Aminopterin inhibition in <i>Aerobacter aerogenes</i>: alanine and valine accumulation during the inhibition and their utilization on recovery

Webb, M.

doi:10.1042/bj0700472

Cited by 28 publications

(9 citation statements)

References 87 publications

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“…Since this induction requires high intracellular alanine (24), it implies that pyruvate increases intracellular alanine. Third, a folic acid antagonist results in accumulation of pyruvate and alanine in Klebsiella pneumoniae (formerly Aerobacter aerogenes and Klebsiella aerogenes), and these studies were extended to show that several antibiotics and a variety of growth transitions also result in accumulation of pyruvate, and possibly alanine (37,38). If pyruvate accumulation increases the concentrations of alanine and possibly valine (via AvtA), then changes in pyruvate concentration might account for reports of alanine and valine accumulation (33,34).…”

Section: Discussionmentioning

confidence: 99%

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Genetics and Regulation of the Major Enzymes of Alanine Synthesis inEscherichia coli

2010

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Genetic analysis of alanine synthesis in the model genetic organism Escherichia coli has implicated avtA, the still uncharacterized alaA and alaB genes, and probably other genes. We identified alaA as yfbQ. We then transferred mutations in several transaminase genes into a yfbQ mutant and isolated a mutant that required alanine for optimal growth. For cells grown with carbon sources other than pyruvate, the major alaninesynthesizing transaminases are AvtA, YfbQ (AlaA), and YfdZ (which we designate AlaC). Growth with pyruvate as the carbon source and multicopy suppression suggest that several other transaminases can contribute to alanine synthesis. Expression studies showed that alanine modestly repressed avtA and yfbQ but had no effect on yfdZ. The leucine-responsive regulatory protein (Lrp) mediated control by alanine. We purified YfbQ and YfdZ and showed that both are dimers with K m s for pyruvate within the intracellular range of pyruvate concentration.

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Section: Discussionmentioning

confidence: 99%

“…The concentrations of pyruvate, alanine, and glutamate vary with environmental changes and growth rate (9,14,20,31,32,34,37,38). Glutamate is usually the most abundant amino acid in pools, and alanine the second most abundant (23,32,42), although for phosphate limitation, the alanine pool appears to exceed the glutamate pool (32).…”

Section: Discussionmentioning

confidence: 99%

Genetics and Regulation of the Major Enzymes of Alanine Synthesis inEscherichia coli

2010

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“…Under aerobic conditions, the central metabolite pyruvate leads to acetyl-coenzyme A (acetyl-CoA) and the citric acid cycle, while under anaerobic conditions, in addition to forming acetyl-CoA, it is the precursor of most fermentation products. Because pyruvate is a precursor of valine (the condensation of two pyruvates leads to one valine) and L-alanine (by transamination), Webb's studies suggest that in the absence of a limited source of carbon, reduced growth or oxygen rarefaction may slow down the citric acid (Kreb's) cycle and consequently lead to an increased drain of pyruvate toward biosynthesis of Lalanine and L-valine, both of which are increased in the CFT073 in-biofilm supernatant (55,56).…”

Section: Discussionmentioning

confidence: 99%

“…These conditions are reminiscent of those within biofilms, raising the possibility that in-biofilm valine production may be a consequence of complex alterations of metabolic pools within biofilms. Webb reported previously that the growth inhibition of Klebsiella pneumoniae by aminopterin led to the production of valine and L-alanine as a metabolic consequence of pyruvate accumulation (55,56). Aminopterin is an inhibitor of dihydrofolate reductase and thus of tetrahydrofolate, a precursor of all one-carbon units.…”

Section: Discussionmentioning

confidence: 99%

The Amino Acid Valine Is Secreted in Continuous-Flow Bacterial Biofilms

Valle

Schmid

et al. 2008

J Bacteriol

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Biofilms are structured communities characterized by distinctive gene expression patterns and profound physiological changes compared to those of planktonic cultures. Here, we show that many gram-negative bacterial biofilms secrete high levels of a small-molecular-weight compound, which inhibits the growth of only Escherichia coli K-12 and a rare few other natural isolates. We demonstrate both genetically and biochemically that this molecule is the amino acid valine, and we provide evidence that valine production within biofilms results from metabolic changes occurring within high-density biofilm communities when carbon sources are not limiting. This finding identifies a natural environment in which bacteria can encounter high amounts of valine, and we propose that in-biofilm valine secretion may be the long-sought reason for widespread but unexplained valine resistance found in most enterobacteria. Our results experimentally validate the postulated production of metabolites that is characteristic of the conditions associated with some biofilm environments. The identification of such molecules may lead to new approaches for biofilm monitoring and control.Biofilms are matrix-encased bacterial communities that develop on most surfaces and often constitute a reservoir of bacterial pathogens. Detrimental biofilms are difficult to eradicate due to a characteristic tolerance to biocides. They cause serious health and economic problems when they form on medical and industrial devices (11,19,24). While biofilmassociated antibiotic tolerance is considered a major physiological trait that distinguishes free cells from surface-attached cells, biofilms are also characterized by gene expression patterns that are distinctive compared to those of planktonic cultures (4,35,40,41,58). These changes are likely to result from modifications of growth conditions within biofilms, and they have been proposed to correspond to ill-understood responses triggered by the biofilm lifestyle (3,26).Recently, studies conducted with simplified, mixed communities composed of two bacterial species revealed that biofilm bacteria express competitive or cooperative behavior that does not take place within classical planktonic cultures (7,22). These results suggested that biofilm-associated weaponry could contribute to the dynamics of bacterial biofilm populations. However, thus far, the production of molecules involved in antagonistic bacterial relationships within biofilms has been poorly investigated.Here we show that the continuous-flow biofilms formed by many Escherichia coli strains and other gram-negative bacteria accumulate high levels of a small-molecular-weight compound with inhibitory activity against E. coli K-12 strains. We demonstrate both genetically and biochemically that this compound is the amino acid valine. We provide evidence that valine secretion within biofilms could be the cause of the longknown but unexplained widespread valine resistance observed for most enterobacteria. Beyond the biological significance of in-biofilm valin...

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“…To this end, the compound, 5-aminoimidazole-4-carboxamide-1-␤ribofuranoside (AICA riboside), was utilized. AICA riboside can be phosphorylated to form AICAR, an interme-several folate antagonists that act at the sites of folate utilization, phosphoribosylglycinamide transformylase, diate in the de novo pathway that is situated at a point distal to the presumed sites of inhibition of azaserine and AICAR transformylase (Webb, 1958;Hakala and Taylor, 1959;McGeer and McGeer, 1963; Baram et al, (see Fig. 1).…”

Section: Role Of De Novo Purine Synthesis and Purinementioning

confidence: 99%

Involvement of purine nucleotide synthetic pathways in gonadotropin-induced meiotic maturation in mouse cumulus cell-enclosed oocytes

Downs

1997

Mol. Reprod. Dev.

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This study was carried out to test the hypothesis that purine nucleotide‐generating pathways are required for ligand‐stimulated oocyte maturation in meiotically arrested cumulus cell‐enclosed oocytes. Oo‐cytes from hormonally primed, immature mice were cultured overnight in Eagle's minimum essential medium containing dibutyryl cyclic AMP (dbcAMP) (to maintain meiotic arrest), plus either mycophenolic acid or alanosine (inhibitors of guanyl and adenyl nucleotide production, respectively). Follicle‐stimulating hormone (FSH) was added either at the outset of culture or after a 3‐hr preincubation period. Under either of these conditions, the inhibitors suppressed FSH induction of germinal vesicle breakdown (GVB). In addition, the potency of FSH as an inducer of GVB was reduced following the 3‐hr preincubation period, but this could be prevented if nucleotide precursors such as hypoxanthine, guanosine, or adenosine were included during the first 3 hr. Furthermore, preincubation had little effect on FSH induction of GVB when hypoxanthine was used to maintain meiotic arrest for the entire culture period. The phosphodiesterase inhibitor, 3‐isobutyl‐l‐methylxanthine, could not mimic this protective effect of hypoxanthine. Azaserine and aminopterin, inhibitors of purine de novo synthesis, blocked hormone‐triggered maturation in dbcAMP‐arrested oocytes, but had little effect on hypoxanthine‐arrested oocytes. The effect of azaserine on dbcAMP‐treated oocytes could be reversed by the inclusion of AICA riboside, a compound that can be taken up by cells and phosphorylated to form AICAR, which can enter the purine de novo pathway at a point distal to the sites of azaserine inhibition. FSH was stimulatory to purine de novo synthesis, while azaserine, aminopterin, hypoxanthine, and AICA riboside all suppressed de novo synthesis in the presence or absence of FSH, with dbcAMP having no effect. HPLC analysis of 14C‐hypoxanthine metabolism in oocyte‐cumulus cell complexes revealed that changes in the pattern of purine metabolism did not mediate the meiosis‐inducing effect of FSH. These data support the conclusion that purine nucleotide‐generating pathways are vital participants in the mechanism(s) regulating hormone‐induced meiotic maturation, and that either the de novo or salvage pathway can fulfill this nucleotide requirement. Mol Reprod Dev 46:155–167, 1997. © 1997 Wiley‐Liss, Inc.

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Aminopterin inhibition in Aerobacter aerogenes: alanine and valine accumulation during the inhibition and their utilization on recovery

Cited by 28 publications

References 87 publications

Genetics and Regulation of the Major Enzymes of Alanine Synthesis inEscherichia coli

Genetics and Regulation of the Major Enzymes of Alanine Synthesis inEscherichia coli

The Amino Acid Valine Is Secreted in Continuous-Flow Bacterial Biofilms

Involvement of purine nucleotide synthetic pathways in gonadotropin-induced meiotic maturation in mouse cumulus cell-enclosed oocytes

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