Levels of glutathione in Escherichia coli

1979 ◽  
Vol 57 (2) ◽  
pp. 107-111 ◽  
Author(s):  
Peter C. Loewen
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Stationary Phase ◽  
Minimal Medium ◽  
Anaerobic Growth ◽  
Glutathione Disulfide ◽  
Intracellular Pool ◽  
Glucose Depletion ◽  
Apparent Correlation ◽  
Cell Densities

Log phase cells of Escherichia coli growing in minimal medium contain a basal level of glutathione (5 pmol/mL per Klett unit) which can increase more than sixfold when the cells reach stationary phase. Since the addition of cysteine alone to log phase cells illicits the same response, the increase in the intracellular pool of glutathione appears to be influenced by the amount of cysteine available for glutathione synthesis. Glucose depletion at low cell densities resulted in a decrease in the glutathione pool while the addition of amino acids other than cysteine did not affect the glutathione pool. Depletion of ammonia or proline as the nitrogen source also resulted in a decrease in the glutathione pool to one-third of the original basal level as did a shift to anaerobic growth. The large glutathione pool in stationary phase cells dropped from 31.5 to 4.5 pmol/mL per Klett unit within 30 min of transfer to fresh medium. There was no apparent correlation between changes in the glutathione and coenzyme A – glutathione disulfide (CoASSG) pools after a variety of metabolic disruptions.

scholarly journals Cooperative Roles of Nitric Oxide-Metabolizing Enzymes To Counteract Nitrosative Stress in Enterohemorrhagic Escherichia coli

2019 ◽  
Vol 87 (9) ◽  
Author(s):  
Takeshi Shimizu ◽  
Akio Matsumoto ◽  
Masatoshi Noda
Keyword(s):  
Nitric Oxide ◽  
Escherichia Coli ◽  
Stationary Phase ◽  
Nitrosative Stress ◽  
Enterohemorrhagic Escherichia Coli ◽  
The Other ◽  
Anaerobic Growth ◽  
Bacterial Cells ◽  
Metabolizing Enzymes ◽  
Microaerobic Conditions

ABSTRACT Enterohemorrhagic Escherichia coli (EHEC) has at least three enzymes, NorV, Hmp, and Hcp, that act independently to lower the toxicity of nitric oxide (NO), a potent antimicrobial molecule. This study aimed to reveal the cooperative roles of these defensive enzymes in EHEC against nitrosative stress. Under anaerobic conditions, combined deletion of all three enzymes significantly increased the NO sensitivity of EHEC determined by the growth at late stationary phase; however, the expression of norV restored the NO resistance of EHEC. On the other hand, the growth of Δhmp mutant EHEC was inhibited after early stationary phase, indicating that NorV and Hmp play a cooperative role in anaerobic growth. Under microaerobic conditions, the growth of Δhmp mutant EHEC was inhibited by NO, indicating that Hmp is the enzyme that protects cells from NO stress under microaerobic conditions. When EHEC cells were exposed to a lower concentration of NO, the NO level in bacterial cells of Δhcp mutant EHEC was higher than those of the other EHEC mutants, suggesting that Hcp is effective at regulating NO levels only at a low concentration. These findings of a low level of NO in bacterial cells with hcp indicate that the NO consumption activity of Hcp was suppressed by Hmp at a low range of NO concentrations. Taken together, these results show that the cooperative effects of NO-metabolizing enzymes are regulated by the range of NO concentrations to which the EHEC cells are exposed.

Effect of ascorbate on oxygen uptake and growth of Escherichia coli B

1988 ◽  
Vol 34 (6) ◽  
pp. 822-824 ◽  
Author(s):  
Holly E. Richter ◽  
Jacek Switala ◽  
Peter C. Loewen
Keyword(s):  
Escherichia Coli ◽  
Oxygen Uptake ◽  
Electron Acceptor ◽  
Minimal Medium ◽  
Anaerobic Growth ◽  
Rich Medium ◽  
Terminal Electron Acceptor ◽  
Subsequent Change ◽  
Escherichia Coli B

The addition of ascorbate to aerobically growing cultures of Escherichia coli B caused only a short pause in growth and no subsequent change in the rate or extent of growth. The effect of ascorbate on oxygen uptake varied from inhibition in minimal medium to stimulation in rich medium. Cyanide-resistant growth and oxygen uptake were stimulated by ascorbate. Both the rate and extent of anaerobic growth were stimulated in proportion to the amount of ascorbate added when fumarate was the terminal electron acceptor. Ascorbate had no effect on any aspect of anaerobic growth in the absence of a terminal electron acceptor or in the presence of nitrate.

scholarly journals Pyruvate Formate Lyase and Acetate Kinase Are Essential for Anaerobic Growth of Escherichia coli on Xylose

2004 ◽  
Vol 186 (22) ◽  
pp. 7593-7600 ◽  
Author(s):  
Adnan Hasona ◽  
Youngnyun Kim ◽  
F. G. Healy ◽  
L. O. Ingram ◽  
K. T. Shanmugam
Keyword(s):  
Escherichia Coli ◽  
Minimal Medium ◽  
High Capacity ◽  
Redox Balance ◽  
Anaerobic Growth ◽  
Acetate Kinase ◽  
Anaerobic Conditions ◽  
Pyruvate Formate Lyase ◽  
E Coli ◽  
Xylose Transport

ABSTRACT During anaerobic growth of bacteria, organic intermediates of metabolism, such as pyruvate or its derivatives, serve as electron acceptors to maintain the overall redox balance. Under these conditions, the ATP needed for cell growth is derived from substrate-level phosphorylation. In Escherichia coli, conversion of glucose to pyruvate yields 2 net ATPs, while metabolism of a pentose, such as xylose, to pyruvate only yields 0.67 net ATP per xylose due to the need for one (each) ATP for xylose transport and xylulose phosphorylation. During fermentative growth, E. coli produces equimolar amounts of acetate and ethanol from two pyruvates, and these reactions generate one additional ATP from two pyruvates (one hexose equivalent) while still maintaining the overall redox balance. Conversion of xylose to acetate and ethanol increases the net ATP yield from 0.67 to 1.5 per xylose. An E. coli pfl mutant lacking pyruvate formate lyase cannot convert pyruvate to acetyl coenzyme A, the required precursor for acetate and ethanol production, and could not produce this additional ATP. E. coli pfl mutants failed to grow under anaerobic conditions in xylose minimal medium without any negative effect on their survival or aerobic growth. An ackA mutant, lacking the ability to generate ATP from acetyl phosphate, also failed to grow in xylose minimal medium under anaerobic conditions, confirming the need for the ATP produced by acetate kinase for anaerobic growth on xylose. Since arabinose transport by AraE, the low-affinity, high-capacity, arabinose/H+ symport, conserves the ATP expended in pentose transport by the ABC transporter, both pfl and ackA mutants grew anaerobically with arabinose. AraE-based xylose transport, achieved after constitutively expressing araE, also supported the growth of the pfl mutant in xylose minimal medium. These results suggest that a net ATP yield of 0.67 per pentose is only enough to provide for maintenance energy but not enough to support growth of E. coli in minimal medium. Thus, pyruvate formate lyase and acetate kinase are essential for anaerobic growth of E. coli on xylose due to energetic constraints.

scholarly journals Role of the sRNA GcvB in regulation of cycA in Escherichia coli

Microbiology ◽  
2009 ◽  
Vol 155 (1) ◽  
pp. 106-114 ◽  
Author(s):  
Sarah C. Pulvermacher ◽  
Lorraine T. Stauffer ◽  
George V. Stauffer
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Minimal Medium ◽  
Luria Bertani ◽  
Cell Physiology ◽  
Rt Pcr ◽  
Luria Bertani Broth ◽  
Increased Sensitivity ◽  
Transcriptional Fusions

In Escherichia coli, the gcvB gene encodes a small non-translated RNA that regulates several genes involved in transport of amino acids and peptides (including sstT, oppA and dppA). Microarray analysis identified cycA as an additional regulatory target of GcvB. The cycA gene encodes a permease for the transport of glycine, d-alanine, d-serine and d-cycloserine. RT-PCR confirmed that GcvB and the Hfq protein negatively regulate cycA mRNA in cells grown in Luria–Bertani broth. In addition, deletion of the gcvB gene resulted in increased sensitivity to d-cycloserine, consistent with increased expression of cycA. A cycA : : lacZ translational fusion confirmed that GcvB negatively regulates cycA expression in Luria–Bertani broth and that Hfq is required for the GcvB effect. GcvB had no effect on cycA : : lacZ expression in glucose minimal medium supplemented with glycine. However, Hfq still negatively regulated the fusion in the absence of GcvB. A set of transcriptional fusions of cycA to lacZ identified a sequence in cycA necessary for regulation by GcvB. Analysis of GcvB identified a region complementary to this region of cycA mRNA. However, mutations predicted to disrupt base-pairing between cycA mRNA and GcvB did not alter expression of cycA : : lacZ. A model for GcvB function in cell physiology is discussed.

scholarly journals Protective Action of ppGpp in Microcin J25-Sensitive Strains

2008 ◽  
Vol 190 (12) ◽  
pp. 4328-4334 ◽  
Author(s):  
María Fernanda Pomares ◽  
Paula A. Vincent ◽  
Ricardo N. Farías ◽  
Raúl A. Salomón
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Stationary Phase ◽  
Efflux Pump ◽  
Protective Action ◽  
Intracellular Concentration ◽  
Peptide Antibiotic ◽  
Intracellular Level ◽  
Toxic Level ◽  
Microcin J25

ABSTRACT As Escherichia coli strains enter the stationary phase of growth they become more resistant to the peptide antibiotic microcin J25. It is known that starvation for nutrients such as amino acids or glucose leads to increases in guanosine 3′,5′-bispyrophosphate (ppGpp) levels and that the intracellular concentration of this nucleotide increases as cells enter the stationary phase of growth. Therefore, we examined the effects of artificially manipulating the ppGpp levels on sensitivity to microcin J25. A direct correlation was found between ppGpp accumulation and microcin resistance. Our results indicate that the nucleotide is required to induce production of YojI, a chromosomally encoded efflux pump which, in turn, expels microcin from cells. This would maintain the intracellular level of the antibiotic below a toxic level.

scholarly journals Mutations Enhancing Amino Acid Catabolism Confer a Growth Advantage in Stationary Phase

1999 ◽  
Vol 181 (18) ◽  
pp. 5800-5807 ◽  
Author(s):  
Erik R. Zinser ◽  
Roberto Kolter
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Amino Acid ◽  
Stationary Phase ◽  
Carbon Starvation ◽  
Amino Acid Catabolism ◽  
Rich Media ◽  
Dying Cells

ABSTRACT Starved cultures of Escherichia coli undergo successive rounds of population takeovers by mutants of increasing fitness. These mutants express the growth advantage in stationary phase (GASP) phenotype. Previous work identified the rpoS819 allele as a GASP mutation allowing cells to take over stationary-phase cultures after growth in rich media (M. M. Zambrano, D. A. Siegele, M. A. Almirón, A. Tormo, and R. Kolter, Science 259:1757–1760, 1993). Here we have identified three new GASP loci from an aged rpoS819 strain: sgaA, sgaB, and sgaC. Each locus is capable of conferring GASP on therpoS819 parent, and they can provide successively higher fitnesses for the bacteria in the starved cultures. All four GASP mutations isolated thus far allow for faster growth on both individual and mixtures of amino acids. Each mutation confers a growth advantage on a different subset of amino acids, and these mutations act in concert to increase the overall catabolic capacity of the cell. We present a model whereby this enhanced ability to catabolize amino acids is responsible for the fitness gain during carbon starvation, as it may allow GASP mutants to outcompete the parental cells when growing on the amino acids released by dying cells.

scholarly journals Complex Physiology and Compound Stress Responses during Fermentation of Alkali-Pretreated Corn Stover Hydrolysate by an Escherichia coli Ethanologen

2012 ◽  
Vol 78 (9) ◽  
pp. 3442-3457 ◽  
Author(s):  
Michael S. Schwalbach ◽  
David H. Keating ◽  
Mary Tremaine ◽  
Wesley D. Marner ◽  
Yaoping Zhang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Escherichia Coli ◽  
Amino Acids ◽  
Stationary Phase ◽  
Gene Expression Profiling ◽  
Stress Responses ◽  
Expression Profiling ◽  
Content Type ◽  
E Coli ◽  
Cell Maintenance

ABSTRACTThe physiology of ethanologenicEscherichia coligrown anaerobically in alkali-pretreated plant hydrolysates is complex and not well studied. To gain insight into howE. coliresponds to such hydrolysates, we studied anE. coliK-12 ethanologen fermenting a hydrolysate prepared from corn stover pretreated by ammonia fiber expansion. Despite the high sugar content (∼6% glucose, 3% xylose) and relatively low toxicity of this hydrolysate,E. coliceased growth long before glucose was depleted. Nevertheless, the cells remained metabolically active and continued conversion of glucose to ethanol until all glucose was consumed. Gene expression profiling revealed complex and changing patterns of metabolic physiology and cellular stress responses during an exponential growth phase, a transition phase, and the glycolytically active stationary phase. During the exponential and transition phases, high cell maintenance and stress response costs were mitigated, in part, by free amino acids available in the hydrolysate. However, after the majority of amino acids were depleted, the cells entered stationary phase, and ATP derived from glucose fermentation was consumed entirely by the demands of cell maintenance in the hydrolysate. Comparative gene expression profiling and metabolic modeling of the ethanologen suggested that the high energetic cost of mitigating osmotic, lignotoxin, and ethanol stress collectively limits growth, sugar utilization rates, and ethanol yields in alkali-pretreated lignocellulosic hydrolysates.

fusB is an allele of nadD, encoding nicotinate mononucleotide adenylyltransferase in Escherichia coli

Microbiology ◽  
2003 ◽  
Vol 149 (9) ◽  
pp. 2427-2433 ◽  
Author(s):  
Martin Stancek ◽  
Leif A. Isaksson ◽  
Monica Rydén-Aulin
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Temperature Sensitivity ◽  
Minimal Medium ◽  
Synthetic Medium ◽  
Normal Growth ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
Atp Binding Site ◽  
Pleiotropic Phenotype

Isolation of the temperature-sensitive Escherichia coli mutant 72c has been described previously. The mutant allele was named fusB and causes a pleiotropic phenotype, the most striking features of which, besides temperature sensitivity, are the inability to grow on synthetic medium and supersensitivity to trimethoprim, an antibiotic that inhibits the C1 metabolism. This work shows that the fusB mutation is a frameshift mutation in the nadD gene that encodes nicotinate mononucleotide adenylyltransferase. The frameshift leads to a change of the last 10 amino acids and an addition of 17 amino acids. This lesion, renamed nadD72, leads to very little NAD+ and NADPH synthesis at the permissive temperature and essentially no synthesis at the non-permissive temperature. As a comparison, a new mutation in the nadD gene, with an amino acid change in the ATP-binding site, has been isolated. Its NAD+ synthesis is decreased at 30 °C but the level is still sufficient to support normal growth. At 42 °C, NAD+ synthesis is reduced further, which leads to temperature sensitivity on minimal medium. This mutation was designated nadD74. Thus, a small decrease in NAD+ levels affects ability to grow on minimal medium at 42 °C, while a large decrease leads to a more pleiotropic phenotype.

THE INFLUENCE OF TIME AND MEDIA CHANGES ON THE INDUCTION AND SUPPRESSION OF β-GALACTOSIDASE SYNTHESIS IN ESCHERICHIA COLI B

1967 ◽  
Vol 13 (3) ◽  
pp. 257-269
Author(s):  
S. J. Webb ◽  
Janet L. Walker
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Minimal Medium ◽  
Sole Source ◽  
Enzyme Synthesis ◽  
Induction Medium ◽  
Nucleic Acid Bases ◽  
Apparent Effect ◽  
Lag Period ◽  
Escherichia Coli B

When cells of Escherichia coli B were grown in a glucose – amino acid medium and then transferred to a minimal medium containing lactose or isopropyl-β-D-thiogalactopyranoside as a sole source of carbon, no induction of β-galactosidase occurred unless one or several amino acids were supplied. Of the amino acids tested, aspartic acid was the most effective and its ability to initiate the synthesis of the enzyme was increased by the addition of arginine. In the presence of these two, or all of the amino acids, there was a lag period of 10 min before enzyme synthesis occurred. The duration of the lag period was unaffected by the addition of nucleic acid bases or succinate to the induction medium. Succinate or glutamate partially inhibited the synthesis of the enzyme, whereas glucose, inositol, or chloramphenicol completely suppressed it. With the exception of that produced by chloramphenicol, inhibition was dependent on the time at which the inhibitor was added. If inhibitors were added after the 10-min lag period, they had no apparent effect until 45 min had elapsed. Cells transferred after 15 min from one induction medium to another displayed for 30 min the induction characteristics of the first medium. It appears that a process occurring during the early 15-min period determines the rate at which enzymes will be synthesized for the next 30 min and that the action of inhibitors is to prevent this process. The process seems to require intact DNA and amino acids and it is suggested that it determines the specificity and quantity of mRNA manufactured.

scholarly journals An N-Terminally Truncated RpoS (σS) Protein in Escherichia coli Is Active In Vivo and Exhibits Normal Environmental Regulation Even in the Absence of rpoS Transcriptional and Translational Control Signals

2002 ◽  
Vol 184 (12) ◽  
pp. 3167-3175 ◽  
Author(s):  
K. Rajkumari ◽  
J. Gowrishankar
Keyword(s):  
Escherichia Coli ◽  
Amino Acids ◽  
Rna Polymerase ◽  
Stationary Phase ◽  
Translational Control ◽  
Sigma Factor ◽  
Receptor Protein ◽  
Wild Type ◽  
Truncated Protein

ABSTRACT RpoS (σS) in Escherichia coli is a stationary-phase-specific primary sigma factor of RNA polymerase which is 330 amino acids long and belongs to the eubacterial σ70 family of proteins. Conserved domain 1.1 at the N-terminal end of σ70 has been shown to be essential for RNA polymerase function, and its deletion has been shown to result in a dominant-lethal phenotype. We now report that a σS variant with a deletion of its N-terminal 50 amino acids (σSΔ1-50), when expressed in vivo either from a chromosomal rpoS::IS10 allele (in rho mutant strains) or from a plasmid-borne arabinose-inducible promoter, is as proficient as the wild type in directing transcription from the proU P1 promoter; at three other σS-dependent promoters that were tested (osmY, katE, and csiD), the truncated protein exhibited a three- to sevenfold reduced range of activities. Catabolite repression at the csiD promoter (which requires both σS and cyclic AMP [cAMP]-cAMP receptor protein for its activity) was also preserved in the strain expressing σSΔ1-50. The intracellular content of σSΔ1-50 was regulated by culture variables such as growth phase, osmolarity, and temperature in the same manner as that described earlier for σS, even when the truncated protein was expressed from a template that possessed neither the transcriptional nor the translational control elements of wild-type rpoS. Our results indicate that, unlike that in σ70, the N-terminal domain in σS may not be essential for the protein to function as a sigma factor in vivo. Furthermore, our results suggest that the induction of σS-specific promoters in stationary phase and during growth under conditions of high osmolarity or low temperature is mediated primarily through the regulation of σS protein degradation.

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