Genetic and biochemical analyses of Escherichia coli strains having a mutation in the structural gene (poxB) for pyruvate oxidase

Chang, Ying; Cronan, John E.

doi:10.1128/jb.154.2.756-762.1983

Cited by 61 publications

(21 citation statements)

References 25 publications

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“…Under laboratory conditions, E. coti Pox is a nonessential enzyme, and poxB null mutant strains grow normally (Chang and Cronan, 1983). However, our finding that poxB expression is tightly controlled by the rpoS gene argues that PoxB may be important for cell survival during the transition between the exponential and stationary grovrth phases.…”

Section: Discussionmentioning

confidence: 82%

Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene

Chang

Wang

Cronan

1994

Molecular Microbiology

111

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The activity of Escherichia coli pyruvate oxidase (PoxB) was shown to be growth-phase dependent; the enzyme activity reaches a maximum at early stationary phase. We report that PoxB activity is dependent on a functional rpoS(katF) gene which encodes a sigma factor required to transcribe a number of stationary-phase-induced genes. PoxB activity as well as the beta-galactosidase encoded by a poxB::lacZ protein fusion was completely abolished in a strain containing a defective rpoS gene. Northern and primer extension analyses showed that poxB expression was regulated at the transcriptional level and was transcribed from a single promoter; the 5' end of the mRNA being located 27 bp upstream of the translational initiation codon of poxB. The poxB gene was expressed at decreased levels under anaerobiosis; however, the anaerobic regulatory genes arcA, arcB or fnr were not involved in anaerobic poxB gene expression. Expression of the rpoS(katF) gene has been reported to be affected by acetate, the product of PoxB reaction. However, we found that poxB null mutations had no effect on rpoS(katF) expression. Inactivation of two genes involved in acetate metabolism, ackA and pta, had no effect on either poxB or rpoS(katF) expression.

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Section: Discussionmentioning

confidence: 82%

Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene

Chang

Wang

Cronan

1994

Molecular Microbiology

111

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“…In principle, carbon fluxes from pyruvate can be distributed over three enzymes: PFL, PDHc, and lactate dehydrogenase (LDH) (the flux through pyruvate oxidase being minor [2,5]). The absence of in vivo flux via the LDH in MC4100 and RM319 under anaerobic conditions is puzzling.…”

Section: Discussionmentioning

confidence: 99%

The Steady-State Internal Redox State (NADH/NAD) Reflects the External Redox State and Is Correlated with Catabolic Adaptation in Escherichia coli

et al. 1999

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Escherichia coli MC4100 was grown in anaerobic glucose-limited chemostat cultures, either in the presence of an electron acceptor (fumarate, nitrate, or oxygen) or fully fermentatively. The steady-state NADH/NAD ratio depended on the nature of the electron acceptor. Anaerobically, the ratio was highest, and it decreased progressively with increasing midpoint potential of the electron acceptor. Similarly, decreasing the dissolved oxygen tension resulted in an increased NADH/NAD ratio. As pyruvate catabolism is a major switch point between fermentative and respiratory behavior, the fluxes through the different pyruvate-consuming enzymes were calculated. Although pyruvate formate lyase (PFL) is inactivated by oxygen, it was inferred that the in vivo activity of the enzyme occurred at low dissolved oxygen tensions (DOT ≤ 1%). A simultaneous flux from pyruvate through both PFL and the pyruvate dehydrogenase complex (PDHc) was observed. In anaerobic cultures with fumarate or nitrate as an electron acceptor, a significant flux through the PDHc was calculated on the basis of the redox balance, the measured products, and the known biochemistry. This result calls into question the common assumption that the complex cannot be active under these conditions. In vitro activity measurements of PDHc showed that the cellular content of the enzyme varied with the internal redox state and revealed an activity for dissolved oxygen tension of below 1%. Whereas Western blots showed that the E3 subunit of PDHc (dihydrolipoamide dehydrogenase) did not vary to a large extent under the conditions tested, the E2 subunit (dihydrolipoamide acetyltransferase) amount followed the trend that was found for the in vitro PDHc activity. From this it is concluded that regulation of the PDHc is exerted at the E1/E2 operon (aceEF). We propose that the external redox state (measured as the midpoint potentials of those terminal acceptors with which the cell has sufficient capacity to react) is reflected by the internal redox state. The latter may subsequently govern both the expression and the activity of the two pyruvate-catabolizing enzymes.

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“…16 The pyruvate oxidase enzyme was later found to be encoded by an unlinked gene, poxB. 17 PoxA appears to be critical for optimal expression of PoxB but its role appears to be post-translational given that mRNA levels in poxA mutants are similar to wild-type. PoxA is an "orphan" member of the lysyl-tRNA synthetases (LysRS) family that shares a high degree of sequence and structural similarity with the catalytic domain of LysRS but lacks the canonical anticodon recognition domain.…”

Section: Introductionmentioning

confidence: 99%

Elongation factor P mediates a novel post-transcriptional regulatory pathway critical for bacterial virulence

et al. 2011

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Bacterial pathogens detect and integrate multiple environmental signals to coordinate appropriate changes in gene expression including the selective expression of virulence factors, changes to metabolism and the activation of stress response systems. Mutations that abolish the ability of the pathogen to respond to external cues are typically attenuating. Here we discuss our recent discovery of a novel post-transcriptional regulatory pathway critical for Salmonella virulence and stress resistance. The enzymes PoxA and YjeK coordinately attach a unique beta-amino acid onto a highly conserved lysine residue in the translation factor elongation factor P (EF-P). Strains in which EF-P is unmodified due to the absence of PoxA or YjeK are attenuated for virulence and display highly pleiotropic phenotypes, including hypersusceptibility to a wide range of unrelated antimicrobial compounds. Work from our laboratory and others now suggests that EF-P, previously thought to be essential, instead plays an ancillary role in translation by regulating the synthesis of a relatively limited subset of proteins. Other observations suggest that the eukaryotic homolog of EF-P, eIF5A, may illicit similar changes in the translation machinery during stress adaptation, indicating that the role of these factors in physiology may be broadly conserved.

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Genetic and biochemical analyses of Escherichia coli strains having a mutation in the structural gene (poxB) for pyruvate oxidase

Cited by 61 publications

References 25 publications

Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene

Expression of Escherichia coli pyruvate oxidase (PoxB) depends on the sigma factor encoded by the rpoS(katF) gene

The Steady-State Internal Redox State (NADH/NAD) Reflects the External Redox State and Is Correlated with Catabolic Adaptation in Escherichia coli

Elongation factor P mediates a novel post-transcriptional regulatory pathway critical for bacterial virulence

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