Effect of the global redox sensing/regulation networks on Escherichia coli and metabolic flux distribution based on C-13 labeling experiments

Zhu, Jiangfeng; Shalel-Levanon, Sagit; Bennett, George N.; San, Ka‐Yiu

doi:10.1016/j.ymben.2006.07.002

Cited by 37 publications

(35 citation statements)

References 51 publications

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“…The regulatory pattern of some fluxes (also observed at both the transcriptional and enzymatic activity levels) significantly departs from that reported for arcA mutants under conditions with restricted O 2 supply (42,73), providing evidence that elimination of the entire arcB coding sequence (or sequences encoding different ArcB domains) has a different effect on cell physiology than does the absence of the cognate response regulator. Cases in point include the pattern of regulation at the pyruvate metabolic node and the split of acetyl-CoA between ethanol and acetate fluxes (see below).…”

Section: Discussionmentioning

confidence: 72%

Manipulation of the Anoxic Metabolism in Escherichia coli by ArcB Deletion Variants in the ArcBA Two-Component System

Bidart

Ruiz

Almeida

et al. 2012

Appl Environ Microbiol

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Bioprocesses conducted under conditions with restricted O 2 supply are increasingly exploited for the synthesis of reduced biochemicals using different biocatalysts. The model facultative anaerobe Escherichia coli has elaborate sensing and signal transduction mechanisms for redox control in response to the availability of O 2 and other electron acceptors. The ArcBA two-component system consists of ArcB, a membrane-associated sensor kinase, and ArcA, the cognate response regulator. The tripartite hybrid kinase ArcB possesses a transmembrane, a PAS, a primary transmitter (H1), a receiver (D1), and a phosphotransfer (H2) domain. Metabolic fluxes were compared under anoxic conditions in a wild-type E. coli strain, its ⌬arcB derivative, and two partial arcB deletion mutants in which ArcB lacked either the H1 domain or the PAS-H1-D1 domains. These analyses revealed that elimination of different segments in ArcB determines a distinctive distribution of D-glucose catabolic fluxes, different from that observed in the ⌬arcB background. Metabolite profiles, enzyme activity levels, and gene expression patterns were also investigated in these strains. Relevant alterations were observed at the P-enol-pyruvate/pyruvate and acetyl coenzyme A metabolic nodes, and the formation of reduced fermentation metabolites, such as succinate, D-lactate, and ethanol, was favored in the mutant strains to different extents compared to the wild-type strain. These phenotypic traits were associated with altered levels of the enzymatic activities operating at these nodes, as well as with elevated NADH/NAD ؉ ratios. Thus, targeted modification of global regulators to obtain different metabolic flux distributions under anoxic conditions is emerging as an attractive tool for metabolic engineering purposes.A noxic fermentation of different carbon sources by Escherichia coli is increasingly gaining momentum in biotechnological setups designed to obtain reduced biochemicals. Relevant examples in this sense include (but are certainly not limited to) the production of ethanol (31, 58, 69), succinate (64), D-lactate (45), and polyhydroxyalkanoates (24, 40), often by using redox and/or regulatory E. coli mutants as the biocatalyst. These metabolic engineering approaches underscore the need for a complete understanding of the cell physiology and metabolic network operativity under anoxic growth conditions. In fact, the relative lack of knowledge on the cellular wiring of these regulatory networks under conditions relevant to both laboratory and industrial applications represents a hurdle that has to be overcome for the efficient design of industrial processes. Metabolic fluxes through the central carbon pathways constitute the backbone of cell metabolism and represent the in vivo reaction rates of cognate enzymatic steps (62). The observed fluxome is the phenotypic consequence of both gene transcription and translation, as well as of the enzymatic activity and the regulation exerted at the metabolite level (48). Fluxome analysis is thus a useful approach to...

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Section: Discussionmentioning

confidence: 72%

Manipulation of the Anoxic Metabolism in Escherichia coli by ArcB Deletion Variants in the ArcBA Two-Component System

Bidart

Ruiz

Almeida

et al. 2012

Appl Environ Microbiol

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“…As stated above, when reducing equivalents are in excess, Pyr can be converted into lactate through NADHdependent LdhA (as was the case for CT1062, the arcA mutant). Zhu et al (76) previously showed that higher fluxes toward lactate synthesis in an arcA background can be attributed to the lower conversion of Pyr into acetyl-CoA and, at the same time, a reducing redox environment. Pyr utilization is important for maintenance of carbon fluxes because when its concentration reaches a threshold value, PEP synthesis is inhibited and growth on phosphotransferase system sugars is impaired (11,37), among other metabolic effects.…”

Section: Discussionmentioning

confidence: 99%

“…Only recently, the impact of metabolic regulation of ArcAB and Fnr (for fumarate and nitrate respiration) on fermentative metabolite production patterns was studied under different oxygen levels (59,76). To understand the global metabolic regulatory functions exerted by CreBC and ArcAB, it would be helpful to look at the overall intracellular flux distribution under microaerobic conditions.…”

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confidence: 99%

Metabolic Flux Analysis of Escherichia coli creB and arcA Mutants Reveals Shared Control of Carbon Catabolism under Microaerobic Growth Conditions

et al. 2009

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Escherichia coli has several elaborate sensing mechanisms for response to availability of oxygen and other electron acceptors, as well as the carbon source in the surrounding environment. Among them, the CreBC and ArcAB two-component signal transduction systems are responsible for regulation of carbon source utilization and redox control in response to oxygen availability, respectively. We assessed the role of CreBC and ArcAB in regulating the central carbon metabolism of E. coli under microaerobic conditions by means of 13 C-labeling experiments in chemostat cultures of a wild-type strain, ⌬creB and ⌬arcA single mutants, and a ⌬creB ⌬arcA double mutant. Continuous cultures were conducted at D ‫؍‬ 0.1 h ؊1 under carbon-limited conditions with restricted oxygen supply. Although all experimental strains metabolized glucose mainly through the EmbdenMeyerhof-Parnas pathway, mutant strains had significantly lower fluxes in both the oxidative and the nonoxidative pentose phosphate pathways. Significant differences were also found at the pyruvate branching point. Both pyruvate-formate lyase and the pyruvate dehydrogenase complex contributed to acetyl-coenzyme A synthesis from pyruvate, and their activity seemed to be modulated by both ArcAB and CreBC. Strains carrying the creB deletion showed a higher biomass yield on glucose compared to the wild-type strain and its ⌬arcA derivative, which also correlated with higher fluxes from building blocks to biomass. Glyoxylate shunt and lactate dehydrogenase were active mainly in the ⌬arcA strain. Finally, it was observed that the tricarboxylic acid cycle reactions operated in a rather cyclic fashion under our experimental conditions, with reduced activity in the mutant strains.Transcriptional regulation comprises a complex network of global and specific regulators. Global regulators normally have pleiotropic effects on metabolism, since they control the transcription of several operons that belong to different functional groups. In Escherichia coli, seven global regulators (ArcA, Crp, Fis, Fnr, Ihf, Lrp, and NarL) directly modulate expression of about one-half of all genes (41). CreBC and ArcAB are two global sensing and regulation systems in E. coli which, along with some other regulatory systems, modulate the expression of genes involved in central metabolism and respiratory pathways allowing rapid adaptive responses to carbon source and oxygen availability in the culture environment.The cre (for carbon source responsive) locus comprises creABCD and was formerly known as the phoM locus (4). Whereas creA is a hypothetical open reading frame and creD, also known as cet, encodes an inner-membrane protein of unknown function, creB and creC encode a two-component system, i.e., a cytoplasmic regulator and a sensor kinase, respectively. After the discovery of CreBC, genes modulated by this system proved elusive. Based on sequence analysis and using bioinformatic tools Avison et al. (5) were able to define a cre tag sequence, to which CreB binds in vitro (8), in order to describe the...

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“…The medium used for shakenflask experiments was M9 minimal medium containing (per liter of deionized H 2 O) 6.0 g of Na 2 HPO 4 , 3.0 g of KH 2 PO 4 , 0.5 g of NaCl, 1.0 g of NH 4 Cl, 0.4 g of MgSO 4 , 0.01 g of CaCl 2 , and 0.06 g of ammonium iron(III) citrate. MgSO 4 , CaCl 2 , and ammonium iron(III) citrate were added to the medium after autoclaving and cooling.…”

Section: Methodsmentioning

confidence: 99%

“…In Escherichia coli, seven global regulators (ArcA, Crp, Fis, Fnr, Ihf, Lrp, and NarL) directly modulate the expression of about one-half of all genes (1). This facultative aerobe is able to adapt its metabolism to different oxygen availability conditions through the concerted actions of a network of regulators, including the global regulators ArcAB and Fnr (2)(3)(4). These regulators affect many metabolic pathways, allowing the cells to reach an adequate redox balance under any given conditions.…”

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confidence: 99%

The CreC Regulator of Escherichia coli, a New Target for Metabolic Manipulations

Godoy

Nikel

Gomez

et al. 2016

Appl Environ Microbiol

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The CreBC (carbon source-responsive) two-component regulation system of Escherichia coli affects a number of functions, including intermediary carbon catabolism. The impacts of different creC mutations (a ⌬creC mutant and a mutant carrying the constitutive creC510 allele) on bacterial physiology were analyzed in glucose cultures under three oxygen availability conditions. Differences in the amounts of extracellular metabolites produced were observed in the null mutant compared to the wild-type strain and the mutant carrying creC510 and shown to be affected by oxygen availability. The ⌬creC strain secreted more formate, succinate, and acetate but less lactate under low aeration. These metabolic changes were associated with differences in AckA and LdhA activities, both of which were affected by CreC. Measurement of the NAD(P)H/NAD(P) ؉ ratios showed that the creC510 strain had a more reduced intracellular redox state, while the opposite was observed for the ⌬creC mutant, particularly under intermediate oxygen availability conditions, indicating that CreC affects redox balance. The null mutant formed more succinate than the wild-type strain under both low aeration and no aeration. Overexpression of the genes encoding phosphoenolpyruvate carboxylase from E. coli and a NADH-forming formate dehydrogenase from Candida boidinii in the ⌬creC mutant further increased the yield of succinate on glucose. Interestingly, the elimination of ackA and adhE did not significantly improve the production of succinate. The diverse metabolic effects of this regulator on the central biochemical network of E. coli make it a good candidate for metabolic-engineering manipulations to enhance the formation of bioproducts, such as succinate.T he survival of an organism depends, at least in part, on its ability to sense and respond to changes in the environment. In bacteria, global regulators control the transcription of genes in response to specific external stimuli and metabolic signals, finely tuning different aspects of their physiology to overcome environmental challenges. In Escherichia coli, seven global regulators (ArcA, Crp, Fis, Fnr, Ihf, Lrp, and NarL) directly modulate the expression of about one-half of all genes (1). This facultative aerobe is able to adapt its metabolism to different oxygen availability conditions through the concerted actions of a network of regulators, including the global regulators ArcAB and Fnr (2-4). These regulators affect many metabolic pathways, allowing the cells to reach an adequate redox balance under any given conditions. There is a very close association between carbon and electron flows, and even small differences in oxygen availability have been observed to elicit profound effects on the distribution of carbon fluxes (5).CreBC (for carbon source responsive) is a global sensing and regulation system affecting genes involved in a variety of functions, including enzymes of intermediary catabolism (6). Previous studies have shown that the creABCD operon is activated (i) during growth in minimal mediu...

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Effect of the global redox sensing/regulation networks on Escherichia coli and metabolic flux distribution based on C-13 labeling experiments

Cited by 37 publications

References 51 publications

Manipulation of the Anoxic Metabolism in Escherichia coli by ArcB Deletion Variants in the ArcBA Two-Component System

Manipulation of the Anoxic Metabolism in Escherichia coli by ArcB Deletion Variants in the ArcBA Two-Component System

Metabolic Flux Analysis of Escherichia coli creB and arcA Mutants Reveals Shared Control of Carbon Catabolism under Microaerobic Growth Conditions

The CreC Regulator of Escherichia coli, a New Target for Metabolic Manipulations

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