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Synthesis of alpha-ketoglutarate by reductive carboxylation of succinate in Veillonella, Selenomonas, and Bacteriodes species.

Evidence for reductive carboxylation of succinate to synthesize alpha-ketoglutarate was sought in anaerobic heterotrophs from the rumen and from other anaerobic habitats. Cultures were grown in media containing unlabeled energy substrates plus [14C]succinate, and synthesis of cellular glutamate with a much higher specific activity than that of cellular asparate was taken as evidence for alpha-ketoglutarate synthase activity. Our results indicate alpha-ketoglutarate synthase functions in Selenomonas ruminantium, Veillonella alcalescens, Bacteroides fragilis, Bacteroides vulgatus, Bacteroides uniformis, Bacteroides distasonis, and Bacteroides multiacidus. Evidence for this carboxylation was not found in strains representative of 10 other species.

Amino Acids↗

Microcalorimetric study of Escherichia coli aerobic growth: theoretical aspects of growth on succinic acid.

Two methods of investigation were used to evaluate the heat quantity associated with anabolic processes (qan) during the aerobic growth of Escherichia coli in a minimal medium containing succinic acid as the sole energy and carbon source. The study of the contribution of biosynthetic reactions from succinic acid and ammonia were investigated by both methods. The two qan values obtained were in excellent agreement and were found to be significant. Thus it was demonstrated that the contribution of anabolism strongly influenced the quantity of heat associated with microbial aerobic growth. The qan calculated as above explained the experimental enthalpy change which was recently reported.

Aerobiosis↗

Cloning and expression in Escherichia coli of sdhA, the structural gene for cytochrome b558 of the Bacillus subtilis succinate dehydrogenase complex.

Bacillus subtilis cytochrome b558 is a transmembrane protein which anchors succinate dehydrogenase (SDH) to the cytoplasmic membrane and is reduced by succinate. The structural gene for this cytochrome was cloned and expressed in Escherichia coli. Random BamHI or BglII fragments of B. subtilis 168 DNA were cloned in the BamHI site of plasmid pHV32. The derived plasmids were used to transform B. subtilis SDH mutants to chloramphenicol resistance by integration of the plasmid via DNA homology. Of some 3,000 transformants tested, 6 were SDH positive and had pHV32 integrated close to the sdh operon. Two plasmids, pKIM2 and pKIM4, with an insert of B. subtilis DNA of 5.7 and 3.4 kilobases, respectively, were generated by transforming E. coli with DNA from the SDH-positive transformants after cleavage with EcoRI or BglII and ligation. In E. coli carrying either of the two plasmids, about 4% of total membrane protein was B. subtilis cytochrome b558. E. coli (pKIM2) also contained antigen which reacted with antibodies specific for the flavoprotein and the iron-sulfur protein subunit of B. subtilis SDH. Enzymatically active, membrane-bound B. subtilis SDH could not be demonstrated in E. coli (pKIM2). The B. subtilis DNA insert in pKIM2 could transform B. subtilis sdhA (cytochrome b558), sdhB (flavoprotein), and sdhC (iron-sulfur protein) mutants to the wild type. The results suggest that pKIM2 carries the whole B. subtilis sdh operon. The data confirm the gene order and the proposed direction of transcription of the B. subtilis sdh operon. Most likely the sdh genes in E. coli(pKIM2) are controlled by their natural promoter.

Antigens, Bacterial↗

Electron-paramagnetic-resonance spectroscopy of Bacillus subtilis cytochrome b558 in Escherichia coli membranes and in succinate dehydrogenase complex from Bacillus subtilis membranes.

Cytochrome b558 of the Bacillus subtilis succinate dehydrogenase complex was studied by electron-paramagnetic-resonance (EPR) spectroscopy. The cytochrome amplified in Escherichia coli membranes by expression of the cloned cytochrome gene and in the succinate dehydrogenase complex immunoprecipitated from solubilized B. subtilis membranes, respectively, is shown to be low spin with a highly anisotropic (gmax approximately equal to 3.5) EPR signal. The amino acid residues most likely forming fifth and sixth axial ligands to heme in cytochrome b558 are discussed on the basis of the EPR signal and the recently determined gene sequence (K. Magnusson, M. Philips, J.R. Guest, and L. Rutberg, J. Bacteriol. 166:1067-1071, 1986) and in comparison with other b-type cytochromes.

Bacillus subtilis↗

Nucleotide sequence encoding the flavoprotein and iron-sulfur protein subunits of the Bacillus subtilis PY79 succinate dehydrogenase complex.

The nucleotide sequence of a 2.7-kilobase segment of DNA containing the sdhA and sdhB genes encoding the flavoprotein (Fp, sdhA) and iron-sulfur protein (Ip, sdhB) subunits of the succinate dehydrogenase of Bacillus subtilis was determined. This sequence extends the previously reported sequence encoding the cytochrome b558 subunit (sdhC) and completes the sequence of the sdh operon, sdhCAB. The predicted molecular weights for the Fp and Ip subunits, 65,186 (585 amino acids) and 28,285 (252 amino acids), agreed with the values determined independently for the labeled Fp and Ip antigens, although it appeared that the B. subtilis Fp was not functional after expression of the sdhA gene in Escherichia coli. Both subunits closely resembled the corresponding Fp and Ip subunits of the succinate dehydrogenase (SDH) and fumarate reductase of E. coli in size, composition, and amino acid sequence. The sequence homologies further indicated that the B. subtilis SDH subunits are equally related to the SDH and fumarate reductase subunits of E. coli but are less closely related than are the corresponding pairs of E. coli subunits. The regions of highest sequence conservation were identifiable as the catalytically significant flavin adenine dinucleotide-binding sites and cysteine clusters of the iron-sulfur centers.

Amino Acid Sequence↗

Effect of diphenylhydantoin on gamma aminobutyric acid (GABA) and succinate activity in rat Purkinje cells.

A study has been made of the effect of diphenylhydantoin (DPH) upon the levels of gamma aminobutyric acid (GABA) and succinic dehydrogenase in rat Purkinje cells. DPH was administered over 26 days in chronic experiments using controls receiving the same injection vehicle without DPH. Animals in this group received daily 1.25 mg/kg body weight, 12.5 mg/kg body weight, and 50 mg/kg body weight DPH. Acute experiments were carried out over the course of not more than four days, three groups of animals receiving 75 mg/kg body weight, 87.5 mg/kg body weight, and 100 mg/kg body weight DPH. No effect upon succinic dehydrogenase could be demonstrated at any dose level. There was a significant progressive loss of GABA with increasing dosage of DPH.

Aminobutyrates↗

Low platelet monoamine oxidase activity in headache: no correlation with phenolsulphotransferase, succinate dehydrogenase, platelet preparation method or smoking.

Platelet monoamine oxidase activity in male migrainous and cluster headache patients was significantly lower than in male controls, confirming our previous study. The activity range showed a normal distribution and low mean values could not be attributed to a subgroup with particularly low activity. When Corash 's platelet preparation method was used, with its high platelet yield, specific enzyme activities of a similar order were obtained. Thus, the low values encountered were not due to abnormal recovery within the platelet population. Two other enzyme activities, phenolsulphotransferase M and succinate dehydrogenase, were also measured in the same platelet samples. Although low succinate dehydrogenase activity was identified in the headache groups, it appeared to represent a separate phenomenon and there was no significant correlation between activity of either enzyme and that of monoamine oxidase. This shows that the low activity of platelet monoamine oxidase in headache is not related to a generalised platelet enzyme deficit. It was also shown that the low monoamine oxidase activity in the headache patients could not be attributed to smoking.

Arylsulfotransferase↗

Ultrastructural localization of succinate dehydrogenase in a self-parasitic isolate of Saprolegnia megasperma.

The enzyme succinate dehydrogenase (SDH, succinate: (acceptor) oxidoreductase, EC 1.3.99.1) was localized by the combined techniques of cytochemistry and electrom microscopy in the hyphae of a self-parasitizing isolate of Saprolegnia megasperma Coker. The enzyme was localized in the mitochondrial membranes; its activity was inhibited by malonate. Electron-dense deposits, whose formation was not prevented by the addition of malonate, appeared outsided of the hyphal cell walls. No evidence was found at the ultrastructural level within the vegetative hyphae for any abnormalities which could be linked to the phenomeonon of self-parasitism.

Fungi↗

Effect of the carbon source and cyclic AMP on isocitrate dehydrogenase, succinate dehydrogenase, and malate dehydrogenase in Klebsiella pneumoniae C3.

When strain C3 of Klebsiella pneumoniae is grown on a minimal medium with excess glucose, isocitrate dehydrogenase, malate dehydrogenase, and succinate dehydrogenase specific activities increase in the last period of the exponential growth phase and in the beginning of the stationary phase. Glucose exhaustion does not alter the development of malate dehydrogenase and succinate dehydrogenase, but specific activities are higher than those obtained with excess glucose. In contrast, glucose exhaustion can be correlated with a decrease of isocitrate dehydrogenase specific activity in the stationary phase. Induction of strain C3 isocitrate dehydrogenase by glucose in complex medium and repression by cAMP in mineral medium were observed. Glucose induction and the NADP/NADPH ratio are suggested as regulatory mechanisms controlling isocitrate dehydrogenase synthesis in the Enterobacteriaceae, but the former appears to be restricted to some Klebsiella strains.

Carbohydrates↗

Differential acute and long term actions of succinic acid monomethyl ester exposure on insulin-secreting BRIN-BD11 cells.

Esters of succinic acid are potent insulin secretagogues, and have been proposed as novel antidiabetic agents for type 2 diabetes. This study examines the effects of acute and chronic exposure to succinic acid monomethyl ester (SAM) on insulin secretion, glucose metabolism and pancreatic beta cell function using the BRIN-BD11 cell line. SAM stimulated insulin release in a dose-dependent manner at both non-stimulatory (1.1 mM) and stimulatory (16.7 mM) glucose. The depolarizing actions of arginine also stimulated a significant increase in SAM-induced insulin release but 2-ketoisocaproic acid (KIC) inhibited SAM induced insulin secretion indicating a possible competition between the preferential oxidative metabolism of these two agents. Prolonged (18 hour) exposure to SAM revealed decreases in the insulin-secretory responses to glucose, KIC, glyceraldehyde and alanine. Furthermore, SAM diminished the effects of non-metabolized secretagogues arginine and 3-isobutyl-1-methylxanthine (IBMX). While the ability of BRIN-BD11 cells to oxidise glucose was unaffected by SAM culture, glucose utilization was substantially reduced. Collectively, these data suggest that while SAM may enhance the secretory potential of non-metabolized secretagogues, it may also serve as a preferential metabolic fuel in preference to other important physiological nutrients and compromise pancreatic beta cell function following prolonged exposure.

1-Methyl-3-isobutylxanthine↗

Single muscle fibre analyses in 2 brothers with succinate dehydrogenase deficiency.

We report 2 brothers with hypertrophic cardiomyopathy, lipid storage and skeletal mitochondrial myopathy. Biochemical analyses detected complex II (succinate dehydrogenase) deficiency of the respiratory chain. Single fiber analyses using a microphotometer were performed and revealed reduced succinate dehydrogenase activity in all muscle fibers. This report shows the validity of kinetic microphotometric analysis in revealing complex II defects.

Adult↗

Comparative aspects of muscle fibre size and succinic dehydrogenase distribution in the longissimus dorsi muscle of several species of East African mammals.

The distribution of succinic dehydrogenase enzyme activity was investigated in frozen sections of longissimus dorsi muscle taken from several species of East African game animal (giraffe, hartebeest, wildebeest, oryx, gerenuk and dik-dik) as well as local zebu cattle. Muscle fibres were classified as red (high succinic dehydrogenase activity), white (low activity) or intermediate. The mean diameter and percentage distribution of each fibre type were noted as well as the overall mean muscle fibre diameter (MFD) for each species. The diameters of red muscle fibres were found to be between 54 and 62% of the diameters of the white muscle fibres for all species with MFD differences between species being up to over 100%. The variation in MFD was found to be significantly and positively correlated with live weight, when zebu values were omitted. It was found that the variation in MFD was significantly related to the diameter of muscle fibre types and not to the percent distribution of these fibre types.

Animals↗

Comparison between succinate dehydrogenase inhibition test and subrenal capsule assay for chemosensitivity testing.

The chemosensitivity result of the succinate dehydrogenase inhibition (SDI) test was compared with that of the subrenal capsule (SRC) assay in 23 human tumor tissues exposed to adriamycin (ADM), mitomycin C (MMC), cisplatin (DDP) and 5-fluorouracil (5-FU). The chemosensitivity was considered as positive when the succinate dehydrogenase (SD) activity of the drug-exposed cells was decreased to below 50% of that of control cells on day 3 in the SDI test, and the tumor size on day 6 was decreased to below -10% of that on day 0 in the SRC assay. Correlation rates between the decrease of SD activity in the SDI test and the decrease of tumor size in the SRC assay, using 23 evaluable cases in both assays, were r = 0.717 for ADM, r = 0.699 for MMC, r = 0.796 for DDP and r = 0.735 for 5-FU. The correlations of the chemosensitivity results were 73.9% for ADM, 73.9% for MMC, 82.6% for DDP and 60.9% for 5-FU. A positive correlation was noted between the in vitro and in vivo chemosensitivity results. This SDI test can serve as an effective tool for chemosensitivity testing.

Animals↗

Human hepatocellular carcinoma sensitivity to antitumor drugs assayed using the succinate dehydrogenase inhibition test.

Chemosensitivity of hepatocellular carcinoma tissues from 72 patients to 6 antitumor agents was assayed using the succinate dehydrogenase inhibition test. Sensitivity was positive in 47.2% of tissues exposed to adriamycin, 53.5% to mitomycin C, 10.3% to 5-fluorouracil, 51.5% to cisplatin and aclacinomycin A. respectively, and 52.9% to carboquone. Eight percent of the tissues were sensitive to all 6 drugs, while the resistance rate to all drugs was 36.5%. The remaining 55.5% were sensitive to only some of the drugs. When a comparison of the sensitivity to the 6 drugs was made between two different areas of tumors in 16 patients, positive or negative sensitivity was in a range of 76.9-92.9%. The hypovascular masses seen on the angiography and the histologically well differentiated tumors were resistant to adriamycin, a drug most commonly prescribed to treat patients with liver cancer. For some of these tumors, mitomycin C or carboquone may be effective. Our observation shows that the succinate dehydrogenase inhibition test is useful for determining which drugs will be effective for a particular tumor.

Aged↗

Use of topical lithium succinate in the treatment of seborrhoeic dermatitis.

Twenty-one patients with seborrhoeic dermatitis were included in an open trial of lithium succinate ointment (LSO) for a total duration of 8 weeks. The same clinician made assessments of the severity of redness, scaling, greasiness and overall clinical impression of the condition every 2 weeks. Because the results appeared to be satisfactory, we decided to perform a double-blind, placebo-controlled trial of LSO. Thirty patients with seborrhoeic dermatitis were included. The results also demonstrated the beneficial effect of LSO. A significantly higher number of patients treated with LSO showed remission or marked improvement compared with placebo. The main adverse events demonstrated consisted of minor transient skin irritation and/or stinging sensation. Studying the in vivo inhibitory effect of LSO on the growth of Pityrosporum revealed that Pityrosporum did not significantly have its growth inhibited by lithium. Topical lithium succinate appears to be a safe and an effective treatment for seborrhoeic dermatitis. The product presumably acts as an anti-inflammatory agent.

Adolescent↗

Maleic acid and succinic acid in fermented alcoholic beverages are the stimulants of gastric acid secretion.

Alcoholic beverages produced by fermentation (e.g., beer and wine) are powerful stimulants of gastric acid output and gastrin release in humans. The aim of this study was to separate and specify the gastric acid stimulatory ingredients in alcoholic beverages produced by fermentation. Yeast-fermented glucose was used as a simple model of fermented alcoholic beverages; it was stepwise separated by different methods of liquid chromatography, and each separated solution was tested in human volunteers for its stimulatory action on gastric acid output and gastrin release. Five substances were detected by high-performance liquid chromatography and were analyzed by mass spectrometry and 1H-13C nuclear magnetic resonance spectroscopy. At the end of the separation process of the five identified substances, only the two dicarboxylic acids, maleic acid and succinic acid, had a significant (P < 0.05) stimulatory action on gastric acid output (76% and 70% of fermented glucose, respectively), but not on gastrin release. When given together, they increased gastric acid output by 100% of fermented glucose and by 95% of maximal acid output. We therefore conclude that maleic acid and succinic acid are the powerful stimulants of gastric acid output in fermented glucose and alcoholic beverages produced by fermentation, and that gastrin is not their mediator of action.

Adult↗

The cytochemical localization of succinic dehydrogenase in the sperm mitochondria of the gastropod Biomphalaria glabrata. A comparison of two methods.

The mitochondrial derivative of the sperm of the gastropod pulmonate Biomphalaria glabrata was studies to ascertain succinic dehydrogenase localization cytochemically. Two techniques were compared. One technique depends on a tetrazolium salt that yields an osmiophilic formazan upon reduction. The other technique is dependent on the reduction of copper ferricyanide. The effects of several electron transport inhibitors were studied. The reaction product observed in the matrix of the mitochondrial derivative using the former technique is sensitive to rotenone and is believed to be nicotinamide adenine dinucleotide-dependent. The reaction product observed in the intracristal spaces using the copper ferricyanide method is insensitive to rotenone and is believed to cytochemically demonstrate succinic dehydrogenase in this material.

Animals↗

The SDH mutation database: an online resource for succinate dehydrogenase sequence variants involved in pheochromocytoma, paraganglioma and mitochondrial complex II deficiency.

BACKGROUND: The SDHA, SDHB, SDHC and SDHD genes encode the subunits of succinate dehydrogenase (succinate: ubiquinone oxidoreductase), a component of both the Krebs cycle and the mitochondrial respiratory chain. SDHA, a flavoprotein and SDHB, an iron-sulfur protein together constitute the catalytic domain, while SDHC and SDHD encode membrane anchors that allow the complex to participate in the respiratory chain as complex II. Germline mutations of SDHD and SDHB are a major cause of the hereditary forms of the tumors paraganglioma and pheochromocytoma. The largest subunit, SDHA, is mutated in patients with Leigh syndrome and late-onset optic atrophy, but has not as yet been identified as a factor in hereditary cancer. DESCRIPTION: The SDH mutation database is based on the recently described Leiden Open (source) Variation Database (LOVD) system. The variants currently described in the database were extracted from the published literature and in some cases annotated to conform to current mutation nomenclature. Researchers can also directly submit new sequence variants online. Since the identification of SDHD, SDHC, and SDHB as classic tumor suppressor genes in 2000 and 2001, studies from research groups around the world have identified a total of 120 variants. Here we introduce all reported paraganglioma and pheochromocytoma related sequence variations in these genes, in addition to all reported mutations of SDHA. The database is now accessible online. CONCLUSION: The SDH mutation database offers a valuable tool and resource for clinicians involved in the treatment of patients with paraganglioma-pheochromocytoma, clinical geneticists needing an overview of current knowledge, and geneticists and other researchers needing a solid foundation for further exploration of both these tumor syndromes and SDHA-related phenotypes.

Codon, Nonsense↗