Localisation of gluconeogenesis and tricarboxylic acid (TCA)-cycle enzymes and first functional analysis of the TCA cycle in Toxoplasma gondii

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.

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TCA Cycle Defects and Cancer: When Metabolism Tunes Redox State

International Journal of Cell Biology ◽

10.1155/2012/161837 ◽

2012 ◽

Vol 2012 ◽

pp. 1-9 ◽

Cited By ~ 82

Author(s):

Simone Cardaci ◽

Maria Rosa Ciriolo

Keyword(s):

Redox State ◽

Fumarate Hydratase ◽

Tca Cycle ◽

Tricarboxylic Acid ◽

Cellular Redox ◽

Redox Biology ◽

Recessive Mutations ◽

Tca Cycle Enzymes ◽

The Tca Cycle ◽

Inborn Defects

Inborn defects of the tricarboxylic acid (TCA) cycle enzymes have been known for more than twenty years. Until recently, only recessive mutations were described which, although resulted in severe multisystem syndromes, did not predispose to cancer onset. In the last ten years, a causal role in carcinogenesis has been documented for inherited and acquired alterations in three TCA cycle enzymes, succinate dehydrogenase (SDH), fumarate hydratase (FH), and isocitrate dehydrogenase (IDH), pointing towards metabolic alterations as the underlying hallmark of cancer. This paper summarizes the neoplastic alterations of the TCA cycle enzymes focusing on the generation of pseudohypoxic phenotype and the alteration of epigenetic homeostasis as the main tumor-promoting effects of the TCA cycle affecting defects. Moreover, we debate on the ability of these mutations to affect cellular redox state and to promote carcinogenesis by impacting on redox biology.

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Metabolism of glycolic acid by Azotobacter chroococcum PRL H62

Canadian Journal of Microbiology ◽

10.1139/m73-053 ◽

1973 ◽

Vol 19 (3) ◽

pp. 321-324 ◽

Cited By ~ 5

Author(s):

W. G. W. Kurz ◽

T. A. G. LaRue

Keyword(s):

Nitrogen Fixation ◽

Dicarboxylic Acid ◽

Glycolic Acid ◽

Tca Cycle ◽

Azotobacter Chroococcum ◽

Acid Cycle ◽

Isocitric Dehydrogenase ◽

Tca Cycle Enzymes ◽

The Tca Cycle ◽

Reduced Nitrogen

When Azotobacter chroococcum grows on glycolic acid as sole C source, it cannot utilize N2 and must be provided with reduced nitrogen. Glycolic acid is metabolized via Kornberg's dicarboxylic acid cycle. The TCA cycle enzymes are low in activity, and isocitric dehydrogenase is absent. It is likely that isocitric dehydrogenase is the source of reductant for nitrogen fixation by Azotobacter nitrogenase.

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Gluconeogenesis from labeled carbon: estimating isotope dilution

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.1986.250.3.e296 ◽

1986 ◽

Vol 250 (3) ◽

pp. E296-E305 ◽

Cited By ~ 3

Author(s):

J. K. Kelleher

Keyword(s):

Steady State ◽

Isotope Dilution ◽

Experimental Tests ◽

Tca Cycle ◽

Tricarboxylic Acid ◽

Complex Model ◽

Acetyl Coa ◽

The Tca Cycle ◽

Carbon Compounds ◽

Mathematical Techniques

To estimate the rate of gluconeogenesis from steady-state incorporation of labeled 3-carbon precursors into glucose, isotope dilution must be considered so that the rate of labeling of glucose can be quantitatively converted to the rate of gluconeogenesis. An expression for the value of this isotope dilution can be derived using mathematical techniques and a model of the tricarboxylic acid (TCA) cycle. The present investigation employs a more complex model than that used in previous studies. This model includes the following pathways that may affect the correction for isotope dilution: 1) flux of 3-carbon precursor to the oxaloacetate pool via acetyl-CoA and the TCA cycle; 2) flux of 4- or 5-carbon compounds into the TCA cycle; 3) reversible flux between oxaloacetate (OAA) and pyruvate and between OAA and fumarate; 4) incomplete equilibrium between OAA pools; and 5) isotope dilution of 3-carbon tracers between the experimentally measured pool and the precursor for the TCA-cycle OAA pool. Experimental tests are outlined which investigators can use to determine whether these pathways are significant in a specific steady-state system. The study indicated that flux through these five pathways can significantly affect the correction for isotope dilution. To correct for the effects of these pathways an alternative method for calculating isotope dilution is proposed using citrate to relate the specific activities of acetyl-CoA and OAA.

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Effect of thermal injury on the TCA cycle enzymes of Staphylococcus aureus MF 31 and Salmonella typhimurium 7136

Canadian Journal of Microbiology ◽

10.1139/m71-121 ◽

1971 ◽

Vol 17 (6) ◽

pp. 759-765 ◽

Cited By ~ 22

Author(s):

Richard I. Tomlins ◽

Merle D. Pierson ◽

Z. John Ordal

Keyword(s):

Lactate Dehydrogenase ◽

Malate Dehydrogenase ◽

Thermal Injury ◽

Specific Activity ◽

Fumarate Hydratase ◽

Tca Cycle ◽

Tca Cycle Enzymes ◽

The Tca Cycle ◽

Oxoglutarate Dehydrogenase ◽

Metabolic Activities

The heating of S. aureus MF-31 and S. typhimurium 7136 at 52C and 48C respectively, produced a sublethal heat injury. When injured cells were placed in fresh growth medium they recovered. The recovery of S. aureus was not inhibited by chloramphenicol. The metabolic activities of tricarboxylic acid (TCA) cycle enzymes, as well as other selected enzymes in crude extracts of normal and heat-injured cells of both microorganisms were assayed. In extracts from S. typhimurium there was some loss of specific activity with fumarate hydratase, glutamate dehydrogenase, fructose diphosphate aldolase, lactate dehydrogenase, and the NAD(P) oxidases as a result of heating. In extracts from S. aureus oxoglutarate dehydrogenase, malate dehydrogenase and lactate dehydrogenase were severely inactivated after heating. Other enzymes in comparison were only moderately sensitive to heat. No significant increase in enzyme activity was observed in extracts from injured cells of either microorganism. Re-naturation of lactate dehydrogenase and malate dehydrogenase occurred during the recovery of S. aureus both in the presence and absence of chloramphenicol. No renaturation of oxoglutarate dehydrogenase was found under the same conditions.

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

Journal of Bacteriology ◽

10.1128/jb.187.9.2967-2973.2005 ◽

2005 ◽

Vol 187 (9) ◽

pp. 2967-2973 ◽

Cited By ~ 71

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.

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Metabolic engineering of Bacillus amyloliquefaciens for enhanced production of S-adenosylmethionine by coupling of an engineered S-adenosylmethionine pathway and the tricarboxylic acid cycle

Biotechnology for Biofuels ◽

10.1186/s13068-019-1554-0 ◽

2019 ◽

Vol 12 (1) ◽

Cited By ~ 3

Author(s):

Liying Ruan ◽

Lu Li ◽

Dian Zou ◽

Cong Jiang ◽

Zhiyou Wen ◽

...

Keyword(s):

Metabolic Engineering ◽

Bacillus Amyloliquefaciens ◽

Tricarboxylic Acid Cycle ◽

Fold Increase ◽

Tca Cycle ◽

Tricarboxylic Acid ◽

Scale Production ◽

Free Medium ◽

Enhanced Production ◽

The Tca Cycle

Abstract Background S-Adenosylmethionine (SAM) is a critical cofactor involved in many biochemical reactions. However, the low fermentation titer of SAM in methionine-free medium hampers commercial-scale production. The SAM synthesis pathway is specially related to the tricarboxylic acid (TCA) cycle in Bacillus amyloliquefaciens. Therefore, the SAM synthesis pathway was engineered and coupled with the TCA cycle in B. amyloliquefaciens to improve SAM production in methionine-free medium. Results Four genes were found to significantly affect SAM production, including SAM2 from Saccharomyces cerevisiae, metA and metB from Escherichia coli, and native mccA. These four genes were combined to engineer the SAM pathway, resulting in a 1.42-fold increase in SAM titer using recombinant strain HSAM1. The engineered SAM pathway was subsequently coupled with the TCA cycle through deletion of succinyl-CoA synthetase gene sucC, and the resulted HSAM2 mutant produced a maximum SAM titer of 107.47 mg/L, representing a 0.59-fold increase over HSAM1. Expression of SAM2 in this strain via a recombinant plasmid resulted in strain HSAM3 that produced 648.99 mg/L SAM following semi-continuous flask batch fermentation, a much higher yield than previously reported for methionine-free medium. Conclusions This study reports an efficient strategy for improving SAM production that can also be applied for generation of SAM cofactors supporting group transfer reactions, which could benefit metabolic engineering, chemical biology and synthetic biology.

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Metabolism of glucose-14C, pyruvate-14C, and mannitol-14C by Melampsora lini. II. Conversion to soluble products

Canadian Journal of Botany ◽

10.1139/b68-069 ◽

1968 ◽

Vol 46 (4) ◽

pp. 453-460 ◽

Cited By ~ 10

Author(s):

D. Mitchell ◽

Michael Shaw

Keyword(s):

Pentose Phosphate Pathway ◽

Tricarboxylic Acid Cycle ◽

Rust Fungus ◽

Tca Cycle ◽

Tricarboxylic Acid ◽

Pentose Phosphate ◽

Soluble Carbohydrates ◽

Melampsora Lini ◽

Carbohydrate Fraction ◽

The Tca Cycle

Mycelium of the flax rust fungus (Melampsora lini (Pers.) Lév.), grown on flax cotyledons in tissue culture, had a mean [Formula: see text]of 4.1 and a mean C6/C1 ratio of 0.14, measured after 4 hours in radioactive glucose. The C6/C1 ratio increased with time and also after treatment with 10−5 M 2,4-dinitrophenol. The relative labelling of the (80%) ethanol-soluble carbohydrates, and organic and amino acid fractions after incubation with glucose-1-, -2-, or -6-14C also indicated preferential release of C1 as 14CO2. Trehalose (unknown A) was tentatively identified in the carbohydrate fraction and was mildly radioactive after incubation of the mycelium with labelled glucose for 3 hours. The principal radioactive products of glucose in this fraction were two unknowns, B and C, which were tentatively identified as mannitol and arabitol. The labelling patterns were consistent with their formation from intermediates of the pentose phosphate pathway. The distribution of radioactivity derived from glucose in alanine, glutamate, and aspartate also indicated that hexose or triose units formed in the pentose phosphate pathway were converted to pyruvate, which either gave rise to alanine or was further oxidized in the tricarboxylic acid cycle. Incubation with pyruvate-1-, -2-, or -3-14C for 3 hours gave rise to 14CO2 and labelled alanine, glutamate, and aspartate in a manner consistent with the operation of the TCA cycle. Mannitol-1-6-14C was not metabolized to any appreciable extent in this period, but did give rise to 14CO2 and to several unidentified compounds in the carbohydrate fraction.

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Host nutrient milieu drives an essential role for aspartate biosynthesis during invasiveStaphylococcus aureusinfection

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1922211117 ◽

2020 ◽

Vol 117 (22) ◽

pp. 12394-12401 ◽

Cited By ~ 2

Author(s):

Aimee D. Potter ◽

Casey E. Butrico ◽

Caleb A. Ford ◽

Jacob M. Curry ◽

Irina A. Trenary ◽

...

Keyword(s):

Metabolic Pathways ◽

Aerobic Glycolysis ◽

Bacterial Pathogen ◽

Tca Cycle ◽

Acid Synthesis ◽

Tricarboxylic Acid ◽

Staphylococcal Infection ◽

Amino Acid Synthesis ◽

The Tca Cycle ◽

Host Tissues

The bacterial pathogenStaphylococcus aureusis capable of infecting a broad spectrum of host tissues, in part due to flexibility of metabolic programs.S. aureus, like all organisms, requires essential biosynthetic intermediates to synthesize macromolecules. We therefore sought to determine the metabolic pathways contributing to synthesis of essential precursors during invasiveS. aureusinfection. We focused specifically on staphylococcal infection of bone, one of the most common sites of invasiveS. aureusinfection and a unique environment characterized by dynamic substrate accessibility, infection-induced hypoxia, and a metabolic profile skewed toward aerobic glycolysis. Using a murine model of osteomyelitis, we examined survival ofS. aureusmutants deficient in central metabolic pathways, including glycolysis, gluconeogenesis, the tricarboxylic acid (TCA) cycle, and amino acid synthesis/catabolism. Despite the high glycolytic demand of skeletal cells, we discovered thatS. aureusrequires glycolysis for survival in bone. Furthermore, the TCA cycle is dispensable for survival during osteomyelitis, andS. aureusinstead has a critical need for anaplerosis. Bacterial synthesis of aspartate in particular is absolutely essential for staphylococcal survival in bone, despite the presence of an aspartate transporter, which we identified as GltT and confirmed biochemically. This dependence on endogenous aspartate synthesis derives from the presence of excess glutamate in infected tissue, which inhibits aspartate acquisition byS. aureus. Together, these data elucidate the metabolic pathways required for staphylococcal infection within bone and demonstrate that the host nutrient milieu can determine essentiality of bacterial nutrient biosynthesis pathways despite the presence of dedicated transporters.

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Pattern of substrate utilization in vascular smooth muscle using 13C isotopomer analysis of glutamate

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.1998.275.6.h2227 ◽

1998 ◽

Vol 275 (6) ◽

pp. H2227-H2235 ◽

Cited By ~ 4

Author(s):

Tara M. Allen ◽

Christopher D. Hardin

Keyword(s):

Smooth Muscle ◽

Vascular Smooth Muscle ◽

Tca Cycle ◽

Tricarboxylic Acid ◽

Acetyl Coa ◽

Contractile State ◽

The Tca Cycle ◽

Direct Measurements ◽

Isotopomer Analysis ◽

13C Isotopomer Analysis

Although vascular smooth muscle (VSM) derives the majority of its energy from oxidative phosphorylation, controversy exists concerning which substrates are utilized by the tricarboxylic acid (TCA) cycle. We used 13C isotopomer analysis of glutamate to directly measure the entry of exogenous [13C]glucose and acetate and unlabeled endogenous sources into the TCA cycle via acetyl-CoA. Hog carotid artery segments denuded of endothelium were superfused with 5 mM [1-13C]glucose and 0–5 mM [1,2-13C]acetate at 37°C for 3–12 h. We found that both resting and contracting VSM preferentially utilize [1,2-13C]acetate compared with [1-13C]glucose and unlabeled substrates. The entry of glucose into the TCA cycle (30–60% of total entry via acetyl-CoA) exhibited little change despite alterations in contractile state or acetate concentrations ranging from 0 to 5 mM. We conclude that glucose and nonglucose substrates are important oxidative substrates for resting and contracting VSM. These are the first direct measurements of relative substrate entry into the TCA cycle of VSM during activation and may provide a useful method to measure alterations in VSM metabolism under physiological and pathophysiological conditions.

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