Succinyl Coenzyme Research Articles

Regulation of citrate synthase from blue-green bacteria by succinyl coenzyme A

Citrate synthase (EC 4.1.3.7) was prepared from nine species of blue-green bacteria. In every case the citrate synthase was of the “large” type otherwise found only in Gram-negative bacteria.

Nuclear magnetic resonance relaxation time studies on the manganese(II) ion complex with succinyl coenzyme A synthetase from Escherichia coli.

Nuclear magnetic resonance, as well as electron paramagnetic resonance, experiments were carried out in a study of the Mn(II) ion complex with phosphorylated succinyl-CoA synthetase. For high specific activity enzyme samples, there are 3.5 +/- 0.7 metal ion binding sites per enzyme molecule with indistinguishable dissociation constants (KD = 6.9 X 10(-4) M). However, for enzyme samples with a lower specific activity yet equivalent purity, there are 1.6 strong metal ion binding sites (KD = 6.6 X 10(-4) M) and 2.0 weak metal ion binding sites (KD = 4.0 X 10(-3) M), a result that is easily reconciled with the alpha2beta2 subunit structure of the enzyme. Water proton relaxation rate measurements indicate that each strongly bound Mn(II) ion is coordinated to three water molecules. The present results strongly suggest that the existence of nearly 4 high-affinity metal binding sites per enzyme molecule is related to the integrity or configuration of that portion of the molecule which interacts with substrates.

Letter: A nonenzymic model intermediate for the coenzyme B 12 dependent isomerization of methymalonyl coenzyme A to succinyl coenzyme A.

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTA nonenzymic model intermediate for the coenzyme B12 dependent isomerization of methylmalonyl coenzyme A to succinyl coenzyme APaul Dowd and Moritz ShapiroCite this: J. Am. Chem. Soc. 1976, 98, 12, 3724–3725Publication Date (Print):June 1, 1976Publication History Published online1 May 2002Published inissue 1 June 1976https://doi.org/10.1021/ja00428a065RIGHTS & PERMISSIONSArticle Views85Altmetric-Citations28LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (317 KB) Get e-Alerts Get e-Alerts

Congenital Methylmalonic Acidemia: a Variant of the B ‘Non-Responsive’ Form with Evidence for Reduced Affinity of Methylmalonyl-CoA Mutase for its B(12)-Coenzyme

Methylmalonate metabolism was investigated in fibroblasts and leukocytes of two unrelated patient with a B12-nonresponsive type of congenital methylmalonic acidemia. Intact fibroblasts from both patients showed a defective metabolism of methyl-14 c-malonate to 14CO2, whereas no such defect was found in their intact peripheral leukocytes. In disrupted fibroblasts, the conversion of methylmalonyl coenzyme A to succinyl coenzyme A was markedly reduced but was completely normalized by the addition of 5'-deoxyadenosylcobalamin (AdoCb1; 10(-5) mol/l), the specific coenzyme of methylmalonyl coenzyme A mutase. Assays with decreasing concentrations of AdoCbl (10(-5)-10(-11) mol/l) suggested a reduced affinity of the mutase apoenzyme for its coenzyme, implicating yet another variant of this heterogeneous disease.

Catalysis of a step of the overall reaction by the alpha subunit of Escherichia coli succinyl coenzyme A synthetase.

The isolated alpha subunit or succinyl-CoA synthetase from Escherichia coli is capable of catalyzing one step of the overall reaction, namely its own phosphorylation by the substrate ATP. The data presented herein also suggest that the binding sites for other substrates (succinate, succinyl-CoA) are located either on the beta subunit or comprise part of both subunit types. From these observations and from our earlier finding that the two subunits species are necessary for the overall reaction, we propose that the active site is assembled at or close to the point of contact of the two subunits in the native alpha2beta2 enzymic structure.

Succinyl coenzyme A and porphobilinogen formation in isolated etio-chloroplasts and greening laminae

A comparative study of succinyl CoA synthetase and δ-ALA dehydratase activities in greening Avena laminae and in isolated etio-chloroplasts showed the former enzyme activity to be non-plastidic in location during all stages of chloroplast morphogenesis and the latter to be mainly associated with the plastids. The maximum rate of formation of porphobilinogen in developing plastids was achieved after 24 hr of illumination.

Isolation of the alpha and beta subunits of Escherichia coli succinyl coenzyme A synthetase and their recombination into active enzyme.

Pure preparations of the alpha and beta subunits of succinyl-CoA synthetase of Escherichia coli have been obtained by preparative gel electrophoresis in the presence of detergent or by gel filtration in acid urea solution. Substantial enzyme activity (50 to 60%) can be recovered following renaturation of an equimolar mixture of the two subunits prepared by gel filtration. The substrate ATP is required for reconstitution of activity, but the presence of our substrates does not appear to influence the renaturation process. Knowing that ATP phosphorylates the alpha subunit in the native enzyme, these data suggest that phosphorylation of this subunit is necessary for correct assembly. Comparison of the amino acid compositions of the stoichiometry of the subunit reassociation in the reconstitution process confirm the alpha2 beta2 structure for the native enzyme.

Succinyl Coenzyme A Synthetase of Escherichia coli: Initial Rate Kinetics of Succinyl-CoA Cleavage and Isotope Exchange Studies

Initial rate kinetic studies of succinyl coenzyme A synthetase of E. coli in the direction of succinyl-CoA cleavage are consistent with the operation of a partially random sequential kinetic mechanism with initial binding of ADP followed by random association of succinyl-CoA and Pi. The mechanism is analogous to that proposed previously for the succinyl-CoA formation reaction, and thus the kinetic mechanism of the overall reversible succinyl-CoA synthetase reaction appears to be symmetrical.Studies of the kinetics of [Formula: see text] isotope exchange at equilibrium show that this partially random sequential kinetic mechanism is not an exclusive pathway. [Formula: see text] isotope exchange rates did not show complete substrate inhibition when CoA or succinate was varied in constant ratio with Pi. However, when CoA or succinate was varied in constant ratio with succinyl-CoA, nearly complete substrate inhibition was observed. These results can be interpreted in terms of a wide variety of minor pathways of substrate binding and product release available to the enzyme under various conditions.

Succinyl Coenzyme A Synthetase of Escherichia coli: EFFECTS OF PHOSPHOENZYME FORMATION AND OF SUBSTRATE BINDING ON THE REACTIVITY AND STABILITY OF THE ENZYME

Abstract Formation of phosphohistidine at only one of an apparent total of two active sites in Escherichia coli succinyl coenzyme A synthetase results in profound changes in the reactivity of the enzyme. On storage, the dephosphorylated form of enzyme loses activity at a rate approximately 100 times that of the monophosphorylated form. Similarly, there is a difference of two orders of magnitude in the rates of inactivation by trypsin of the phosphorylated and dephosphorylated enzymes. There is direct correlation between both the rate and extent of inactivation and the status of phosphorylation of the enzyme. In keeping with the ability of substrates to interconvert the phosphorylated and dephosphorylated forms of enzyme, ATP protects the previously dephosphorylated enzyme from proteolytic attack, whereas the presence of succinate and Mg++ renders previously phosphorylated enzyme susceptible to proteolysis. These effects are observed not only with trypsin, but with a variety of proteases having different specificities. The results indicate that phosphorylation of 1 active site histidine residue per oligomeric enzyme molecule results in a significant change in the three-dimensional structure of the enzyme to a tighter conformation, with concomitant stability and, perhaps, distortion of the second active site in the molecule so that its phosphorylation is unfavorable.

Succinyl coenzyme A synthetase of pig heart: studies of the subunit structure and of phosphoenzyme formation.

Succinyl coenzyme A synthetase from pig heart (molecular weight approximately 75 000) gives rise to two species of subunits when subjected to polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The molecular weights of the constituent subunits are estimated to be about 42 500 and 34 500, suggesting that the enzyme is a dimer. The presence of saturating concentrations of guanosine 5′-triphosphate (GTP) results in the formation of one phosphohistidine residue per molecule of enzyme; the site of phosphorylation is in the smaller subunit. The standard free energy change for phosphorylation of the enzyme by GTP is approximately −400 cal/mol. The results point to both interesting similarities and differences in succinyl-CoA synthetases isolated from mammalian and bacterial sources.

Regulation of pyruvate kinases by succinyl coenzyme A

Summary The fructose-1,6-diphosphate (FDP) activated pyruvate kinases from Escherichia coli , rat liver and yeast are inhibited by succinyl-CoA. In the case of the enzyme from E. coli , the inhibition by succinyl-CoA only occurs in the presence of the activator, FDP. Succinyl-CoA and ATP in combination show cooperative inhibition.

Succinyl coenzyme A synthetase of Escherichia coli. Sequence of a peptide containing the active-site phosphohistidine residue.

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTSuccinyl coenzyme A synthetase of Escherichia coli. Sequence of a peptide containing the active-site phosphohistidine residueT. Wang, L. Jurasek, and W. A. BridgerCite this: Biochemistry 1972, 11, 11, 2067–2070Publication Date (Print):May 1, 1972Publication History Published online1 May 2002Published inissue 1 May 1972https://doi.org/10.1021/bi00761a011RIGHTS & PERMISSIONSArticle Views48Altmetric-Citations25LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (468 KB) Get e-Alerts Get e-Alerts

Evidence for two types of subunits in succinyl coenzyme a synthetase

Two species of subunits are derived from Escherichia coli succinyl CoA synthetase by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Their molecular weights are approximately 38,500 and 29,500. The active-site phosphohistidine residue occurs in the smaller subunit. The data strongly suggest that the two subunit types are present in equimolar amounts in the native enzyme (molecular weight ca . 140,000).

Induction of Hepatic δ-Aminolevulinic Acid Synthetase Activity in Strains of Inbred Mice

Abstract δ-Aminolevulinic acid synthetase activity is increased in hepatic homogenates from mice that have been fasted or treated (or both) with 3,5-dicarbethoxy-1,4-dihydro-2,4,6-trimethylpyridine (DDC). Comparison of δ-aminolevulinic acid synthetase activity in livers from adult female mice of 15 inbred strains showed significant differences (p l 0.01) among the strains in the amount of enzyme activity normally present and in the amount present after induction. After DDC administration, δ-aminolevulinic acid synthetase activity increased at a linear rate for 12 hours and then reached a plateau at a level that was maintained until at least 20 hours after drug administration. Strain differences were maintained over a range of DDC doses from 28 mg per kg to 500 mg per kg (the highest dose tested). No significant differences in plasma DDC levels were noted between strains of and induced activity. Representative strains with high and low DDC-induced enzyme activity were compared in detail. Neither modification of the glycine, pyridoxal 5'-phosphate, or EDTA concentrations nor substitution of other buffers for the usual Tris buffer changed the relative enzyme activities of homogenates. Addition of succinyl coenzyme A or a succinyl CoA generating system increased total homogenate activity to the same extent in high and low strains. In animals from both classes, added succinate inhibited δ-aminolevulinate production. When hepatic homogenates of DDC-treated mice were fractionated by differential centrifugation, no substantial differences were noted among strains in the subcellular distribution of δ-aminolevulinic acid synthetase activity. Our data suggest that among inbred strains of mice there are either differences in the number of δ-aminolevulinic acid synthetase enzyme molecules present or differences in the catalytic activity of a structurally modified enzyme produced in response to fasting or DDC administration (or both) and that genetic factors are responsible for this heterogeneity of response.

Subunits, composition, and related properties of succinyl coenzyme A synthetase.

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTSubunits, composition, and related properties of succinyl coenzyme A synthetaseClaus Leitzmann, Jang-Yen Wu, and Paul D. BoyerCite this: Biochemistry 1970, 9, 11, 2338–2346Publication Date (Print):May 26, 1970Publication History Published online1 May 2002Published inissue 26 May 1970https://doi.org/10.1021/bi00813a018RIGHTS & PERMISSIONSArticle Views110Altmetric-Citations43LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (2 MB) Get e-Alerts Get e-Alerts

The Kinetics of Succinyl Coenzyme A Synthetase from Escherichia coli: A REACTION WITH A COVALENT ENZYME-SUBSTRATE INTERMEDIATE NOT EXHIBITING "PING-PONG" KINETICS

Succinyl coenzyme A synthetase from Escherichia coli does not exhibit a steady state kinetic pattern indicative of kinetics, despite the fact that catalytic participation of a phosphorylated enzyme covalent intermediate has been established. Instead, kinetic patterns are consistent with the sequential addition of all substrates, to form a quaternary complex, before the release of any product. Double reciprocal plots are intersecting with ATP and succinate varied at nonsaturating concentration of the third substrate, CoA. These plots become parallel at higher CoA concentrations, suggesting that CoA can be the second substrate to add in the binding sequence. Analogous reciprocal plots with ATP and CoA varied at both nonsaturating and saturating succinate concentrations are also intersecting and parallel, respectively. The data may be reconciled with a binding sequence in which ATP binds to the enzyme first, followed by the random addition of CoA and succinate. The finding of a sequential mechanism for the addition of substrates to this enzyme, whose catalysis is known to involve a covalent intermediate, serves to illustrate a possible danger in the use of initial rate kinetics. It is not possible to discount chemical mechanisms involving covalent intermediates by failure to observe the ping-pong kinetic pattern which may be anticipated for these catalytic routes.

Quantitative appraisals of possible catalytic intermediates in the succinyl coenzyme A synthetase reaction.

Quantitative appraisals of possible catalytic intermediates in the succinyl coenzyme A synthetase reaction.

Dephosphorylation of succinyl coenzyme A synthetase as related to enzyme specificity and catalytic intermediates

Dephosphorylation of succinyl coenzyme A synthetase as related to enzyme specificity and catalytic intermediates

Apparent Respiratory Control in Uncoupled Mitochondria

Abstract Respiration by coupled mitochondria from heart was enhanced by acetate, butyrate, and octanoate in a controlled manner, limited by exogenous ADP. When uncoupling agents were added, the ADP requirement for oxidation of pyruvate, α-ketoglutarate, succinate, or β-hydroxybutyrate was abolished. In contrast, respiration by uncoupled mitochondria incubated with acetate, butyrate, or octanoate remained dependent on the addition of ADP. The ratio of ADP to oxygen was approximately unity in uncoupled mitochondria incubated with short chain fatty acids. The controlled increase in respiration by mitochondria associated with ADP addition was abolished or inhibited by malonate or arsenite, indicating that operation of the tricarboxylic acid cycle was required for manifestation of the respiratory control in both coupled and uncoupled mitochondria. The data were interpreted to indicate that ATP required for fatty acid activation was generated from exogenous ADP indirectly via operation of the succinyl coenzyme A synthetase and the nucleoside diphosphokinase reactions. Addition of the nonmetabolized inhibitors of carnitine acetyltransferase ((+)-acetylcarnitine and dl-bromoacetylcarnitine) to bovine heart mitochondria elicited effects similar to those observed after the addition of acetate, butyrate, or octanoate. Either of these transferase inhibitors induced an increase in respiration, dependent on ADP, by both coupled and uncoupled mitochondria. The ADP to oxygen ratio in uncoupled mitochondria was approximately unity, and the effect was completely abolished by malonate or arsenite. The nature of the endogenous substrate whose oxidation was enhanced in the presence of ADP by the nonmetabolized inhibitors of carnitine acetyltransferase was not proven, but evidence was presented suggesting the importance of endogenous fatty acids in these processes.

Succinyl Phosphate and the Succinyl Coenzyme A Synthetase Reaction

Trilithium succinyl phosphate of a high degree of purity has been synthesized chemically. It is active in the synthesis of ATP from ADP in a reaction catalyzed by succinyl coenzyme A synthetase from Escherichia coli (succinate: CoA ligase (ADP); EC 6.2.1.5). With desulfo-CoA present, 1 unit of enzyme catalyzes the conversion of 1 µmole of succinyl phosphate into 0.5 µmole of ATP in 60 min at 37°. Kinetically, the rate of ATP synthesis from succinyl phosphate is one-fifth of that from succinyl-CoA plus inorganic phosphate. Omitting desulfo-CoA lowers the rate of reaction to one-quarter and the conversion to one-half. The reactivity of succinyl-CoA synthetase with 13 other acyl phosphates is reported. Succinyl-CoA is rapidly synthesized from succinyl phosphate and coenzyme A in a nonenzymatic reaction, the rate and extent of which depend on temperature, pH, and concentration of reactants.