Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.3.3.1 (citrate synthase)
4,488 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have studied the growth on acetate, the metabolism of acetate enzymes, and respiration of a series of citrate synthase mutants of Saccharomyces cerevisiae. The results confirmed and extended our previous observation that cytosolic citrate synthase is not necessary for growth on acetate. Deletion of mitochondrial citrate synthase (CS1) protein resulted in changes in metabolites, decrease in the amounts of pyruvate and alpha-ketoglutarate dehydrogenase complexes, reduced mitochondrial respiration of citrate and isocitrate, and an inability to grow on acetate. Using site-directed mutagensis, we constructed two separate CS1 proteins with mutations in the enzyme's active site. The mitochondria of cells carrying either site-directed mutagenized CS1 contained the inactive citrate synthase protein. With one mutant in which His313 was replaced with a glycine (CS1/H313G), growth on acetate was restored, and mitochondrial respiration of citrate and isocitrate increased toward parental levels as did the levels of several enzymes. With the other mutant CS1 in which Asp414 was replaced with a glycine (CS1/D414G), no growth on acetate or changes in other parameters was observed. We propose that the characteristics of the strain carrying the CS1 with a H313G mutation result from the formation of an intact Krebs cycle complex by the inactive but structurally unchanged H313G protein.
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PMID:Metabolic studies on citrate synthase mutants of yeast. A change in phenotype following transformation with an inactive enzyme. 266 55

Binding experiments indicate that mitochondrial aspartate aminotransferase can associate with the alpha-ketoglutarate dehydrogenase complex and that mitochondrial malate dehydrogenase can associate with this binary complex to form a ternary complex. Formation of this ternary complex enables low levels of the alpha-ketoglutarate dehydrogenase complex, in the presence of the aminotransferase, to reverse inhibition of malate oxidation by glutamate. Thus, glutamate can react with the aminotransferase in this complex without glutamate inhibiting production of oxalacetate by the malate dehydrogenase in the complex. The conversion of glutamate to alpha-ketoglutarate could also be facilitated because in the trienzyme complex, oxalacetate might be directly transferred from malate dehydrogenase to the aminotransferase. In addition, association of malate dehydrogenase with these other two enzymes enhances malate dehydrogenase activity due to a marked decrease in the Km of malate. The potential ability of the aminotransferase to transfer directly alpha-ketoglutarate to the alpha-ketoglutarate dehydrogenase complex in this multienzyme system plus the ability of succinyl-CoA, a product of this transfer, to inhibit citrate synthase could play a role in preventing alpha-ketoglutarate and citrate from accumulating in high levels. This would maintain the catalytic activity of the multienzyme system because alpha-ketoglutarate and citrate allosterically inhibit malate dehydrogenase and dissociate this enzyme from the multienzyme system. In addition, citrate also competitively inhibits fumarase. Consequently, when the levels of alpha-ketoglutarate and citrate are high and the multienzyme system is not required to convert glutamate to alpha-ketoglutarate, it is inactive. However, control by citrate would be expected to be absent in rapidly dividing tumors which characteristically have low mitochondrial levels of citrate.
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PMID:Regulation of malate dehydrogenase activity by glutamate, citrate, alpha-ketoglutarate, and multienzyme interaction. 289 80

Interaction between the alpha-ketoglutarate dehydrogenase complex and NAD+-dependent isocitrate dehydrogenase was detected with a variety of techniques including polyethylene glycol precipitation, ultracentrifugation, and centrifugal gel filtration on a Sepharose 6B column. The interaction was specific in that citrate synthase, cytosolic malate dehydrogenase, and NADP-dependent isocitrate dehydrogenase did not interact with alpha-ketoglutarate dehydrogenase complex. The interaction was not inhibited by either 0.1 M KCl or 0.4 M (NH4)2SO4, but was completely prevented by 5% glycerol. A new method for the preparation of NADH: ubiquinone oxidoreductase resulted in an enzyme having a protein subunit composition similar to that of classical complex I preparation. Evidence is given for the existence of ternary complexes containing NADH:ubiquinone oxidoreductase-alpha-ketoglutarate dehydrogenase complex-NAD-dependent isocitrate dehydrogenase and NADH: ubiquinone oxidoreductase-alpha-ketoglutarate dehydrogenase complex-succinate thiokinase. These data suggest that a part of the citric acid cycle may be located in the vicinity of NADH: ubiquinone oxidoreductase. These complexes may facilitate the transport of metabolites among these enzymes without their equilibrating with the whole compartment.
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PMID:Interaction between NAD-dependent isocitrate dehydrogenase, alpha-ketoglutarate dehydrogenase complex, and NADH:ubiquinone oxidoreductase. 311 Jan 60

The yeast, Saccharomyces cerevisiae, contains two citrate synthase isoenzymes, mitochondrial (CS1) and cytosolic (CS2). In this study, we have examined the metabolic consequences of the absence of CS1, CS2, and both isoenzymes in the respective mutant strains CS1-, CS2-, and CS1-CS2-. No significant differences were found in the growth rates of the parental, CS1-, or CS2- strains when grown in the single carbon sources galactose, glycerol, lactate, pyruvate, or glutamate. However, in nonfermentable carbon sources, the lag period in growth of CS1- was approximately 4 times that of the parental strain and the CS2- mutant. This difference was found even in glutamate. The CS1- mutant failed to grow on acetate in either complete or minimal liquid medium. Total cellular citrate concentration in the CS1- compared to the parental strain was higher when the cells were grown in lactate or pyruvate. On these same substrates, the malate concentration was 2-fold higher in the CS1-mutant when compared to the parental or CS2- strains. The production of 14CO2 by CS1- from [1-14C]acetate was 36% and that from [2-14C]acetate was 9.2% of the amount from the parental or CS2- strains. The 14CO2 production from [1-14C]glutamate was 28% and 20% in CS1- and CS1-CS2-, respectively, compared to the parental strain. Since these results are not easily explained solely by the absence of mitochondrial citrate synthase enzyme, we also determined the activity of some other enzymes of the citric acid cycle and electron transport chain. We found decreased activity of pyruvate dehydrogenase complex, alpha-ketoglutarate dehydrogenase complex, and aconitase, while the rest of the citric acid cycle enzymes and oxidative enzymes did not change significantly. The same changes in enzyme activities were found in two different yeast strains carrying the same citrate synthase mutations.
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PMID:Metabolic changes in Saccharomyces cerevisiae strains lacking citrate synthases. 313 54

Crude extracts of both vegetative cells and glycerol-induced microcysts of Myxococcus xanthus contained the following enzyme activities: phosphofructokinase, phosphoglucoisomerase, fructose-1,6-diphosphatase, fructosediphosphate aldolase, glyceraldehyde-3-phosphate dehydrogenase, phosphopyruvate carboxylase, citrate synthase, isocitrate dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, malate dehydrogenase, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, phosphoglucomutase, and uridine diphosphate glucose pyrophosphorylase. With the exception of isocitrate dehydrogenase, which was present at a fivefold higher concentration in microcysts, all activities in extracts from both types of cells were essentially equal. Hexokinase and pyruvate kinase could not be detected in extracts from either type of cell. Microcysts metabolized acetate at a lower rate than did vegetative cells. Most of this decrease was reflected in a substantial decrease in ability of microcysts to oxidize acetate to CO(2). In addition, microcysts and vegetative cells showed a different distribution of (14)C-label from incorporated acetate.
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PMID:Comparative intermediary metabolism of vegetative cells and microcysts of Myxococcus xanthus. 430 96

A technique was developed for the detection, on agar, of mutants of Bacillus subtilis that lacked a functional tricarboxylic acid cycle. Mutants devoid of detectable levels of aconitase, isocitric dehydrogenase, alpha-ketoglutarate dehydrogenase, succinic dehydrogenase, fumarase, and malate dehydrogenase have been isolated and characterized. Several mutants with conditionally expressible lesions, including a mutant with a heat-sensitive citrate synthase, have also been isolated. All of the mutants examined express all the biochemical markers normally absent in early-stage sporulation mutants except elastase, and some of these mutants sporulated nearly as well as the prototroph.
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PMID:Isolation and characterization of tricarboxylic acid cycle mutants of Bacillus subtilis. 499 41

During exponential growth, ordinary colorless (OC) plants of Blastocladiella emersonii consumed little glucose and produced no lactic acid. Similarly, resistant sporangial (RS) plants did not utilize glucose or produce lactic acid during the first 24 hr of exponential growth. During the next 24 hr of RS development, glucose was consumed with the concomitant production of lactic acid which was then reutilized. Lactic acid gradually accumulated again at maturity. Enzyme studies on cell-free extracts indicated the presence of all tricarboxylic cycle enzymes except alpha-ketoglutarate dehydrogenase at all stages of development of both RS and OC plants. Included among the enzymes detected were an adenosine monophosphate-stimulated, nicotinamide adenine dinucleotide-isocitric dehydrogenase, and citrate-condensing enzyme. When measured on a per plant basis, tricarboxylic cycle enzyme levels increased during the exponential growth of both kinds of plants. Only after the bicarbonate ceased to have effect on RS plant morphogenesis was there a decrease in the levels of the tricarboxylic cycle enzymes when measured on a per plant basis. Specific activity measurements indicated some differences in the differential rates of synthesis among the enzymes studied previous to 36 hr. Preliminary studies utilizing short periods of (14)C-bicarbonate fixation in young RS plants indicated that during the first 4 min most of the label was located in aspartic acid. These results are discussed in terms of previous results and particularly Cantino's hypothesis concerning the relationship between bicarbonate induction and tricarboxylic-cycle enzymes in the morphogenesis of B. emersonii.
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PMID:Tricarboxylic acid cycle enzymes and morphogenesis in Blastocladiella emersonii. 580 5

Burton, Sheril D. (Institute of Marine Science, University of Alaska, College), Richard Y. Morita, and Wayne Miller. Utilization of acetate by Beggiatoa. J. Bacteriol. 91:1192-1200. 1966.-A proposed system which would permit acetate incorporation into four-carbon compounds without the presence of key enzymes of the citric acid cycle or glyoxylate cycle is described. In this system, acetyl-coenzyme A (CoA) is condensed with glyoxylate to form malate, which, in turn, is converted to oxaloacetate. Oxaloacetate then reacts with glutamate to produce alpha-ketoglutarate, which is subsequently converted to isocitrate. Cleavage of isocitrate produces glyoxylate and succinate. Thus, the proposed system is similar to the glyoxylate bypass in that malate is produced from glyoxylate and acetyl-CoA, but differs from both the citric acid cycle and the glyoxylate bypass, since citrate and fumarate are not involved. Fumarase, aconitase, catalase, citritase, pyruvate kinase, enolase, phosphoenolpyruvate carboxylase, lactic dehydrogenase, alpha-ketoglutarate dehydrogenase, and condensing enzyme were not detectable in crude extracts of Beggiatoa. Succinate was oxidized by a soluble enzyme not associated with an electron-transport particle. Isocitrate was identified as the sole compound labeled when C(14)O(2) was added to a reduced nicotinamide adenine dinucleotide, CO(2) generating system (crystalline glucose-6-phosphate dehydrogenase and glucose-6-phosphate) in the presence of alpha-ketoglutarate.
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PMID:Utilization of acetate by Beggiatoa. 592 51

Acetate oxidation by sulphate was studied with desulfobacter postgatei. Cell extracts of the organism were found to contain high activities of the following enzymes: citrate synthase, aconitase, isocitrate dehydrogenase, alpha-ketoglutarate dehydrogenase, succinate dehydrogenase, fumarase, malate dehydrogenase and pyruvate synthase. It is concluded that acetate oxidation with sulphate in D. postgatei proceeds via the citric acid cycle with the synthesis of pyruvate from acetyl CoA and CO2 as an anaplerotic reaction. The apparent Ks for acetate oxidation by D. postgatei as determined in vivo was near 0.2 mM. The apparent Ks for acetate fermentation to methane and CO2 by methanosarcina barkeri was 3 mM. The significantly lower ks for acetate of the sulphate reducer explains why methane formation from acetate in natural habitats is apparently inhibited by sulphate.
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PMID:Dissimilatory sulphate reduction with acetate as electron donor. 612 36

The alpha-ketoglutarate dehydrogenase complex of either pig heart or Escherichia coli catalyzes a NAD- and CoASH-dependent oxidation of 2-keto-4-hydroxyglutarate which is stereoselective toward the L-isomer of this hydroxyketo acid. L-Malyl-CoA is the product of the reaction; the evidence includes observing (a) a steady increase in absorbance at 230 nm during the oxidation of 2-keto-4-hydroxyglutarate, (b) a positive response of oxidation reaction mixtures to neutral hydroxylamine, (c) loss of the two foregoing results concomitant with release of thiol-reacting material and the formation of free malate when reaction mixtures are heated, (d) formation of a hydroxamate which has chromatographic mobilities identical to that of chemically synthesized malate hydroxamate, (e) enzymatic formation of a radioactive product from 14C-labeled 2-keto-4-hydroxyglutarate which co-migrates with chemically synthesized malyl-CoA, and (f) hydrolysis of the product by citrate synthase, an enzyme absolutely specific for citryl-CoA and L-malyl-CoA. A 1:1:1 stoichiometric relationship exists between the amount of 2-keto-4-hydroxyglutarate oxidized, NAD reduced, and malate (or malyl-CoA) formed. Results from studies in which either 14C-labeled 2-keto-4-hydroxyglutarate, pyruvate, or glyoxylate is incubated with mixtures of purified enzymes or extracts of E. coli support the suggestion that the aldolase which preferentially catalyzes formation of L-2-keto-4-hydroxyglutarate from pyruvate plus glyoxylate in E. coli is coupled with the oxidative decarboxylation of this substrate, as reported here, and other enzymes in a multistep pyruvate-catalyzed cyclic oxidation of glyoxylate.
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PMID:Malyl-CoA formation in the NAD-, CoASH-, and alpha-ketoglutarate dehydrogenase-dependent oxidation of 2-keto-4-hydroxyglutarate. Possible coupled role of this reaction with 2-keto-4-hydroxyglutarate aldolase activity in a pyruvate-catalyzed cyclic oxidation of glyoxylate. 638 79


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