scholarly journals Analysis of tricarboxylic acid-cycle metabolism of hepatoma cells by comparison of 14CO2 ratios

1987 ◽  
Vol 246 (3) ◽  
pp. 633-639 ◽  
Author(s):  
J K Kelleher ◽  
B M Bryan ◽  
R T Mallet ◽  
A L Holleran ◽  
A N Murphy ◽  
...  
Keyword(s):  
Steady State ◽  
Tricarboxylic Acid Cycle ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Hepatoma Cells ◽  
Acid Cycle ◽  
The Tca Cycle ◽  
14Co2 Production ◽  
Metabolic Properties ◽  
Tca Cycle Intermediates

The CO2-ratios method is applied to the analysis of abnormalities of TCA (tricarboxylic acid)-cycle metabolism in AS-30D rat ascites-hepatoma cells. This method utilizes steady-state 14CO2-production rates from pairs of tracers of the same compound to evaluate TCA-cycle flux patterns. Equations are presented that quantitatively convert CO2 ratios into estimates of probability of flux through TCA-cycle-related pathways. Results of this study indicated that the ratio of 14CO2 produced from [1,4-14C]succinate to 14CO2 produced from [2,3-14C]succinate was increased by the addition of glutamine (5 mM) to the medium. An increase in the succinate CO2 ratio is quantitatively related to an increased flux of unlabelled carbon into the TCA-cycle-intermediate pools. Analysis of 14C distribution in [14C]citrate derived from [2,3-14C]succinate indicated that flux from the TCA cycle to the acetyl-CoA-derived carbons of citrate was insignificant. Thus the increased succinate CO2 ratio observed in the presence of glutamine could only result from an increased flux of carbon into the span of the TCA cycle from citrate to oxaloacetate. This result is consistent with increased flux of glutamine to alpha-oxoglutarate in the incubation medium containing exogenous glutamine. Comparison of the pyruvate CO2 ratio, steady-state 14CO2 production from [2-14C]pyruvate versus [3-14C]pyruvate, with the succinate 14CO2 ratio detected flux of pyruvate to C4 TCA-cycle intermediates in the medium containing glutamine. This result was consistent with the observation that [14C]aspartate derived from [2-14C]pyruvate was labelled in C-2 and C-3. 14C analysis also produced evidence for flux of TCA-cycle carbon to alanine. This study demonstrates that the CO2-ratios method is applicable in the analysis of the metabolic properties of AS-30D cells. This methodology has verified that the atypical TCA-cycle metabolism previously described for AS-30D-cell mitochondria occurs in intact AS-30D rat hepatoma cells.

scholarly journals Staphylococcus epidermidis Polysaccharide Intercellular Adhesin Production Significantly Increases during Tricarboxylic Acid Cycle Stress

2005 ◽  
Vol 187 (9) ◽  
pp. 2967-2973 ◽  
Author(s):  
Cuong Vuong ◽  
Joshua B. Kidder ◽  
Erik R. Jacobson ◽  
Michael Otto ◽  
Richard A. Proctor ◽  
...  
Keyword(s):  
Staphylococcus Epidermidis ◽  
Tricarboxylic Acid Cycle ◽  
Tca Cycle ◽  
Redox Status ◽  
Tricarboxylic Acid ◽  
Polysaccharide Intercellular Adhesin ◽  
Low Oxygen ◽  
Acid Cycle ◽  
The Tca Cycle ◽  
High Concentrations

ABSTRACT Staphylococcal polysaccharide intercellular adhesin (PIA) is important for the development of a mature biofilm. PIA production is increased during growth in a nutrient-replete or iron-limited medium and under conditions of low oxygen availability. Additionally, stress-inducing stimuli such as heat, ethanol, and high concentrations of salt increase the production of PIA. These same environmental conditions are known to repress tricarboxylic acid (TCA) cycle activity, leading us to hypothesize that altering TCA cycle activity would affect PIA production. Culturing Staphylococcus epidermidis with a low concentration of the TCA cycle inhibitor fluorocitrate dramatically increased PIA production without impairing glucose catabolism, the growth rate, or the growth yields. These data lead us to speculate that one mechanism by which staphylococci perceive external environmental change is through alterations in TCA cycle activity leading to changes in the intracellular levels of biosynthetic intermediates, ATP, or the redox status of the cell. These changes in the metabolic status of the bacteria result in the attenuation or augmentation of PIA production.

scholarly journals Tricarboxylic Acid (TCA) Cycle Intermediates: Regulators of Immune Responses

2020 ◽  
Author(s):  
Inseok Choi ◽  
Hyewon Son ◽  
Jea-Hyun Baek
Keyword(s):  
Immune Responses ◽  
Molecular Mechanisms ◽  
Tricarboxylic Acid Cycle ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Cell Stress ◽  
Danger Signals ◽  
Cellular Processes ◽  
The Tca Cycle ◽  
Tca Cycle Intermediates

Tricarboxylic acid cycle (TCA) is a series of chemical reactions in aerobic organisms used to generate energy via the oxidation of acetyl-CoA derived from carbohydrates, fatty acids, and proteins. In the eukaryotic system, the TCA cycle completely occurs in mitochondria, while the intermediates of the TCA cycle are retained in mitochondria due to their polarity and hydrophilicity. Under conditions of cell stress, mitochondria become disrupted and release their contents, which act as danger signals in the cytosol. Of note, the TCA cycle intermediates may also leak from dysfunctioning mitochondria and regulate cellular processes. Increasing evidence shows that the metabolites of the TCA cycle are substantially involved in the regulation of immune responses. In this review, we aimed to provide a comprehensive systematic overview of the molecular mechanisms of each TCA cycle intermediate that may play key roles in regulating cellular immunity in cell stress and discuss their implications for immune activation and suppression.

scholarly journals Tricarboxylic Acid (TCA) Cycle Intermediates: Regulators of Immune Responses

Life ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 69
Author(s):  
Inseok Choi ◽  
Hyewon Son ◽  
Jea-Hyun Baek
Keyword(s):  
Immune Responses ◽  
Molecular Mechanisms ◽  
Tricarboxylic Acid Cycle ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Cell Stress ◽  
Danger Signals ◽  
Cellular Processes ◽  
The Tca Cycle ◽  
Tca Cycle Intermediates

The tricarboxylic acid cycle (TCA) is a series of chemical reactions used in aerobic organisms to generate energy via the oxidation of acetylcoenzyme A (CoA) derived from carbohydrates, fatty acids and proteins. In the eukaryotic system, the TCA cycle occurs completely in mitochondria, while the intermediates of the TCA cycle are retained inside mitochondria due to their polarity and hydrophilicity. Under cell stress conditions, mitochondria can become disrupted and release their contents, which act as danger signals in the cytosol. Of note, the TCA cycle intermediates may also leak from dysfunctioning mitochondria and regulate cellular processes. Increasing evidence shows that the metabolites of the TCA cycle are substantially involved in the regulation of immune responses. In this review, we aimed to provide a comprehensive systematic overview of the molecular mechanisms of each TCA cycle intermediate that may play key roles in regulating cellular immunity in cell stress and discuss its implication for immune activation and suppression.

scholarly journals New Insights into Heme Utilization Pathways

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. SCI-25-SCI-25
Author(s):  
Emanuela Tolosano
Keyword(s):  
Respiratory Chain ◽  
Tricarboxylic Acid Cycle ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Energetic Metabolism ◽  
Acid Cycle ◽  
Heme Synthesis ◽  
Chain Complexes ◽  
The Tca Cycle

Heme, an iron-containing porphyrin, plays pivotal functions in cell energetic metabolism, serving as a cofactor for most of the respiratory chain complexes and interacting with the translocases responsible for the ADP/ATP exchange between mitochondria and cytosol. Moreover, heme biosynthesis is considered a cataplerotic pathway for the tricarboxylic acid cycle (TCA) cycle, as the process consumes succynil-CoA, an intermediate of the TCA cycle. Finally, heme synthesis is one of the major cellular iron-consuming processes, thus competing with mitochondrial biogenesis of iron-sulfur (Fe-S) clusters, the crucial cofactors of electron transport chain complexes and of some TCA cycle enzymes. The process of heme synthesis consists of eight enzymatic reactions starting in mitochondria with the condensation of glycine and succynil-CoA to form δ-aminolevulinic acid (ALA), catalyzed by amino levulinic acid synthase (ALAS), the rate-limiting enzyme in heme biosynthetic pathway. Two isoforms of ALAS exist, ALAS1, ubiquitously expressed and controlled by heme itself through a negative feedback, and ALAS2, specifically expressed in the erythroid cells and mainly controlled by iron availability. ALA is exported from mitochondria to cytosol and converted to coproporphyrinogenIII that is imported back into the mitochondrial intermembrane space and converted to protoporphyrinogen IX. The latter is oxidized to porphyrin IX. Finally, ferrous iron is inserted into porphyrin IX by ferrochelatase, a Fe-S cluster-containing enzyme. Heme is incorporated into mitochondrial heme-containing proteins including complexes of the respiratory chain or exported to cytosol for incorporation into cytosolic apo-hemoproteins. Cytosolic heme level is maintained by the rate of hemoprotein production, the activity of heme transporters, including both heme importers and exporters, and the rate of heme degradation mediated by heme oxygenases. The concerted action of all these mechanisms regulates heme level that in turn controls its own synthesis by regulating the expression and activity of ALAS1. During differentiation of erythroid progenitors, cells bypass the heme-mediated negative regulation of its production by expressing ALAS2 that is responsible for the high rate of heme synthesis required to sustain hemoglobin production. We showed that the process of heme efflux through the plasma membrane heme exporter Feline Leukemia Virus C Receptor (FLVCR)1a is required to sustain ALAS1-catalyzed heme synthesis. In tumor cells, the potentiation of heme synthesis/export axis contributes to the down-modulation of tricarboxylic acid cycle (TCA) cycle favoring a glycolysis- compared to an oxidative-based metabolism. Our data indicate that the heme synthesis/export axis slow down the TCA cycle through two mechanisms, on one hand, by consuming succynil-CoA, an intermediate of the cycle, and, on the other, by consuming mitochondrial iron thus limiting the production of Fe-S clusters, essential co-factors of complexes of the respiratory chain as well as of key enzymes of the cycle. The importance of heme synthesis/export axis in metabolic rewiring occurring during tumorigenesis is highlighted by the impaired proliferation and survival observed in FLVCR1a-silenced cancer cells. We speculate that the heme synthesis/export axis plays a role in metabolic adaptation also in proliferating cells in physiologic conditions, especially when oxygen concentration is limiting, as suggested by the phenotype of murine models of Flvcr1a deficiency. Finally, in post-mitotic cells the heme synthesis/export axis might contribute to modulate mitochondrial activity. This conclusion is supported by the observation that FLVCR1 gene was found mutated in human pathologies characterized by impaired function of neuronal cell populations strongly dependent on mitochondrial oxidative metabolism. In conclusion, our data highlight the crucial role of heme synthesis/export axis in the control of cell energetic metabolism. Future work is required to elucidate the role of exported heme in the extracellular environment. Disclosures No relevant conflicts of interest to declare.

Analysis of tricarboxylic acid cycle using [14C]citrate specific activity ratios

1985 ◽  
Vol 248 (2) ◽  
pp. E252-E260 ◽  
Author(s):  
J. K. Kelleher
Keyword(s):  
Steady State ◽  
Specific Activity ◽  
Tricarboxylic Acid Cycle ◽  
Specific Radioactivity ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
State Equations ◽  
Futile Cycle ◽  
Net Flux ◽  
Tca Cycle Intermediates

The information content of citrate carbon specific radioactivity ratios in steady-state 14C isotopic studies has been analyzed. Sixteen steady-state equations resulted containing five unknowns, 14 equations in terms of citrate carbon specific radioactivity ratios, and two modified forms of the “CO2 ratio” equation. Although each of the 16 equations is not independent, there is more than one independent equation for each variable. These additional equations may be used to test the assumptions on which the model is based. Each of the five unknowns is defined as the probability of flux around a complete cycle, either the tricarboxylic acid (TCA) cycle or a futile cycle such as pyruvate--oxaloacetate--pyruvate. To solve these equations for the five unknowns, an investigator need only measure the specific radioactivity of various citrate carbons and the 14CO2 production rate. The study did not yield a direct expression for net flux between pyruvate and 4-carbon TCA cycle intermediates. However, these equations do place certain constraints on the net flux through this important pathway.

scholarly journals Metabolic Engineering of the Tricarboxylic Acid Cycle for Improved Lysine Production by Corynebacterium glutamicum

2009 ◽  
Vol 75 (24) ◽  
pp. 7866-7869 ◽  
Author(s):  
Judith Becker ◽  
Corinna Klopprogge ◽  
Hartwig Schröder ◽  
Christoph Wittmann
Keyword(s):  
Metabolic Engineering ◽  
Corynebacterium Glutamicum ◽  
Tricarboxylic Acid Cycle ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Start Codon ◽  
Flux Analysis ◽  
Lysine Production ◽  
Acid Cycle ◽  
The Tca Cycle

ABSTRACT In the present work, lysine production by Corynebacterium glutamicum was improved by metabolic engineering of the tricarboxylic acid (TCA) cycle. The 70% decreased activity of isocitrate dehydrogenase, achieved by start codon exchange, resulted in a >40% improved lysine production. By flux analysis, this could be correlated to a flux shift from the TCA cycle toward anaplerotic carboxylation.

scholarly journals Inactivation of the Tricarboxylic Acid Cycle Aconitase Gene from Streptomyces viridochromogenesTü494 Impairs Morphological and Physiological Differentiation

1999 ◽  
Vol 181 (22) ◽  
pp. 7131-7135 ◽  
Author(s):  
D. Schwartz ◽  
S. Kaspar ◽  
G. Kienzlen ◽  
K. Muschko ◽  
W. Wohlleben
Keyword(s):  
Escherichia Coli ◽  
Tricarboxylic Acid Cycle ◽  
Aerial Mycelium ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Acid Cycle ◽  
The Tca Cycle

ABSTRACT The tricarboxylic acid (TCA) cycle aconitase gene acnAfrom Streptomyces viridochromogenes Tü494 was cloned and analyzed. AcnA catalyzes the isomerization of citrate to isocitrate in the TCA cycle, as indicated by the ability of acnA to complement the aconitase-deficient Escherichia coli mutant JRG3259. An acnA mutant was unable to develop aerial mycelium and to sporulate, resulting in a bald phenotype. Furthermore, the mutant did not produce the antibiotic phosphinothricin tripeptide, demonstrating that AcnA also affects physiological differentiation.

Tricarboxylic acid cycle enzymes of a pseudophyllid cestode Penetrocephalus ganapatii

1990 ◽  
Vol 64 (1) ◽  
pp. 51-53
Author(s):  
S. Dhandayuthapani ◽  
K. Nellaiappan
Keyword(s):  
Isocitrate Dehydrogenase ◽  
Citrate Synthase ◽  
Tricarboxylic Acid Cycle ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Reverse Direction ◽  
Acid Cycle ◽  
Tca Cycle Enzymes ◽  
The Tca Cycle ◽  
Atp Generation

ABSTRACTStudies on the tricarboxylic acid cycle (TCA cycle) enzymes of Penetrocephalus ganapatii reveal that the TCA cycle is only partially operative, as some of the enzymes at the start of the cycle viz. citrate synthase, aconitase and isocitrate dehydrogenase are found to be low in their activities. The high activities of malate dehydrogenase and fumarase, showing affinity towards a reverse direction, indicate that the TCA cycle operates in the reverse direction resulting in the formation of fumarate. The low succinate dehydrogenase/fumarate reductase ratio suggests that ATP generation may occur at site I of the respiratory chain during the reduction of fumarate into succinate.

Biosynthesis of Pyoluteorin: A Mixed Polyketide-Tricarboxylic Acid Cycle Origin Demonstrated by [1,2-13C2]Acetate Incorporation

1986 ◽  
Vol 41 (5-6) ◽  
pp. 532-536 ◽  
Author(s):  
D. A. Cuppels ◽  
C. R. Howell ◽  
R. D. Stipanovic ◽  
A. Stoessl ◽  
J. B. Stothers
Keyword(s):  
Nmr Spectroscopy ◽  
Tricarboxylic Acid Cycle ◽  
Tca Cycle ◽  
Tricarboxylic Acid ◽  
Acetate Incorporation ◽  
Acid Cycle ◽  
The Tca Cycle ◽  
Starter Unit ◽  
C Nmr

The incorporation of [1,2-13C2]acetate into the bacterial phytotoxin pyoluteorin (1) as determined by 13C NMR spectroscopy occurs in a pattern consistent with the biosynthesis of the molecule as a tetraketide in which proline, or an equivalent precursor derived from the TCA cycle, serves as the starter unit.

Sepsis does not impair tricarboxylic acid cycle in the heart

1991 ◽  
Vol 260 (1) ◽  
pp. C50-C57 ◽  
Author(s):  
R. S. Hotchkiss ◽  
S. K. Song ◽  
J. J. Neil ◽  
R. D. Chen ◽  
J. K. Manchester ◽  
...  
Keyword(s):  
Tricarboxylic Acid Cycle ◽  
Cecal Ligation ◽  
Tca Cycle ◽  
High Energy ◽  
Tricarboxylic Acid ◽  
Female Rats ◽  
Resonance Spectroscopy ◽  
High Energy Phosphates ◽  
The Tca Cycle ◽  
Tca Cycle Intermediates

Sepsis has been reported to cause mitochondrial dysfunction and inhibition of key enzymes that regulate the tricarboxylic acid (TCA) cycle. We investigated the effect of sepsis on high-energy phosphates, glycolytic and TCA cycle intermediates, and specific amino acids that are involved in regulating the size of the TCA cycle pool during changes in metabolic state of the heart. Sepsis was induced in 12 female rats by the cecal ligation and perforation technique under halothane anesthesia; seven control rats underwent cecal manipulation without ligation. At 36-42 h postsurgery, the rats were reanesthetized, the chest was opened, and the hearts were freeze-clamped. Perchloric acid extracts of the hearts were analyzed with fluorometric enzymatic methods and 31P nuclear magnetic resonance spectroscopy. There were no significant differences in the levels of the TCA cycle intermediates or high-energy phosphates between the septic and control rats. The major metabolic changes were the 28% decrease in alanine and the 31% decrease in glutamate in the septic hearts compared with control (P less than 0.05 and P less than 0.005, respectively). Phosphocholine, a component of membrane phospholipids, was increased by 91% in the septic hearts (P less than 0.01). We conclude that sepsis does not impair the TCA cycle or induce significant cellular ischemia in the heart. The increase in phosphocholine may represent significant cellular membrane disruption during sepsis.

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