Plant Mitochondrial Carriers: Molecular Gatekeepers That Help to Regulate Plant Central Carbon Metabolism

The evolution of membrane-bound organelles among eukaryotes led to a highly compartmentalized metabolism. As a compartment of the central carbon metabolism, mitochondria must be connected to the cytosol by molecular gates that facilitate a myriad of cellular processes. Members of the mitochondrial carrier family function to mediate the transport of metabolites across the impermeable inner mitochondrial membrane and, thus, are potentially crucial for metabolic control and regulation. Here, we focus on members of this family that might impact intracellular central plant carbon metabolism. We summarize and review what is currently known about these transporters from in vitro transport assays and in planta physiological functions, whenever available. From the biochemical and molecular data, we hypothesize how these relevant transporters might play a role in the shuttling of organic acids in the various flux modes of the TCA cycle. Furthermore, we also review relevant mitochondrial carriers that may be vital in mitochondrial oxidative phosphorylation. Lastly, we survey novel experimental approaches that could possibly extend and/or complement the widely accepted proteoliposome reconstitution approach.

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Comparative metabolomics of Phialemonium curvatum as an omnipotent fungus cultivated on crude palm oil versus glucose

Microbial Cell Factories ◽

10.1186/s12934-020-01434-w ◽

2020 ◽

Vol 19 (1) ◽

Author(s):

Arief Izzairy Zamani ◽

Susann Barig ◽

Sarah Ibrahim ◽

Hirzun Mohd. Yusof ◽

Julia Ibrahim ◽

...

Keyword(s):

Carbon Source ◽

Organic Acids ◽

Vegetable Oil ◽

Carbon Metabolism ◽

Sole Carbon Source ◽

Carbon Sources ◽

Tca Cycle ◽

Central Carbon Metabolism ◽

Targeted Metabolomics ◽

Central Carbon

Abstract Background Sugars and triglycerides are common carbon sources for microorganisms. Nonetheless, a systematic comparative interpretation of metabolic changes upon vegetable oil or glucose as sole carbon source is still lacking. Selected fungi that can grow in acidic mineral salt media (MSM) with vegetable oil had been identified recently. Hence, this study aimed to investigate the overall metabolite changes of an omnipotent fungus and to reveal changes at central carbon metabolism corresponding to both carbon sources. Results Targeted and non-targeted metabolomics for both polar and semi-polar metabolites of Phialemonium curvatum AWO2 (DSM 23903) cultivated in MSM with palm oil (MSM-P) or glucose (MSM-G) as carbon sources were obtained. Targeted metabolomics on central carbon metabolism of tricarboxylic acid (TCA) cycle and glyoxylate cycle were analysed using LC–MS/MS-TripleQ and GC–MS, while untargeted metabolite profiling was performed using LC–MS/MS-QTOF followed by multivariate analysis. Targeted metabolomics analysis showed that glyoxylate pathway and TCA cycle were recruited at central carbon metabolism for triglyceride and glucose catabolism, respectively. Significant differences in organic acids concentration of about 4- to 8-fold were observed for citric acid, succinic acid, malic acid, and oxaloacetic acid. Correlation of organic acids concentration and key enzymes involved in the central carbon metabolism was further determined by enzymatic assays. On the other hand, the untargeted profiling revealed seven metabolites undergoing significant changes between MSM-P and MSM-G cultures. Conclusions Overall, this study has provided insights on the understanding on the effect of triglycerides and sugar as carbon source in fungi global metabolic pathway, which might become important for future optimization of carbon flux engineering in fungi to improve organic acids production when vegetable oil is applied as the sole carbon source.

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Immunosuppressive activity enhances central carbon metabolism and bioenergetics in myeloid-derived suppressor cells in vitro models

BMC Cell Biology ◽

10.1186/1471-2121-13-18 ◽

2012 ◽

Vol 13 (1) ◽

pp. 18 ◽

Cited By ~ 33

Author(s):

Ines Hammami ◽

Jingkui Chen ◽

Frederic Murschel ◽

Vincenzo Bronte ◽

Gregory De Crescenzo ◽

...

Keyword(s):

Carbon Metabolism ◽

Suppressor Cells ◽

Central Carbon Metabolism ◽

In Vitro Models ◽

Myeloid Derived Suppressor Cells ◽

Immunosuppressive Activity ◽

Central Carbon

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Dissection of Central Carbon Metabolism of Hemoglobin-ExpressingEscherichia coli by 13C Nuclear Magnetic Resonance Flux Distribution Analysis in Microaerobic Bioprocesses

Applied and Environmental Microbiology ◽

10.1128/aem.67.2.680-687.2001 ◽

2001 ◽

Vol 67 (2) ◽

pp. 680-687 ◽

Cited By ~ 27

Author(s):

Alexander D. Frey ◽

Jocelyne Fiaux ◽

Thomas Szyperski ◽

Kurt Wüthrich ◽

James E. Bailey ◽

...

Keyword(s):

Nuclear Magnetic Resonance ◽

Magnetic Resonance ◽

Carbon Metabolism ◽

Tca Cycle ◽

Central Carbon Metabolism ◽

Distribution Analysis ◽

E Coli ◽

13C Nuclear Magnetic Resonance ◽

Central Carbon ◽

Nuclear Magnetic

ABSTRACT Escherichia coli MG1655 cells expressingVitreoscilla hemoglobin (VHb), Alcaligenes eutrophus flavohemoprotein (FHP), the N-terminal hemoglobin domain of FHP (FHPg), and a fusion protein which comprises VHb and theA. eutrophus C-terminal reductase domain (VHb-Red) were grown in a microaerobic bioreactor to study the effects of low oxygen concentrations on the central carbon metabolism, using fractional13C-labeling of the proteinogenic amino acids and two-dimensional [13C, 1H]-correlation nuclear magnetic resonance (NMR) spectroscopy. The NMR data revealed differences in the intracellular carbon fluxes between E. coli cells expressing either VHb or VHb-Red and cells expressingA. eutrophus FHP or the truncated heme domain (FHPg).E. coli MG1655 cells expressing either VHb or VHb-Red were found to function with a branched tricarboxylic acid (TCA) cycle. Furthermore, cellular demands for ATP and reduction equivalents in VHb- and VHb-Red-expressing cells were met by an increased flux through glycolysis. In contrast, in E. coli cells expressingA. eutrophus hemeproteins, the TCA cycle is running cyclically, indicating a shift towards a more aerobic regulation. Consistently, E. coli cells displaying FHP and FHPg activity showed lower production of the typical anaerobic by-products formate, acetate, and d-lactate. The implications of these observations for biotechnological applications are discussed.

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Metabolic Flux Ratio Analysis of Genetic and Environmental Modulations of Escherichia coli Central Carbon Metabolism

Journal of Bacteriology ◽

10.1128/jb.181.21.6679-6688.1999 ◽

1999 ◽

Vol 181 (21) ◽

pp. 6679-6688 ◽

Cited By ~ 263

Author(s):

Uwe Sauer ◽

Daniel R. Lasko ◽

Jocelyne Fiaux ◽

Michel Hochuli ◽

Ralf Glaser ◽

...

Keyword(s):

Escherichia Coli ◽

Carbon Metabolism ◽

Metabolic Flux ◽

Flux Ratio ◽

Tca Cycle ◽

Central Carbon Metabolism ◽

E Coli ◽

Chemostat Cultures ◽

Central Carbon ◽

Pep Carboxykinase

ABSTRACT The response of Escherichia coli central carbon metabolism to genetic and environmental manipulation has been studied by use of a recently developed methodology for metabolic flux ratio (METAFoR) analysis; this methodology can also directly reveal active metabolic pathways. Generation of fluxome data arrays by use of the METAFoR approach is based on two-dimensional13C-1H correlation nuclear magnetic resonance spectroscopy with fractionally labeled biomass and, in contrast to metabolic flux analysis, does not require measurements of extracellular substrate and metabolite concentrations. METAFoR analyses of E. coli strains that moderately overexpress phosphofructokinase, pyruvate kinase, pyruvate decarboxylase, or alcohol dehydrogenase revealed that only a few flux ratios change in concert with the overexpression of these enzymes. Disruption of both pyruvate kinase isoenzymes resulted in altered flux ratios for reactions connecting the phosphoenolpyruvate (PEP) and pyruvate pools but did not significantly alter central metabolism. These data indicate remarkable robustness and rigidity in central carbon metabolism in the presence of genetic variation. More significant physiological changes and flux ratio differences were seen in response to altered environmental conditions. For example, in ammonia-limited chemostat cultures, compared to glucose-limited chemostat cultures, a reduced fraction of PEP molecules was derived through at least one transketolase reaction, and there was a higher relative contribution of anaplerotic PEP carboxylation than of the tricarboxylic acid (TCA) cycle for oxaloacetate synthesis. These two parameters also showed significant variation between aerobic and anaerobic batch cultures. Finally, two reactions catalyzed by PEP carboxykinase and malic enzyme were identified by METAFoR analysis; these had previously been considered absent in E. colicells grown in glucose-containing media. Backward flux from the TCA cycle to glycolysis, as indicated by significant activity of PEP carboxykinase, was found only in glucose-limited chemostat culture, demonstrating that control of this futile cycle activity is relaxed under severe glucose limitation.

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Characterisation of putative lactate synthetic pathways of Coxiella burnetii

PLoS ONE ◽

10.1371/journal.pone.0255925 ◽

2021 ◽

Vol 16 (8) ◽

pp. e0255925

Author(s):

Janine Hofmann ◽

Mebratu A. Bitew ◽

Miku Kuba ◽

David P. De Souza ◽

Hayley J. Newton ◽

...

Keyword(s):

Enzyme Activity ◽

Coxiella Burnetii ◽

Carbon Metabolism ◽

Metabolic Pathways ◽

Malic Enzyme ◽

Lactate Production ◽

Central Carbon Metabolism ◽

Malolactic Enzyme ◽

Central Carbon

The zoonotic pathogen Coxiella burnetii, the causative agent of the human disease Q fever, is an ever-present danger to global public health. Investigating novel metabolic pathways necessary for C. burnetii to replicate within its unusual intracellular niche may identify new therapeutic targets. Recent studies employing stable isotope labelling established the ability of C. burnetii to synthesize lactate, despite the absence of an annotated synthetic pathway on its genome. A noncanonical lactate synthesis pathway could provide a novel anti-Coxiella target if it is essential for C. burnetii pathogenesis. In this study, two C. burnetii proteins, CBU1241 and CBU0823, were chosen for analysis based on their similarities to known lactate synthesizing enzymes. Recombinant GST-CBU1241, a putative malate dehydrogenase (MDH), did not produce measurable lactate in in vitro lactate dehydrogenase (LDH) activity assays and was confirmed to function as an MDH. Recombinant 6xHis-CBU0823, a putative NAD+-dependent malic enzyme, was shown to have both malic enzyme activity and MDH activity, however, did not produce measurable lactate in either LDH or malolactic enzyme activity assays in vitro. To examine potential lactate production by CBU0823 more directly, [13C]glucose labelling experiments compared label enrichment within metabolic pathways of a cbu0823 transposon mutant and the parent strain. No difference in lactate production was observed, but the loss of CBU0823 significantly reduced 13C-incorporation into glycolytic and TCA cycle intermediates. This disruption to central carbon metabolism did not have any apparent impact on intracellular replication within THP-1 cells. This research provides new information about the mechanism of lactate biosynthesis within C. burnetii, demonstrating that CBU1241 is not multifunctional, at least in vitro, and that CBU0823 also does not synthesize lactate. Although critical for normal central carbon metabolism of C. burnetii, loss of CBU0823 did not significantly impair replication of the bacterium inside cells.

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Implications of central carbon metabolism in SARS-CoV-2 replication and disease severity

10.1101/2021.02.24.432759 ◽

2021 ◽

Cited By ~ 1

Author(s):

Shuba Krishnan ◽

Hampus Nordqvist ◽

Anoop T. Ambikan ◽

Soham Gupta ◽

Maike Sperk ◽

...

Keyword(s):

Amino Acid ◽

Disease Severity ◽

Carbon Metabolism ◽

Metabolic Pathways ◽

Central Carbon Metabolism ◽

Lung Epithelial Cells ◽

Metabolic Adaptation ◽

Altered Expression ◽

Central Carbon

AbstractViruses hijack host metabolic pathways for their replicative advantage. Several observational trans-omics analyses associated carbon and amino acid metabolism in coronavirus disease 2019 (COVID-19) severity in patients but lacked mechanistic insights. In this study, using patient- derived multi-omics data and in vitro infection assays, we aimed to understand i) role of key metabolic pathways in severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) reproduction and ii) its association with disease severity. Our data suggests that monocytes are key to the altered immune response during COVID-19. COVID-19 infection was associated with increased plasma glutamate levels, while glucose and mannose levels were determinants of the disease severity. Monocytes showed altered expression pattern of carbohydrate and amino acid transporters, GLUT1 and xCT respectively in severe COVID-19. Furthermore, lung epithelial cells (Calu-3) showed a strong acute metabolic adaptation following infection in vitro by modulating central carbon metabolism. We found that glycolysis and glutaminolysis are essential for virus replication and blocking these metabolic pathways caused significant reduction in virus production. Taken together, our study highlights that the virus utilizes and re-wires pathways governing central carbon metabolism leading to metabolic toxicity. Thus, the host metabolic perturbation could be an attractive strategy to limit the viral replication and disease severity.

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