13C-flux Analysis Reveals NADPH-balancing Transhydrogenation Cycles in Stationary Phase of Nitrogen-starving Bacillus subtilis

Rühl, Martin; Coq, Dominique Le; Aymerich, Stéphane; Sauer, Uwe

doi:10.1074/jbc.m112.366492

Cited by 54 publications

(58 citation statements)

References 64 publications

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“…Our data and that obtained from nitrogen-starved B. subtilis (37), demonstrate that bacteria can have low transcript levels for metabolic genes and yet remain metabolically active. In the case of R. palustris, metabolic activity can persist in the absence of growth for months (3).…”

Section: Discussionmentioning

confidence: 78%

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Non-growing Rhodopseudomonas palustris Increases the Hydrogen Gas Yield from Acetate by Shifting from the Glyoxylate Shunt to the Tricarboxylic Acid Cycle

McKinlay

Oda

Rühl

et al. 2014

Journal of Biological Chemistry

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Background:The metabolism of non-growing microbes is poorly understood. Results: In a nitrogen-starved and non-growing photoheterotrophic bacterium, metabolic flow was diverted to mobilize electrons for H 2 production. Conclusion: During starvation bacteria decouple their metabolism from biosynthesis. Significance: An understanding of metabolic activities of non-growing cells can be used to engineer improved biocatalysts.

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Section: Discussionmentioning

confidence: 78%

“…Oxidation of carbon sources to CO 2 may be a widespread bacterial response to nitrogen deprivation, even in non-photosynthetic, respiring bacteria. 13 C-metabolic flux analysis of nitrogen-starved B. subtilis fed with glucose also showed elevated fluxes through CO 2 -producing oxidative pathways (37).…”

Section: Discussionmentioning

confidence: 99%

Non-growing Rhodopseudomonas palustris Increases the Hydrogen Gas Yield from Acetate by Shifting from the Glyoxylate Shunt to the Tricarboxylic Acid Cycle

McKinlay

Oda

Rühl

et al. 2014

Journal of Biological Chemistry

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“…The cofactor is typically formed through the pentose phosphate pathway (glucose-6-phosphate dehydrogenase) and the TCA cycle (isocitrate dehydrogenase) (65)(66)(67). Since the pentose phosphate pathway is poorly active in A. pasteurianus NCC 316, we wondered how A. pasteurianus covers its biosynthetic demand for NADPH ϩ H ϩ .…”

Section: Discussionmentioning

confidence: 99%

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

Adler

Frey

Berger

et al. 2014

Appl Environ Microbiol

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c Acetic acid bacteria (AAB) play an important role during cocoa fermentation, as their main product, acetate, is a major driver for the development of the desired cocoa flavors. Here, we investigated the specialized metabolism of these bacteria under cocoa pulp fermentation-simulating conditions. A carefully designed combination of parallel 13 C isotope labeling experiments allowed the elucidation of intracellular fluxes in the complex environment of cocoa pulp, when lactate and ethanol were included as primary substrates among undefined ingredients. We demonstrate that AAB exhibit a functionally separated metabolism during coconsumption of two-carbon and three-carbon substrates. Acetate is almost exclusively derived from ethanol, while lactate serves for the formation of acetoin and biomass building blocks. Although this is suboptimal for cellular energetics, this allows maximized growth and conversion rates. The functional separation results from a lack of phosphoenolpyruvate carboxykinase and malic enzymes, typically present in bacteria to interconnect metabolism. In fact, gluconeogenesis is driven by pyruvate phosphate dikinase. Consequently, a balanced ratio of lactate and ethanol is important for the optimum performance of AAB. As lactate and ethanol are individually supplied by lactic acid bacteria and yeasts during the initial phase of cocoa fermentation, respectively, this underlines the importance of a well-balanced microbial consortium for a successful fermentation process. Indeed, AAB performed the best and produced the largest amounts of acetate in mixed culture experiments when lactic acid bacteria and yeasts were both present.A cetic acid bacteria (AAB) play an important role in cocoa fermentation (1). During fermentation of the pulp that surrounds the cocoa beans, they form acetate. Acetate then diffuses into the beans (2-4), where it initiates a cascade of chemical and biochemical reactions leading to precursor molecules for cocoa flavor (2, 5, 6). Potential substrates for AAB are lactate and ethanol, which are individually produced by lactic acid bacteria (LAB; mainly Lactobacillus fermentum) and yeasts (diverse yeasts such as Saccharomyces cerevisiae, Hanseniaspora opuntiae, and Candida krusei), respectively, during the fermentation process (6-12). Hereby, the degradation of lactate by AAB is desired, since the remaining lactate may provide an off flavor in the final cocoa product (11,13,14). In recent years, AAB have been extensively analyzed for their contribution to cocoa fermentation. Obviously, the most prevalent AAB species is Acetobacter pasteurianus (13,(15)(16)(17). In addition, Acetobacter ghanensis and Acetobacter senegalensis are found during spontaneous cocoa bean fermentation (9,13,17,18). Further studies provided first insights into the basic microbiological properties of such strains and macroscopic dynamics during cocoa pulp fermentation (12,15,(18)(19)(20). At this point, it appears to be relevant to resolve the metabolic contribution of AAB in greater detail. The basic know...

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“…For the many bacteria that use carbon catabolism for energy generation, carbon starvation implies a lack of energy, and consequently, the cell has a limited array of stationary-phase responses. On the other hand, if noncarbon nutrients are limiting for growth, a full catabolic program is possible (10), and the harvested energy could be invested in a variety of ways. Still, metabolic activity without growth could have potential downsides, such as the accumulation of toxic intermediates or decreased resistance to antibiotics, and it is not obvious how microbes resolve this trade-off (11).…”

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confidence: 99%

Environmental Dependence of Stationary-Phase Metabolism in Bacillus subtilis and Escherichia coli

Chubukov

Sauer

2014

Appl Environ Microbiol

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102

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When microbes lack the nutrients necessary for growth, they enter stationary phase. In cases when energy sources are still present in the environment, they must decide whether to continue to use their metabolic program to harvest the available energy. Here we characterized the metabolic response to a variety of types of nutrient starvation in Escherichia coli and Bacillus subtilis. We found that E. coli exhibits a range of phenotypes, with the lowest metabolic rates under nitrogen starvation and highest rates under magnesium starvation. In contrast, the phenotype of B. subtilis was dominated by its decision to form metabolically inactive endospores. While its metabolic rates under most conditions were thus lower than those of E. coli, when sporulation was suppressed by a genetic perturbation or an unnatural starvation condition, the situation was reversed. To further probe stationary-phase metabolism, we used quantitative metabolomics to investigate possible small-molecule signals that may regulate the metabolic rate of E. coli and initiate sporulation in B. subtilis. We hypothesize a role for phosphoenolpyruvate (PEP) in regulating E. coli glucose uptake and for the redox cofactors NAD(H) and NADP(H) in initiation of sporulation. Our work is directly relevant to synthetic biology and metabolic engineering, where active metabolism during stationary phase, which uncouples production from growth, remains an elusive goal.

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13C-flux Analysis Reveals NADPH-balancing Transhydrogenation Cycles in Stationary Phase of Nitrogen-starving Bacillus subtilis

Cited by 54 publications

References 64 publications

Non-growing Rhodopseudomonas palustris Increases the Hydrogen Gas Yield from Acetate by Shifting from the Glyoxylate Shunt to the Tricarboxylic Acid Cycle

Non-growing Rhodopseudomonas palustris Increases the Hydrogen Gas Yield from Acetate by Shifting from the Glyoxylate Shunt to the Tricarboxylic Acid Cycle

The Key to Acetate: Metabolic Fluxes of Acetic Acid Bacteria under Cocoa Pulp Fermentation-Simulating Conditions

Environmental Dependence of Stationary-Phase Metabolism in Bacillus subtilis and Escherichia coli

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