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Bioconversion of fumarate to succinate using glycerol as a carbon source.

In this study, a facultative bacterium that converts fumarate to succinate at a high yield was isolated. The yield of bioconversion was enhanced about 1.2 times by addition of glucose into culture medium at an initial concentration of 6 g/L. When the initial cell density was high (2 g/L), the succinate produced at pH 7.0 for initial fumarate concentrations of 30, 50, 80, and 100 g/L were 29.3, 40.9, 63.6, and 82.5 g/L, respectively, showing an increase with the initial fumarate concentration. The high yield of 96.8%/mole of fumarate in just 4 h was obtained at the initial fumarate concentration of 30 g/L. Comparing these values to those obtained with low cell culture (0.2 g/L), we found that the amount of succinate produced was similar, but the production rate in the high cell culture was about three times higher than was the case in the low cell culture. This strain converted fumarate to succinate at a rate of 3.5 g/L.h under the sparge of CO2.

Biodegradation, Environmental↗

Altered distribution pattern of Na+-K+-ATPase and succinate dehydrogenase activities along the nephron in human acute post-transplant renal failure.

The catalytic activities of Na+-K+-ATPase and succinate dehydrogenase, marker enzymes for active salt reabsorptive capacity of renal basolateral plasma membranes and for respiratory capacity of mitochondrial cristae membranes, were studied in the maintenance phase of human acute post-transplant renal failure. Biopsies of 4 kidney-allografts taken at transplantation operation and additionally at different post-transplantation periods, either with good function or in various stages of dysfunction, were compared with the unaffected part of a human kidney nephrectomized due to hypernephroma. In single nephron segments, Na+-K+-ATPase activity was determined after microdissection by microfluorometry, and succinate dehydrogenase activity was determined by a microphotometric procedure in stained cryosections. In intraoperative and postoperative biopsies of a well-functioning allograft, both Na+-K+-ATPase and succinate dehydrogenase activities did not differ from those of normal renal tissue. In contrast, the catalytic activities were found to be decreased in the distal tubules of 2 anuric allografts when compared with their intraoperative controls. In addition, succinate dehydrogenase activity was reduced in distal tubules of a recovering allograft. Catalytic activities appeared to be unaffected in glomeruli, proximal tubules, and collecting ducts. It is suggested that the predominant distal tubular alterations with regard to these parameters are a consequence of increased distal tubular vulnerability due to circulatory and metabolic conditions.

Acute Kidney Injury↗

Inhibition of succinic semialdehyde dehydrogenase by N-formylglycine.

N-formylglycine was developed as a dead-end inhibitor of the succinic semialdehyde dehydrogenase reaction. At 4 mM, it inhibited Aspergillus niger succinic semialdehyde dehydrogenase by 40%. N-formylglycine is a reversible, complete inhibitor; the inhibition is competitive with succinic semialdehyde and uncompetitive with respect to NAD+ and the Ki values are 4.9 and 10.4 mM respectively. Potato succinic semialdehyde dehydrogenase is also inhibited by N-formylglycine to a similar extent, the nature of the inhibition being identical to that observed with the A. niger enzyme.

Aldehyde Oxidoreductases↗

[Anaerobic formation of succinate and facilitation of its oxidation -- possible mechanisms of cell adaptation to oxygen deficiency].

An important role of anaerobic formation of succinate in anoxic and hypoxic states and the activation of succinate oxidation under hypoxia were shown. It was concluded that, for maintaining the energetics of animal cells under conditions of oxygen deficiency, it is advisable to use substrates capable of participating in the anaerobic formation of succinate, whereas under hypoxia it is reasonable to use succinate itself.

Adaptation, Biological↗

[Diabetes mellitus in the elderly: succinic acid compounds in treating diabetic neuropathies].

To investigate the influence of the succinic acid treatment on geriatric patients with type 2 diabetes. Succinic Acid has some positive biological properties. One of its is a neglecting of an aerobic glycolysis. In this study we evaluated the efficacy of the combination of the succinic acid ("MITOMIN") on treating of diabetic neuropathy of geriatric patients with type 2 diabetes. The analysis was carried out using 26 patients (aged 60-76 years). The duration of diabetes was 9.15 +/- 1.43 years. Biomedical parameters were measured by standard methods; microalbuminuria was measured by "Micral-Test". Quality of life (psychosocial disorders) was estimated with the help of "SANDOZ"-scale for geriatric assessment. The therapy was assigned 1.5 g of mitomin per day during a month. All patients were examined on having late diabetic complications: 7.69%--had diabetic retinopathy; 11.54%--diabetic nephropathy; 73.08%--diabetic neuropathy; 46.15%--chronic failure of brain vessels; 11.5%--macroangiopathy of lower extremities and 100%--had ischeamic heart disease of different levels. Mitomin therapy improved basal and postprandial glycemic control (NS), variance of pallesthesia (p < 0.001), parameters of quality of life, i.e. depression (p < 0.001), anxiety (p < 0.01), short memory (p < 0.05) and emotionality (p < 0.001). Mitomin therapy plays a positive role in management of elderly patients with type 2 diabetes. It improves glycemic control, pallestesia and quality of life. Combination of succinic acid renders central and peripheral neuropathy protective efficacy.

Aged↗

The sequence of the flavoprotein subunit of bovine heart succinate dehydrogenase.

The cDNA sequence of the flavoprotein subunit of bovine heart succinate dehydrogenase is reported. This is the first complete eukaryotic sequence of the flavoprotein subunit to be characterized, and it encodes a 665-amino acid protein that consists of a presequence and a 621-residue mature protein. The deduced bovine sequence shows homology to the corresponding peptides of prokaryotic succinate dehydrogenase and the related fumarate reductases; in particular, there is good overall homology (48%) to the flavoprotein subunit of Escherichia coli succinate dehydrogenase. The conserved sequences comprising the active site and those involved in FAD binding are also found in the bovine protein. The active site of the bovine polypeptide contains a cysteine that confers sensitivity of the enzyme to sulfhydryl reagents; this cysteine is only present in some sequences and thus provides a discriminatory biochemical marker. A putative flavoprotein subunit of human placental succinate dehydrogenase (partial sequence) that lacks this critical cysteine (Malcovati, M., Marchetti, T., Zanelli, T., and Tenchini, M. L. (1991) in Flavins and Flavoproteins 1990 (Curti, B., Ronchi, S., and Zanetti, G., eds) pp. 727-730, Walter de Gruyter & Co., Berlin) has only 16% homology to the bovine heart flavoprotein subunit. However, we show that the enzyme from human placenta is as sensitive to N-ethylmaleimide as that from bovine tissues. In addition, a transcript in human placenta and muscle hybridizes to the bovine heart flavoprotein cDNA and is the same size as that in bovine tissues.

Amino Acid Sequence↗

Cytochrome b560 (QPs1) of mitochondrial succinate-ubiquinone reductase. Immunochemistry, cloning, and nucleotide sequencing.

Mitochondrial succinate-ubiquinone reductase is composed of two parts, a water-soluble succinate dehydrogenase and a two-polypeptide membrane-anchoring protein fraction (QPs). The larger polypeptide of QPs is believed to be associated with cytochrome b560 (QPs1). The structure of QPs1 was studied by immunochemistry and molecular cloning and sequencing. Antibodies against QPs1 were raised in rabbits, purified, and characterized by enzyme-linked immunosorbent assay and Western blotting. The purified antibodies inhibited 75% of the reconstitutive activity of QPs and reacted with both submitochondrial particles (SMP) and mitoplasts. The binding of these antibodies to SMP was greatly increased when succinate dehydrogenase was removed from SMP by alkaline treatment, indicating that QPs1 is a transmembranous protein and that some of its specific epitopes are covered by succinate dehydrogenase. Anti-QPs1 antibodies were used to screen one cDNA clone encoding QPs1 from a bovine heart cDNA lambda gt11 expression library. The cDNA insert is 946 base pairs with an open reading frame of 396 base pairs that encodes for 132 amino acid residues. The molecular weight of QPs1, calculated from the deduced amino acid sequence, is 14,320. Although the apparent molecular weight of QPs1, estimated by high resolution SDS-polyacrylamide gel electrophoresis, is approximately 11,000, the existence of a presequence was ruled out by mass spectrometric analysis of protein fragments. QPs1 is a very hydrophobic protein. Three probable membrane-spanning segments were revealed by a hydropathy plot of the sequence. QPs1 has a higher sequence similarity to the sdhC peptide of Escherichia coli than to the sdhC peptide (cytochrome b558) of Bacillus subtilis. Like the bacterial proteins, QPs1 has 2 conserved histidines at positions 34 and 90. The conserved nature and similar location of these 2 histidines, on the matrix-side surface of the membrane, suggest that they are involved in heme ligation of cytochrome b560.

Amino Acid Sequence↗

[Use of sodium succinate in complex treatment of ischemic stroke].

The results of the study of sodium succinate efficacy in the treatment of 60 patients with ischemic stroke in comparison to the control group, not receiving antioxidants, are presented. The best results of sodium succinate were achieved in regress of local and general brain symptoms. The authors emphasize a possibility of the treatment at home and in outpatient clinic as well as cheaper price of sodium succinate versus other succinates.

Adult↗

Isolation and nucleotide sequence of the Saccharomyces cerevisiae gene for the succinate dehydrogenase flavoprotein subunit.

Succinate dehydrogenase (EC 1.3.99.1) of the mitochondrial inner membrane is a four-subunit membrane-bound enzyme that catalyzes the oxidation of succinate to fumarate and the transfer of electrons into the electron transport chain to oxygen. The catalytic domain of the enzyme is composed of a flavoprotein subunit which contains a covalently attached FAD cofactor and an iron-sulfur subunit with three nonidentical iron-sulfur clusters. We have isolated a complete genomic clone for the flavoprotein subunit of the succinate dehydrogenase from Saccharomyces cerevisiae and determined its nucleotide sequence. The sequence predicts a protein of 70,185 Da (640 amino acids) that shows more similarity to the Escherichia coli succinate dehydrogenase flavoprotein subunit than it does to the only other mitochondrial homologue, the human flavoprotein subunit. The yeast flavoprotein subunit precursor was synthesized in a cell-free translation system and shown to possess a mitochondrial targeting sequence that directs its import into isolated, energized mitochondria where it is processed by the matrix-localized protease. The genes for the flavoprotein and the iron-sulfur subunits reside on different chromosomes and hence form different transcriptional units.

Amino Acid Sequence↗

[The effect of succinate on respiration, transamination, and pyruvate formation in cells of the yeast Dipodascus magnusii].

The effect of succinate on the growth and respiration of the yeast Dipodascus magnusii VKM Y-1072, which is auxotrophic for thiamine and biotin, was studied. The addition of succinate to a culture grown on glucose was found to activate the respiration of cells on various substrates by enhancing the processes related to transamination reactions. In this case, aerobic fermentation (ethanol production) decreased, whereas pyruvate production increased. When succinate was added to the medium as the sole carbon source, it supported yeast growth in the absence of one of the two vitamins, thiamine or biotin, but not both. The yeast metabolism was completely respiratory, without any signs of aerobic fermentation. A drastic rise in pyruvate production in the yeast grown on glucose in the presence of succinate and the absence of biotin are also indicative of metabolic changes.

Biotin↗

Deficient utilization of succinate in a sterol 14 alpha-demethylation mutant of Candida albicans.

Growth of a wild-type strain (KD14) started after a lag of approximately 15 h when glucose-grown cells of Candida albicans were incubated at 25 degrees C, with succinate as a sole energy/carbon source. No growth was observed, however, for a sterol 14 alpha-demethylation mutant (KD4900). During the lag period, the wild-type cells retained their respiratory activity, as measured with glucose as an added substrate, and developed the capacity to incorporate labelled succinate. In contrast, incubation with succinate for 15 h resulted in diminished respiratory activity and failed to induce the succinate uptake capacity of the mutant cells. These two differences from the wild-type seem to account for the inability of the mutant to grow in the presence of a non-fermentable substrate.

Candida albicans↗

Acetate kinase from Veillonella alcalescens. Regulation by succinate and substrates.

The kinetic properties of acetate kinase from Veillonella alcalescens were investigated. In the presence of high concentrations of nucleotide both forward and reverse reactions were observed. In the presence of succinate the degree of cooperativity between subunits of the homodimer decreased, i.e. the Hill coefficient, n, decreased from 2.5 to 1.4 for acetyl phosphate in the presence of succinate. At low substrate concentrations hyperbolic kinetic data were observed with succinate. We have proposed a modified version of the concerted symmetry model to describe the kinetics observed with this enzyme. The primary differentiating feature of the proposed model is the requirement for activator ligand binding for catalysis. In the absence of succinate, the substrate (acetate or acetyl phosphate) also functions as an activating ligand.

Acetate Kinase↗

X-ray studies on crystalline complexes involving amino acids and peptides. XIX. Effects of change in chirality in the complexes of succinic acid with DL- and L-arginine.

Crystals of DL-arginine hemisuccinate dihydrate (I)(monoclinic; P2(1)/c; a = 5.292, b = 16.296. c = 15.203 A; beta = 92.89 degrees; Z = 4) and L-arginine hemisuccinate hemisuccinic acid monohydrate (II) (triclinic; P1; a = 5.099; b = 10.222, c = 14.626 A; alpha = 77.31, beta = 89.46, gamma = 78.42 degrees; Z = 2) were grown under identical conditions from aqueous solutions of the components in molar proportions. The structures were solved by direct methods and refined to R = 0.068 for 2585 observed reflections in the case of (I) and R = 0.036 for 2154 observed reflections in the case of (II). Two of the three crystallographically independent arginine molecules in the complexes have conformations different from those observed so far in the crystal structures containing arginine. The succinic acid molecules and the succinate ions in the structures are centrosymmetric and planar. The crystal structure of (II) is highly pseudosymmetric. Arginine-succinate interactions in both the complexes involve specific guanidyl-carboxylate interactions. The basic elements of aggregation in both the structures are ribbons made up of alternating arginine dimers and succinate ions. However, the ribbons pack in different ways in the two structures. (II) presents an interesting case in which two ionisation states of the same molecule coexist in a crystal. The two complexes provide a good example of the effect of change in chirality on stoichiometry, conformation, aggregation, and ionisation state in the solid state.

Amino Acids↗

Direct interaction between yeast NADH-ubiquinone oxidoreductase, succinate-ubiquinone oxidoreductase, and ubiquinol-cytochrome c oxidoreductase in the reduction of exogenous quinones.

The reduction of the following exogenous quinones by succinate and NADH was studied in mitochondria isolated from both wild type and ubiquinone (Q)-deficient strains of yeast: ubiquinone-0 (Q0), ubiquinone-1 (Q1), ubiquinone-2 (Q2), and its decyl analogue 2,3-dimethoxy-5-methyl-6-decyl-1,4-benzoquinone (DB), duroquinone (DQ), menadione (MQ), vitamin K1 (2-methyl-3-phytyl-1,4-naphthoquinone), the plastoquinone analogue 2,3,6-trimethyl-1,4-benzoquinone (PQOc1), plastoquinone-2 (PQ2), and its decyl analogue (2,3-dimethyl-6-decyl-1,4-benzoquinone). Reduction of the small quinones DQ, Q0, Q1, and PQOc1 by NADH occurred in both wild type and Q-deficient mitochondria in a reaction inhibited more than 50% by myxothiazol and less than 20% by antimycin. The reduction of these small quinones by succinate also occurred in wild type mitochondria in a reaction inhibited more than 50% by antimycin but did not occur in Q-deficient mitochondria suggesting that endogenous Q6 is involved in their reduction. In addition, the inhibitory effects of antimycin and myxothiazol, specific inhibitors of the cytochrome b-c1 complex, on the reduction of these small quinones suggest the involvement of this complex in the electron transfer reaction. By contrast, the reduction of Q2 and DB by succinate was insensitive to inhibitors and by NADH was 20-30% inhibited by myxothiazol suggesting that these analogues are directly reduced by the primary dehydrogenases. The dependence of the sensitivity to the inhibitors on the substrate used suggests that succinate-ubiquinone oxidoreductase interacts specifically with center i (the antimycin-sensitive site) and NADH ubiquinone oxidoreductase preferentially with center o (the myxothiazol-sensitive site) of the cytochrome b-c1 complex. The NADH dehydrogenase involved in the myxothiazol-sensitive quinone reduction faces the matrix side of the inner membrane suggesting that center o may be localized within the membrane at a similar depth as center i.

Electron Transport Complex II↗

A 1H NMR study of succinate synthesis from exogenous precursors in oxygen-deprived rat heart mitochondria.

Succinate synthesis from exogenous malate, alpha-ketoglutarate, oxaloacetate and L-glutamate in isolated oxygen-deprived rat heart mitochondria was studied using 1H NMR. The highest rate of succinate synthesis was observed during incubation of mitochondria with a mixture of L-glutamate and oxaloacetate. When mitochondria were incubated with [U-13C] glutamate and oxaloacetate the [U-13C] succinate/succinate and aspartate/succinate ratios were equal to 2. This suggests that the succinate produced from [U-13C] alpha-keto-glutarate formed via transamination of [U-13C] glutamate with oxaloacetate by aspartate aminotransferase exceeds twofold that synthesized via oxaloacetate reduction. It may thus be expected that GTP yield in a reaction catalyzed by the succinic thiokinase will be 2 times higher that of ATP production coupled with NADH-dependent fumarate reduction.

Anaerobiosis↗

[Succinate dehydrogenase activity of the retina and occipital lobe of the brain during emotional stress].

The emotional stress induces changes in redox processes in the visual analyzer at the level of the succinic acid transformation in the Krebs cycle. The stress evokes a particularly sharp decrease in the succinate dehydrogenase activity in the retina under conditions of dark adaptation whereas in the occipital lobes under the same condition a considerable intensification of the succinic acid oxidation is observed. Stress action under ordinary conditions of the day and night change is accompanied by the succinate dehydrogenase activation in the visual analyzer. However, the light load increases the enzyme activity in test animals the same way as in intact animals.

Animals↗

[Comparison between the succinate dehydrogenase inhibition test and ATP assay for in vitro chemosensitivity testing].

The predicted level of cell viability was compared between the succinate dehydrogenase inhibition (SDI) test and the adenosine triphosphate (ATP) assay, both of which are used for in vitro human tumor chemosensitivity testing. After HeLa cells had been exposed to various concentrations of 5-FU for 1, 2, 3 or 4 days, the decrease occurring in viable cell number correlated with that of succinate dehydrogenase (EC 1.3.99.1) activity and that of the intracellular ATP level of the viable cells. In dead cells, the ATP level was extensively decreased, but the succinate dehydrogenase activity remained at a level of 11% of that of 5-FU-untreated viable cells, even on day 4. The cell viability correlated well with the intracellular ATP level, as compared with the succinate dehydrogenase activity. The activity remaining in dead cells must thus be taken into consideration for the prediction of chemosensitivity in the SDI test, but not in the ATP assay.

Adenosine Triphosphate↗

The anaerobic heart: succinate formation and mechanical performance of cat papillary muscle.

Quiescent or paced, isometrically contracting right ventricular papillary muscles of cat heart were incubated under oxygenated or hypoxic (Po2 less than 10 Torr) conditions in order to investigate lactate and succinate formation. Both compounds were produced and released into the incubation medium during oxygen deficiency. Mechanical performance stimulated synthesis of both compounds under hypoxic as well as oxygenated conditions. Pacing led to the accumulation of large amounts of glutamine both under oxygenated and hypoxic conditions. Hypoxia resulted in a marked depletion of high-energy phosphates and concomitantly mechanical performance was impaired, i.e. developed tension fell rapidly and relaxation rate decreased. Supplying hypoxic, contracting muscles with aspartate (2 mM) resulted in maintenance of muscular function to some extent and led to augmented release of succinate and lactate. The data indicate that anaerobic succinate formation is correlated to the energy requiring processes of the myocardium. Maintenance of myocardial function by the supply of amino acids may be related to their conversion into succinate and to the stimulation of glycolysis.

Alanine↗