Regulation of catalase level in Escherichia coli K12

1977 ◽  
Vol 23 (1) ◽  
pp. 84-91 ◽  
Author(s):  
Yona Yoshpe-Purer ◽  
Yigal Henis ◽  
Jacob Yashphe
Keyword(s):  
Escherichia Coli ◽  
Catabolite Repression ◽  
Nitrogen Starvation ◽  
Carbon Sources ◽  
Tca Cycle ◽  
Enzyme Synthesis ◽  
Anaerobic Growth ◽  
Nutrient Agar ◽  
Tca Cycle Intermediates ◽  
Transient Repression

The synthesis of catalase (H2O2:H2O2 oxidoreductase, EC 1.11.1.6) in Escherichia coli K12 was regulated by repression–induction and catabolite repression. Transient repression by glucose was also demonstrated. Under certain conditions, preferential synthesis was observed. Enzyme synthesis was induced in glycerol-grown cells by 0.15 to 0.2 μmol H2O2 (5–6 μg) per millilitre of culture, added at 10 to 15-min intervals, which were shortened progressively as the level of catalase in the cells rose. Catabolite repression of catalase synthesis was demonstrated with glucose, glycerol, maltose, lactose, xylose, mannitol, sorbitol, and trehalose, and to a lesser extent with arabinose, galactose, sucrose, rhamnose, and dulcitol as sole carbon sources in mineral medium. Catabolite repression was prevented by anaerobic growth and nitrogen starvation. All TCA-cycle intermediates examined afforded high catalase levels when serving as sole carbon sources. High catalase levels were also obtained when cells were grown on nutrient broth, nutrient agar, casitone, and peptone.

scholarly journals Catabolite Repression Control of napF (Periplasmic Nitrate Reductase) Operon Expression in Escherichia coli K-12

2008 ◽  
Vol 191 (3) ◽  
pp. 996-1005 ◽  
Author(s):  
Valley Stewart ◽  
Peggy J. Bledsoe ◽  
Li-Ling Chen ◽  
Amie Cai
Keyword(s):  
Escherichia Coli ◽  
Carbon Source ◽  
Nitrate Reductase ◽  
Catabolite Repression ◽  
Carbon Sources ◽  
Anaerobic Growth ◽  
Periplasmic Nitrate Reductase ◽  
Membrane Bound ◽  
Operon Expression ◽  
K 12

ABSTRACT Escherichia coli, a facultative aerobe, expresses two distinct respiratory nitrate reductases. The periplasmic NapABC enzyme likely functions during growth in nitrate-limited environments, whereas the membrane-bound NarGHI enzyme functions during growth in nitrate-rich environments. Maximal expression of the napFDAGHBC operon encoding periplasmic nitrate reductase results from synergistic transcription activation by the Fnr and phospho-NarP proteins, acting in response to anaerobiosis and nitrate or nitrite, respectively. Here, we report that, during anaerobic growth with no added nitrate, less-preferred carbon sources stimulated napF operon expression by as much as fourfold relative to glucose. Deletion analysis identified a cyclic AMP receptor protein (Crp) binding site upstream of the NarP and Fnr sites as being required for this stimulation. The napD and nrfA operon control regions from Shewanella spp. also have apparent Crp and Fnr sites, and expression from the Shewanella oneidensis nrfA control region cloned in E. coli was subject to catabolite repression. In contrast, the carbon source had relatively little effect on expression of the narGHJI operon encoding membrane-bound nitrate reductase under any growth condition tested. Carbon source oxidation state had no influence on synthesis of either nitrate reductase. The results suggest that the Fnr and Crp proteins may act synergistically to enhance NapABC synthesis during growth with poor carbon sources to help obtain energy from low levels of nitrate.

scholarly journals Identification of Enzymatic Bottlenecks for the Aerobic Production of Malate from Glycerol by the Systematic Gene Overexpression of Anaplerotic Enzymes in Escherichia coli

2021 ◽  
Vol 22 (5) ◽  
pp. 2266
Author(s):  
Zamira E. Soto-Varela ◽  
Gema Cabrera ◽  
Agustin Romero ◽  
Domingo Cantero ◽  
Antonio Valle ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Anaerobic Fermentation ◽  
Waste Product ◽  
Carbon Sources ◽  
Dicarboxylic Acids ◽  
Tca Cycle ◽  
Biotechnological Production ◽  
Phosphoenol Pyruvate ◽  
Valuable Compounds ◽  
C4 Production

The biotechnological production of dicarboxylic acids (C4) from renewable carbon sources represents an attractive approach for the provision of these valuable compounds by green chemistry means. Glycerol has become a waste product of the biodiesel industry that serves as a highly reduced carbon source for some microorganisms. Escherichia coli is capable of consuming glycerol to produce succinate under anaerobic fermentation, but with the deletion of some tricarboxylic acid (TCA) cycle genes, it is also able to produce succinate and malate in aerobiosis. In this study, we investigate possible rate-limiting enzymes by overexpressing the C-feeding anaplerotic enzymes Ppc, MaeA, MaeB, and Pck in a mutant that lacks the succinate dehydrogenase (Sdh) enzyme. The overexpression of the TCA enzyme Mdh and the activation of the glyoxylate shunt was also examined. Using this unbiased approach, we found that phosphoenol pyruvate carboxylase (Ppc) overexpression enhances an oxidative pathway that leads to increasing succinate, while phosphoenol pyruvate carboxykinase (Pck) favors a more efficient reductive branch that produces mainly malate, at 57.5% of the theoretical maximum molar yield. The optimization of the culture medium revealed the importance of bicarbonate and pH in the production of malate. An additional mutation of the ppc gene highlights its central role in growth and C4 production.

scholarly journals Procyclic trypanosomes recycle glucose catabolites and TCA cycle intermediates to stimulate growth in the presence of physiological amounts of proline

2021 ◽  
Vol 17 (3) ◽  
pp. e1009204
Author(s):  
Oriana Villafraz ◽  
Marc Biran ◽  
Erika Pineda ◽  
Nicolas Plazolles ◽  
Edern Cahoreau ◽  
...  
Keyword(s):  
Carbon Sources ◽  
Tca Cycle ◽  
Central Carbon Metabolism ◽  
Tsetse Fly ◽  
Growth Defect ◽  
Insect Vector ◽  
Metabolic Intermediates ◽  
The Tca Cycle ◽  
Production And Consumption ◽  
Tca Cycle Intermediates

Trypanosoma brucei, a protist responsible for human African trypanosomiasis (sleeping sickness), is transmitted by the tsetse fly where the procyclic forms of the parasite develop in the proline-rich (1–2 mM) and glucose-depleted digestive tract. Proline is essential for the midgut colonization of the parasite in the insect vector, however other carbon sources could be available and used to feed its central metabolism. Here we show that procyclic trypanosomes can consume and metabolize metabolic intermediates, including those excreted from glucose catabolism (succinate, alanine and pyruvate), with the exception of acetate, which is the ultimate end-product excreted by the parasite. Among the tested metabolites, tricarboxylic acid (TCA) cycle intermediates (succinate, malate and α-ketoglutarate) stimulated growth of the parasite in the presence of 2 mM proline. The pathways used for their metabolism were mapped by proton-NMR metabolic profiling and phenotypic analyses of thirteen RNAi and/or null mutants affecting central carbon metabolism. We showed that (i) malate is converted to succinate by both the reducing and oxidative branches of the TCA cycle, which demonstrates that procyclic trypanosomes can use the full TCA cycle, (ii) the enormous rate of α-ketoglutarate consumption (15-times higher than glucose) is possible thanks to the balanced production and consumption of NADH at the substrate level and (iii) α-ketoglutarate is toxic for trypanosomes if not appropriately metabolized as observed for an α-ketoglutarate dehydrogenase null mutant. In addition, epimastigotes produced from procyclics upon overexpression of RBP6 showed a growth defect in the presence of 2 mM proline, which is rescued by α-ketoglutarate, suggesting that physiological amounts of proline are not sufficient per se for the development of trypanosomes in the fly. In conclusion, these data show that trypanosomes can metabolize multiple metabolites, in addition to proline, which allows them to confront challenging environments in the fly.

scholarly journals Effect of point mutations in the lac promoter on transient and severe catabolite repression of the lac operon of Escherichia coli

1971 ◽  
Vol 123 (4) ◽  
pp. 579-584 ◽  
Author(s):  
M. D. Yudkin
Keyword(s):  
Escherichia Coli ◽  
Catabolite Repression ◽  
Point Mutations ◽  
Severe Form ◽  
Lac Operon ◽  
Wild Type ◽  
Diploid Strain ◽  
Transient Repression ◽  
Lac Promoter

1. Experiments were devised to show whether the point mutations L8 and L29 in the lac promoter alleviate transient repression. 2. Several recombinants were picked from matings between a single F−p+strain and Hfr strains carrying mutations L8 and L29. All of the 19 p−recombinants tested proved to suffer no transient repression, whereas all of the eight p+recombinants tested suffered prolonged transient repression. 3. A diploid strain was constructed in which more than 90% of the thiogalactoside transacetylase is synthesized from the episome with a wild-type lac promoter, whereas 100% of the β-galactosidase is synthesized from the chromosome with a promoter carrying mutation L8. In this diploid the synthesis of thiogalactoside transacetylase suffered transient repression but the synthesis of β-galactosidase did not. 4. Exactly similar results were obtained with a diploid strain in which the chromosomal promoter carried mutation L29. 5. The same diploid strains were used in experiments to show whether mutations L8 and L29 alleviate the severe catabolite repression caused by growth in glucose plus gluconate. In both strains glucose+gluconate repressed the synthesis of β-galactosidase much less than the synthesis of thiogalactoside transacetylase. 6. These and previously reported results can be explained by assuming (a) that both mutations L8 and L29 render the lac promoter partially, but not completely, insensitive to catabolite repression, and (b) that transient repression is an exceptionally severe form of catabolite repression.

scholarly journals The role of the regulator-gene product (repressor) in catabolite repression of β-galactosidase synthesis in Escherichia coli

1968 ◽  
Vol 106 (2) ◽  
pp. 339-343 ◽  
Author(s):  
J. Palmer ◽  
V. Moses
Keyword(s):  
Escherichia Coli ◽  
Gene Product ◽  
Catabolite Repression ◽  
Lac Repressor ◽  
Temperature Sensitive ◽  
Sensitive Mutant ◽  
Amber Suppressor ◽  
Transient Repression ◽  
I Gene

1. The specific role of the lac repressor (i-gene product) in transient catabolite repression evoked by the introduction of glucose into the medium has been investigated in Escherichia coli by using mutants of the i-gene. 2. A temperature-sensitive mutant (iTL) is normally inducible and demonstrates transient repression when grown at 32°. At 42° it is about 20% constitutive and transient catabolite repression is abolished. 3. A strain carrying an amber suppressor-sensitive mutation in the i-gene is phenotypically constitutive and also fails to show transient catabolite repression. 4. Insertion of Flaci+ into this strain restores both inducibility and transient repression. 5. It is concluded that the i-gene product interacts with the catabolite co-repressor in such a way that its affinity for the operator is increased.

scholarly journals Use of a Bacterial Luciferase Monitoring System To Estimate Real-Time Dynamics of Intracellular Metabolism in Escherichia coli

2016 ◽  
Vol 82 (19) ◽  
pp. 5960-5968 ◽  
Author(s):  
Tomohiro Shimada ◽  
Kan Tanaka
Keyword(s):  
Escherichia Coli ◽  
Real Time ◽  
Monitoring System ◽  
Carbon Sources ◽  
Tca Cycle ◽  
Bacterial Cells ◽  
Bacterial Luciferase ◽  
Content Type ◽  
Time Dynamics ◽  
The Tca Cycle

ABSTRACTRegulation of central carbon metabolism has long been an important research subject in every organism. While the dynamics of metabolic flows during changes in available carbon sources have been estimated based on changes in metabolism-related gene expression, as well as on changes in the metabolome, the flux change itself has scarcely been measured because of technical difficulty, which has made conclusions elusive in many cases. Here, we used a monitoring system employingVibrio fischeriluciferase to probe the intracellular metabolic condition inEscherichia coli. Using a batch culture provided with a limited amount of glucose, we performed a time course analysis, where the predominant carbon source shifts from glucose to acetate, and identified a series of sequential peaks in the luciferase activity (peaks 1 to 4). Two major peaks, peaks 1 and 3, were considered to correspond to the glucose and acetate consuming phases, respectively, based on the glucose, acetate, and dissolved oxygen concentrations in the medium. The pattern of these peaks was changed by the addition of a different carbon source or by an increasing concentration of glucose, which was consistent with the present model. Genetically, mutations involved in glycolysis or the tricarboxylic acid (TCA) cycle/gluconeogenesis specifically affected peak 1 or peak 3, respectively, as expected from the corresponding metabolic phase. Intriguingly, mutants for the acetate excretion pathway showed a phenotype of extended peak 2 and delayed transition to the TCA cycle/gluconeogenesis phase, which suggests that peak 2 represents the metabolic transition phase. These results indicate that the bacterial luciferase monitoring system is useful to understand the real-time dynamics of metabolism in living bacterial cells.IMPORTANCEIntracellular metabolic flows dynamically change during shifts in available carbon sources. However, because of technical difficulty, the flux change has scarcely been measured in living cells. Here, we used aVibrio fischeriluciferase monitoring system to probe the intracellular metabolic condition inEscherichia coli. Using a limited amount of glucose batch culture, a series of sequential peaks (peaks 1 to 4) in the luciferase activity was observed. Changes in the pattern of these peaks by the addition of extra carbon sources and in mutant strains involved in glycolysis or the TCA cycle/gluconeogenesis gene assigned the metabolic phase corresponding to peak 1 as the glycolysis phase and peak 3 as the TCA cycle/gluconeogenesis phase. Intriguingly, the acetate excretion pathway engaged in peak 2 represents the metabolic transition phase. These results indicate that the bacterial luciferase monitoring system is useful to understand the real-time dynamics of metabolism in living bacterial cells.

scholarly journals Fly stage trypanosomes recycle glucose catabolites and TCA cycle intermediates to stimulate growth in near physiological conditions

2020 ◽  
Author(s):  
Oriana Villafraz ◽  
Marc Biran ◽  
Erika Pineda ◽  
Nicolas Plazolles ◽  
Edern Cahoreau ◽  
...  
Keyword(s):  
Carbon Sources ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Tsetse Fly ◽  
Growth Defect ◽  
Insect Vector ◽  
Metabolic Potential ◽  
The Tca Cycle ◽  
Organ Systems ◽  
Tca Cycle Intermediates

AbstractTrypanosoma brucei, a protist responsible for human African trypanosomiasis (sleeping sickness), is transmitted by the tsetse fly, where the procyclic forms of the parasite develop in the proline-rich (1-2 mM) and glucose-depleted digestive tract. Proline is essential for the midgut colonization of the parasite in the insect vector, however other carbon sources could be available and used to feed its central metabolism. Here we show that procyclic trypanosomes can consume and metabolize metabolic intermediates, including those excreted from glucose catabolism (succinate, alanine and pyruvate), with the exception of acetate, which is the ultimate end-product excreted by the parasite. Among the tested metabolites, tricarboxylic acid (TCA) cycle intermediates (succinate, malate and α-ketoglutarate) stimulated growth of the parasite in the presence of 2 mM proline. The pathways used for their metabolism were mapped by proton-NMR metabolic profiling and phenotypic analyses of a dozen RNAi and/or null mutants affecting central carbon metabolism. We showed that (i) malate is converted to succinate by both the reducing and oxidative branches of the TCA cycle, which demonstrates that procyclic trypanosomes can use the full TCA cycle, (ii) the enormous rate of α-ketoglutarate consumption (15-times higher than glucose) is possible thanks to the balanced production and consumption of NADH at the substrate level and (iii) α-ketoglutarate is toxic for trypanosomes if not appropriately metabolized as observed for an α-ketoglutarate dehydrogenase null mutant. In addition, epimastigotes produced from procyclics upon overexpression of RBP6, showed a growth defect in the presence of 2 mM proline, which is rescued by α-ketoglutarate, suggesting that physiological amounts of proline are not sufficient per se for the development of trypanosomes in the fly. In conclusion, these data show that trypanosomes can metabolize multiple metabolites, in addition to proline, which allows them to confront challenging environments in the fly.Author SummaryIn the midgut of its insect vector, trypanosomes rely on proline to feed their energy metabolism. However, the availability of other potential carbon sources that can be used by the parasite is currently unknown. Here we show that tricarboxylic acid (TCA) cycle intermediates, i.e. succinate, malate and α-ketoglutarate, stimulate growth of procyclic trypanosomes incubated in medium containing 2 mM proline, which is in the range of the amounts measured in the midgut of the fly. Some of these additional carbon sources are needed for the development of epimastigotes, which differentiate from procyclics in the midgut of the fly, since their growth defect observed in the presence of 2 mM proline is rescued by addition of α-ketoglutarate. In addition, we have implemented new approaches to study a poorly explored branch of the TCA cycle converting malate to α-ketoglutarate, which was previously described as non-functional in the parasite, regardless of the glucose levels available. The discovery of this branch reveals that a full TCA cycle can operate in procyclic trypanosomes. Our data broaden the metabolic potential of trypanosomes and pave the way for a better understanding of the parasite’s metabolism in various organ systems of the tsetse fly, where it evolves.

scholarly journals The PalkBFGHJKL Promoter Is under Carbon Catabolite Repression Control in Pseudomonas oleovorans but Not in Escherichia coli alk+Recombinants

1999 ◽  
Vol 181 (5) ◽  
pp. 1610-1616 ◽  
Author(s):  
Ivo E. Staijen ◽  
Rosanna Marcionelli ◽  
Bernard Witholt
Keyword(s):  
Escherichia Coli ◽  
Catabolite Repression ◽  
Carbon Catabolite Repression ◽  
Carbon Sources ◽  
Transcriptional Level ◽  
Pseudomonas Oleovorans ◽  
Hydroxylase Activity ◽  
Alkane Hydroxylase ◽  
Alkane Degradation ◽  
E Coli

ABSTRACT The alk genes are located on the OCT plasmid ofPseudomonas oleovorans and encode an inducible pathway for the utilization of n-alkanes as carbon and energy sources. We have investigated the influence of alternative carbon sources on the induction of this pathway in P. oleovorans andEscherichia coli alk + recombinants. In doing so, we confirmed earlier reports that induction of alkane hydroxylase activity in pseudomonads is subject to carbon catabolite repression. Specifically, synthesis of the monooxygenase component AlkB is repressed at the transcriptional level. The alk genes have been cloned into plasmid pGEc47, which has a copy number of about 5 to 10 per cell in both E. coli and pseudomonads.Pseudomonas putida GPo12 is a P. oleovoransderivative cured of the OCT plasmid. Upon introduction of pGEc47 in this strain, carbon catabolite repression of alkane hydroxylase activity was reduced significantly. In cultures of recombinant E. coli HB101 and W3110 carrying pGEc47, induction of AlkB and transcription of the alkB gene were no longer subject to carbon catabolite repression. This suggests that carbon catabolite repression of alkane degradation is regulated differently inPseudomonas and in E. coli strains. These results also indicate that P alkBFGHJKL , the P alk promoter, might be useful in attaining high expression levels of heterologous genes in E. coligrown on inexpensive carbon sources which normally trigger carbon catabolite repression of native expression systems in this host.

Synthesis and Breakdown of the Universal Precursors to Biological Metabolism Promoted by Ferrous Iron

2018 ◽  
Author(s):  
Kamila B. Muchowska ◽  
Sreejith Jayasree VARMA ◽  
Joseph Moran
Keyword(s):  
Amino Acids ◽  
Chemical Reaction ◽  
Metabolic Pathways ◽  
Ferrous Iron ◽  
Reaction Network ◽  
Tca Cycle ◽  
Chemical Reaction Network ◽  
Metabolic Precursors ◽  
Tca Cycle Intermediates

How core biological metabolism initiated and why it uses the intermediates, reactions and pathways that it does remains unclear. Life builds its molecules from CO<sub>2 </sub>and breaks them down to CO<sub>2 </sub>again through the intermediacy of just five metabolites that act as the hubs of biochemistry. Here, we describe a purely chemical reaction network promoted by Fe<sup>2+ </sup>in which aqueous pyruvate and glyoxylate, two products of abiotic CO<sub>2 </sub>reduction, build up nine of the eleven TCA cycle intermediates, including all five universal metabolic precursors. The intermediates simultaneously break down to CO<sub>2 </sub>in a life-like regime resembling biological anabolism and catabolism. Introduction of hydroxylamine and Fe<sup>0 </sup>produces four biological amino acids. The network significantly overlaps the TCA/rTCA and glyoxylate cycles and may represent a prebiotic precursor to these core metabolic pathways.

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