-type strain.…”

Section: Resultsmentioning

confidence: 99%

FolX and FolM Are Essential for Tetrahydromonapterin Synthesis in Escherichia coli and Pseudomonas aeruginosa

Pribat¹,

Blaby²,

Lara‐Núñez

et al. 2010

J Bacteriol

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Tetrahydromonapterin is a major pterin in Escherichia coli and is hypothesized to be the cofactor for phenylalanine hydroxylase (PhhA) in Pseudomonas aeruginosa, but neither its biosynthetic origin nor its cofactor role has been clearly demonstrated. A comparative genomics analysis implicated the enigmatic folX and folM genes in tetrahydromonapterin synthesis via their phyletic distribution and chromosomal clustering patterns. folX encodes dihydroneopterin triphosphate epimerase, which interconverts dihydroneopterin triphosphate and dihydromonapterin triphosphate. folM encodes an unusual short-chain dehydrogenase/ reductase known to have dihydrofolate and dihydrobiopterin reductase activity. The roles of FolX and FolM were tested experimentally first in E. coli, which lacks PhhA and in which the expression of P. aeruginosa PhhA plus the recycling enzyme pterin 4a-carbinolamine dehydratase, PhhB, rescues tyrosine auxotrophy. This rescue was abrogated by deleting folX or folM and restored by expressing the deleted gene from a plasmid. The folX deletion selectively eliminated tetrahydromonapterin production, which far exceeded folate production. Purified FolM showed high, NADPH-dependent dihydromonapterin reductase activity. These results were substantiated in P. aeruginosa by deleting tyrA (making PhhA the sole source of tyrosine) and folX. The ⌬tyrA strain was, as expected, prototrophic for tyrosine, whereas the ⌬tyrA ⌬folX strain was auxotrophic. As in E. coli, the folX deletant lacked tetrahydromonapterin. Collectively, these data establish that tetrahydromonapterin formation requires both FolX and FolM, that tetrahydromonapterin is the physiological cofactor for PhhA, and that tetrahydromonapterin can outrank folate as an end product of pterin biosynthesis.

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“…We recently introduced tomato (Solanum lycopersicum) as a model to explore folate metabolism and its engineering (Díaz de la Garza et al, 2007;Noiriel et al, 2007). Accordingly, in this study, we characterized tomato GGH genes and their recombinant products, emphasizing heterodimerization and dimer stability, and investigated heterodimer formation in planta.…”

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Tomato γ-Glutamylhydrolases: Expression, Characterization, and Evidence for Heterodimer Formation

et al. 2008

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Folates typically have γ-linked polyglutamyl tails that make them better enzyme substrates and worse transport substrates than the unglutamylated forms. The tail can be shortened or removed by the vacuolar enzyme γ-glutamyl hydrolase (GGH). It is known that GGH is active only as a dimer and that plants can have several GGH genes whose homodimeric products differ functionally. However, it is not known whether GGH dimers dissociate under in vivo conditions, whether heterodimers form, or how heterodimerization impacts enzyme activity. These issues were explored using the GGH system of tomato (Solanum lycopersicum). Tomato has three GGH genes that, like those in other eudicots, apparently diverged recently. LeGGH1 and LeGGH2 are expressed in fruit and all other organs, whereas LeGGH3 is expressed mainly in flower buds. LeGGH1 and LeGGH2 homodimers differ in bond cleavage preference; the LeGGH3 homodimer is catalytically inactive. Homodimers did not dissociate in physiological conditions. When coexpressed in Escherichia coli, LeGGH1 and LeGGH2 formed heterodimers with an intermediate bond cleavage preference, whereas LeGGH3 formed heterodimers with LeGGH1 or LeGGH2 that had one-half the activity of the matching homodimer. E. coli cells expressing LeGGH2 showed approximately 85% reduction in folate polyglutamates, but cells expressing LeGGH3 did not, confirming that LeGGH2 can function in vivo and LeGGH3 cannot. The formation of LeGGH1-LeGGH2 heterodimers was demonstrated in planta using bimolecular fluorescence complementation. Plant GGH heterodimers thus appear to form wherever different GGH genes are expressed simultaneously and to have catalytic characteristics midway between those of the corresponding homodimers.

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Pterin and Folate Salvage. Plants and Escherichia coli Lack Capacity to Reduce Oxidized Pterins

Cited by 24 publications

References 37 publications

Comparative genomics of bacterial and plant folate synthesis and salvage: predictions and validations

Comparative genomics of bacterial and plant folate synthesis and salvage: predictions and validations

FolX and FolM Are Essential for Tetrahydromonapterin Synthesis in Escherichia coli and Pseudomonas aeruginosa

Tomato γ-Glutamylhydrolases: Expression, Characterization, and Evidence for Heterodimer Formation

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