Tricarboxylic acid-cycle and related enzymes in restricted facultative methylotrophs.
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ABSTRACT:
SUBMITTER: Colby J
PROVIDER: S-EPMC1165569 | biostudies-other | 1975 Jun
REPOSITORIES: biostudies-other
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Project description:Extracts of trimethylamine-grown W6A and W3A1 (type M restricted facultative methylotrophs) contain trimethylamine dehydrogenase whereas similar extracts of Bacillus PM6 and Bacillus S2A1 (type L restricted facultative methylotrophs) contain trimethylamine mono-oxygenase and trimethylamine N-oxide demethylase but no trimethylamine dehydrogenase. Extracts of the restricted facultatives and of the obligate methylotroph C2A1 contain hexulose phosphate synthase-hexulose phosphate isomerase activity; hydroxypyruvate reductase was not detected. Neither the restricted facultatives nor the obligates 4B6 and C2A1 contain all the enzymes of the hexulose phosphate cycle of formaldehyde assimilation as originally proposed by Kemp & Quayle (1967). Organisms PM6 and S2A1 lack transaldolase and use a modified cycle involving sedoheptulose 1,7-diphosphate and sedoheptulose diphosphatase. The obligates 4B6 and C2A1, and the type M organisms W6A and W3A1, use a different modification of the assimilatory hexulose phosphate cycle involving the Entner-Doudoroff-pathway enzymes phosphogluconate dehydratase and phospho-2-keto-3-deoxygluconate aldolase. The lack of fructose diphosphate aldolase and hexose diphosphatase in these organisms may be a partial explanation of their restricted growth-substrate range. Enzymological evidence suggests that all the obligates and the restricted facultatives use a dissimilatory hexulose phosphate cycle to accomplish the complete oxidation of formaldehyde to CO2 and water.
| S-EPMC1165570 | biostudies-other
Project description:BackgroundTricarboxylic acid (TCA) cycle deregulation may predispose to cardiovascular diseases, but the role of TCA cycle-related metabolites in the development of atrial fibrillation (AF) and heart failure (HF) remains unexplored. This study sought to investigate the association of TCA cycle-related metabolites with risk of AF and HF.MethodsWe used two nested case-control studies within the PREDIMED study. During a mean follow-up for about 10 years, 512 AF and 334 HF incident cases matched by age (±5 years), sex and recruitment center to 616 controls and 433 controls, respectively, were included in this study. Baseline plasma levels of citrate, aconitate, isocitrate, succinate, malate and d/l-2-hydroxyglutarate were measured with liquid chromatography-tandem mass spectrometry. Multivariable conditional logistic regression models were fitted to estimate odds ratios (OR) and 95% confidence intervals (95% CI) for metabolites and the risk of AF or HF. Potential confounders included smoking, family history of premature coronary heart disease, physical activity, alcohol intake, body mass index, intervention groups, dyslipidemia, hypertension, type 2 diabetes and medication use.ResultsComparing extreme quartiles of metabolites, elevated levels of succinate, malate, citrate and d/l-2-hydroxyglutarate were associated with a higher risk of AF [ORQ4 vs. Q1 (95% CI): 1.80 (1.21-2.67), 2.13 (1.45-3.13), 1.87 (1.25-2.81) and 1.95 (1.31-2.90), respectively]. One SD increase in aconitate was directly associated with AF risk [OR (95% CI): 1.16 (1.01-1.34)]. The corresponding ORs (95% CI) for HF comparing extreme quartiles of malate, aconitate, isocitrate and d/l-2-hydroxyglutarate were 2.15 (1.29-3.56), 2.16 (1.25-3.72), 2.63 (1.56-4.44) and 1.82 (1.10-3.04), respectively. These associations were confirmed in an internal validation, except for aconitate and AF.ConclusionThese findings underscore the potential role of the TCA cycle in the pathogenesis of cardiac outcomes.
| S-EPMC9206868 | biostudies-literature
Project description:Using a closed-head impact acceleration model of mild or severe traumatic brain injury (mTBI or sTBI, respectively) in rats, we evaluated the effects of graded head impacts on the gene and protein expressions of pyruvate dehydrogenase (PDH), as well as major enzymes of mitochondrial tricarboxylic acid cycle (TCA). TBI was induced in anaesthetized rats by dropping 450 g from 1 (mTBI) or 2 m height (sTBI). After 6 h, 12 h, 24 h, 48 h, and 120 h gene expressions of enzymes and subunits of PDH. PDH kinases and phosphatases (PDK1-4 and PDP1-2, respectively), citrate synthase (CS), isocitrate dehydrogenase (IDH), oxoglutarate dehydrogenase (OGDH), succinate dehydrogenase (SDH), succinyl-CoA synthase (SUCLG), and malate dehydrogenase (MDH) were determined in whole brain extracts (n = 6 rats at each time for both TBI levels). In the same samples, the high performance liquid chromatographic (HPLC) determination of acetyl-coenzyme A (acetyl-CoA) and free coenzyme A (CoA-SH) was performed. Sham-operated animals (n = 6) were used as controls. After mTBI, the results indicated a general transient decrease, followed by significant increases, in PDH and TCA gene expressions. Conversely, permanent PDH and TCA downregulation occurred following sTBI. The inhibitory conditions of PDH (caused by PDP1-2 downregulations and PDK1-4 overexpression) and SDH appeared to operate only after sTBI. This produced almost no change in acetyl-CoA and free CoA-SH following mTBI and a remarkable depletion of both compounds after sTBI. These results again demonstrated temporary or steady mitochondrial malfunctioning, causing minimal or profound modifications to energy-related metabolites, following mTBI or sTBI, respectively. Additionally, PDH and SDH appeared to be highly sensitive to traumatic insults and are deeply involved in mitochondrial-related energy metabolism imbalance.
| S-EPMC6888669 | biostudies-literature
Project description:Alterations of activity and expression in tricarboxylic acid (TCA) cycle key enzymes have been indicated in several malignancies, including hepatocellular carcinoma (HCC). They play an important role in the progression of cancer. However, the impact of single nucleotide polymorphisms (SNPs) in genes encoding these key enzymes on the recurrence of HCC has not been investigated. In this study, we genotyped 17 SNPs in genes encoding TCA cycle key enzymes and analyzed their association with recurrence-free survival (RFS) in a cohort of 492 Chinese HCC patients by Cox proportional hazard model and survival tree analysis. We identified 7 SNPs in SDHC, SDHD, FH, and IDH2 genes to be significantly associated with the RFS of HCC patients. Moreover, all these SNPs were associated with the early recurrence (within 2 years after surgery) risk of diseases. Cumulative effect analysis showed that these SNPs exhibited a dose-dependent effect on the overall and early recurrence. Further stratified analysis suggested that number of risk genotypes modified the protective effect on HCC recurrence conferred by transcatheter arterial chemoembolization treatment. Finally, the survival tree analysis revealed that SNP rs10789859 in SDHD gene was the primary factor contributing to HCC recurrence in our population. To the best of our knowledge, we for the first time observed the association between SNPs in genes encoding TCA cycle key enzymes and HCC recurrence risk. Further observational and functional studies are needed to validate our findings and generalize its clinical usage.
| S-EPMC4404327 | biostudies-literature
Project description:Background: Glycolysis/gluconeogenesis and tricarboxylic acid (TCA) cycle metabolites have been associated with type 2 diabetes (T2D). However, the associations of these metabolites with T2D incidence and the potential effect of dietary interventions remain unclear.Objectives: We aimed to evaluate the association of baseline and 1-y changes in glycolysis/gluconeogenesis and TCA cycle metabolites with insulin resistance and T2D incidence, and the potential modifying effect of Mediterranean diet (MedDiet) interventions.Methods: We included 251 incident T2D cases and 638 noncases in a nested case-cohort study within the PREDIMED Study during median follow-up of 3.8 y. Participants were allocated to MedDiet + extra-virgin olive oil, MedDiet + nuts, or control diet. Plasma metabolites were measured using a targeted approach by LC-tandem MS. We tested the associations of baseline and 1-y changes in glycolysis/gluconeogenesis and TCA cycle metabolites with subsequent T2D risk using weighted Cox regression models and adjusting for potential confounders. We designed a weighted score combining all these metabolites and applying the leave-one-out cross-validation approach.Results: Baseline circulating concentrations of hexose monophosphate, pyruvate, lactate, alanine, glycerol-3 phosphate, and isocitrate were significantly associated with higher T2D risk (17-44% higher risk for each 1-SD increment). The weighted score including all metabolites was associated with a 30% (95% CI: 1.12, 1.51) higher relative risk of T2D for each 1-SD increment. Baseline lactate and alanine were associated with baseline and 1-y changes of homeostasis model assessment of insulin resistance. One-year increases in most metabolites and in the weighted score were associated with higher relative risk of T2D after 1 y of follow-up. Lower risks were observed in the MedDiet groups than in the control group although no significant interactions were found after adjusting for multiple comparisons.Conclusions: We identified a panel of glycolysis/gluconeogenesis-related metabolites that was significantly associated with T2D risk in a Mediterranean population at high cardiovascular disease risk. A MedDiet could counteract the detrimental effects of these metabolites.This trial was registered at controlled-trials.com as ISRCTN35739639.
| S-EPMC7138680 | biostudies-literature
Project description:1. Twelve bacterial isolates that grow with trimethylamine as sole source of carbon and energy were obtained in pure culture. All the isolates grow on methylamine, dimethylamine and trimethylamine. One isolate, bacterium 4B6, grows only on these methylamines whereas another isolate, bacterium C2A1, also grows on methanol but neither grows on methane; these two organisms are obligate methylotrophs. The other ten isolates grow on a variety of C(i) and other organic compounds and are therefore facultative methylotrophs. 2. Washed suspensions of the obligate methylotrophs bacteria 4B6 and C2A1, and of the facultative methylotrophs bacterium 5B1 and Pseudomonas 3A2, all grown on trimethylamine, oxidize trimethylamine, dimethylamine, formaldehyde and formate; only bacterium 5B1 and Ps. 3A2 oxidize trimethylamine N-oxide; only bacterium 4B6 does not oxidize methylamine. 3. Cell-free extracts of trimethylamine-grown bacteria 4B6 and C2A1 contain a trimethylamine dehydrogenase that requires phenazine methosulphate as primary hydrogen acceptor, and evidence is presented that this enzyme is important for the growth of bacterium 4B6 on trimethylamine. 4. Cell-free extracts of eight facultative methylotrophs, including bacterium 5B1 and Ps. 3A2, do not contain trimethylamine dehydrogenase but contain instead a trimethylamine monooxygenase and trimethylamine N-oxide demethylase. It is concluded that two different pathways for the oxidation of trimethylamine occur amongst the isolates.
| S-EPMC1177564 | biostudies-other
Project description:The tricarboxylic acid (TCA) cycle is a central hub of cellular metabolism, oxidizing nutrients to generate reducing equivalents for energy production and critical metabolites for biosynthetic reactions. Despite the importance of the products of the TCA cycle for cell viability and proliferation, mammalian cells display diversity in TCA-cycle activity. How this diversity is achieved, and whether it is critical for establishing cell fate, remains poorly understood. Here we identify a non-canonical TCA cycle that is required for changes in cell state. Genetic co-essentiality mapping revealed a cluster of genes that is sufficient to compose a biochemical alternative to the canonical TCA cycle, wherein mitochondrially derived citrate exported to the cytoplasm is metabolized by ATP citrate lyase, ultimately regenerating mitochondrial oxaloacetate to complete this non-canonical TCA cycle. Manipulating the expression of ATP citrate lyase or the canonical TCA-cycle enzyme aconitase 2 in mouse myoblasts and embryonic stem cells revealed that changes in the configuration of the TCA cycle accompany cell fate transitions. During exit from pluripotency, embryonic stem cells switch from canonical to non-canonical TCA-cycle metabolism. Accordingly, blocking the non-canonical TCA cycle prevents cells from exiting pluripotency. These results establish a context-dependent alternative to the traditional TCA cycle and reveal that appropriate TCA-cycle engagement is required for changes in cell state.
2021-12-01 | GSE183434 | GEO
Project description:BackgroundC-type cytoplasmic male sterility (CMS-C) is one of the three major types of cytoplasmic male sterility (CMS) in maize. Rf4 is a dominant restorer gene for CMS-C and has great value in hybrid maize breeding, but little information concerning its functional mechanism is known.ResultsTo reveal the functional mechanism of Rf4, we developed a pair of maize near-isogenic lines (NILs) for the Rf4 locus, which included a NIL_rf4 male-sterile line and a NIL_Rf4 male fertility-restored line. Genetic analysis and molecular marker detection indicated that the male fertility of NIL_Rf4 was controlled by Rf4. Whole-genome sequencing demonstrated genomic differences between the two NILs was clustered in the Rf4 mapping region. Unmapped reads of NILs were further assembled to uncover Rf4 candidates. RNA-Seq was then performed for the developing anthers of the NILs to identify critical genes and pathways associated with fertility restoration. A total of 7125 differentially expressed genes (DEGs) were identified. These DEGs were significantly enriched in 242 Gene Ontology (GO) categories, wherein 100 DEGs were involved in pollen tube development, pollen tube growth, pollen development, and gametophyte development. Homology analysis revealed 198 male fertility-related DEGs, and pathway enrichment analysis revealed that 58 DEGs were enriched in cell energy metabolism processes involved in glycolysis, the pentose phosphate pathway, and pyruvate metabolism. By querying the Plant Reactome Pathway database, we found that 14 of the DEGs were involved in the mitochondrial tricarboxylic acid (TCA) cycle and that most of them belonged to the isocitrate dehydrogenase (IDH) and oxoglutarate dehydrogenase (OGDH) enzyme complexes. Transcriptome sequencing and real-time quantitative PCR (qPCR) showed that all the above TCA cycle-related genes were up-regulated in NIL_Rf4. The results of our subsequent enzyme-linked immunosorbent assay (ELISA) experiments pointed out that the contents of both the IDH and OGDH enzymes accumulated more in the spikelets of NIL_Rf4 than in those of NIL_rf4.ConclusionThe present research provides valuable genomic resources for deep insight into the molecular mechanism underlying CMS-C male fertility restoration. Importantly, our results indicated that genes involved in energy metabolism, especially some mitochondrial TCA cycle-related genes, were associated with maize CMS-C male fertility restoration.
| S-EPMC6136215 | biostudies-literature
Project description:Methylmalonic acidemia (MMA) is a propionate pathway disorder caused by dysfunction of the mitochondrial enzyme methylmalonyl-CoA mutase (MMUT). MMUT catalyzes the conversion of methylmalonyl-CoA to succinyl-CoA, an anaplerotic reaction which feeds into the tricarboxylic acid (TCA) cycle. As part of the pathological mechanisms of MMA, previous studies have suggested there is decreased TCA activity due to a "toxic inhibition" of TCA cycle enzymes by MMA related metabolites, in addition to reduced anaplerosis. Here, we have utilized mitochondria isolated from livers of a mouse model of MMA (Mut-ko/ki) and their littermate controls (Ki/wt) to examine the amounts and enzyme functions of most of the TCA cycle enzymes. We have performed mRNA quantification, protein semi-quantitation, and enzyme activity quantification for TCA cycle enzymes in these samples. Expression profiling showed increased mRNA levels of fumarate hydratase in the Mut-ko/ki samples, which by contrast had reduced protein levels as detected by immunoblot, while all other mRNA levels were unaltered. Immunoblotting also revealed decreased protein levels of 2-oxoglutarate dehydrogenase and malate dehydrogenase 2. Interesting, the decreased protein amount of 2-oxoglutarate dehydrogenase was reflected in decreased activity for this enzyme while there is a trend towards decreased activity of fumarate hydratase and malate dehydrogenase 2. Citrate synthase, isocitrate dehydrogenase 2/3, succinyl-CoA synthase, and succinate dehydrogenase are not statistically different in terms of quantity of enzyme or activity. Finally, we found decreased activity when examining the function of methylmalonyl-CoA mutase in series with succinate synthase and succinate dehydrogenase in the Mut-ko/ki mice compared to their littermate controls, as expected. This study demonstrates decreased activity of certain TCA cycle enzymes and by corollary decreased TCA cycle function, but it supports decreased protein quantity rather than "toxic inhibition" as the underlying mechanism of action. SUMMARY: Methylmalonic acidemia (MMA) is an inborn metabolic disorder of propionate catabolism. In this disorder, toxic metabolites are considered to be the major pathogenic mechanism for acute and long-term complications. However, despite optimized therapies aimed at reducing metabolite levels, patients continue to suffer from late complications, including metabolic stroke and renal insufficiency. Since the propionate pathway feeds into the tricarboxylic acid (TCA) cycle, we investigated TCA cycle function in a constitutive MMA mouse model. We demonstrated decreased amounts of the TCA enzymes, Mdh2 and Ogdh as semi-quantified by immunoblot. Enzymatic activity of Ogdh is also decreased in the MMA mouse model compared to controls. Thus, when the enzyme amounts are decreased, we see the enzymatic activity also decreased to a similar extent for Ogdh. Further studies to elucidate the structural and/or functional links between the TCA cycle and propionate pathways might lead to new treatment approaches for MMA patients.
| S-EPMC6903684 | biostudies-literature
Project description:New antimalarial drugs are urgently needed to control drug-resistant forms of the malaria parasite Plasmodium falciparum. Mitochondrial electron transport is the target of both existing and new antimalarials. Herein, we describe 11 genetic knockout (KO) lines that delete six of the eight mitochondrial tricarboxylic acid (TCA) cycle enzymes. Although all TCA KOs grew normally in asexual blood stages, these metabolic deficiencies halted life-cycle progression in later stages. Specifically, aconitase KO parasites arrested as late gametocytes, whereas ?-ketoglutarate-dehydrogenase-deficient parasites failed to develop oocysts in the mosquitoes. Mass spectrometry analysis of (13)C-isotope-labeled TCA mutant parasites showed that P. falciparum has significant flexibility in TCA metabolism. This flexibility manifested itself through changes in pathway fluxes and through altered exchange of substrates between cytosolic and mitochondrial pools. Our findings suggest that mitochondrial metabolic plasticity is essential for parasite development.
| S-EPMC4394047 | biostudies-literature