ISSN 0031-3998 (print), Succinyl-CoA Ligase Deficiency: A Mitochondrial Hepatoencephalomyopathy, https://doi.org/10.1203/PDR.0b013e3181e5c3a4, SUCLG1 mutations and mitochondrial encephalomyopathy: a case study and review of the literature, Loss of succinyl-CoA synthase ADP-forming subunit disrupts mtDNA stability and mitochondrial dynamics in neurons, Mitochondrial dysfunction in liver failure requiring transplantation, Succinate-CoA ligase deficiency due to mutations in SUCLA2 and SUCLG1: phenotype and genotype correlations in 71 patients, Liver X receptors in lipid metabolism: opportunities for drug discovery. This is similar to previous patients. KINETIC STUDIES: INITIAL VELOCITY, PRODUCT INHIBITION, AND EFFECT OF ARSENATE. GDP appears to function by neutralizing the action of a specific inhibitory protein. In further refining this model for catalysis, we have shown that the monothiophosphorylated E. coli enzyme does not catalyse exchange of 18O from the beta, gamma-bridge to the beta-non-bridge position of ATP, indicating that the enzyme does not undergo even transient bis-phosphorylation. As a first step in studying the in vivo synthesis and assembly of the enzyme in the mammalian mitochondrial matrix, we have cloned and sequenced a 900 bp cDNA fragment that encodes most of the alpha subunit of rat liver succinyl-CoA synthetase. J Biol Chem. Before 1. a ligase reversibly reacting succinate and CoA with ATP to produce ADP, inorganic phosphate, and succinyl-CoA; The proteins were eluted by a step gradient with increasing sodium chloride concentrations at a flow rate of 4 ml/min, as follows: 0 to 20 min, 0 mM NaCl; 20 to 65 min, 50 mM NaCl; 65 to 110 min, 75 mM NaCl; 110 to 155 min, 100 mM NaCl; and 155 to 210 min, 150 mM NaCl. [10]. Eur J Biochem 241: 779786, Welch EM, Jacobson A 1999 An internal open reading frame triggers nonsense-mediated decay of the yeast SPT10 mRNA. In this strain, DTDP is metabolized via the intermediate product 3-sulfinopropionate (3SP) (29,31). Previous studies had suggested that SucCD enzymes might be promiscuous toward succinate analogues, such as itaconate and 3-sulfinopropionate (3SP). Its central importance to many biochemical pathways suggests that it was one of the earliest components of metabolism. ATP modulation of catalytic steps in the succinyl-CoA synthetase reaction. Plasmid pET-23a(+)::sucCDAboHis was generated by PCR-based mutagenesis using 5-phosphorylated oligonucleotides P_forward_XhoI_Histag_Abo and P_Abo_rev_mutagenesis and pET-23a(+)::sucCDAbo as the template. These findings are strongly suggestive for succinyl-CoA ligase deficiency when present in a patient with lactic acidosis. A detailed structural description of Escherichia coli succinyl-CoA synthetase, Crystallization of succinyl-CoA synthetase from Escherichia coli, Novel mechanisms of Escherichia coli succinyl-coenzyme A synthetase regulation. The proteins were eluted by a linear gradient of increasing sodium chloride concentrations at a flow rate of 3 ml/min, as follows: 0 to 40 min, 0 mM NaCl, and 40 to 240 min, 0 to 1 M NaCl (, 5 mmol/min). Substrate-level phosphorylation is a metabolism reaction that results in the production of ATP or GTP supported by the energy released from another high-energy bond that leads to phosphorylation of ADP or GDP to ATP or GTP (note that the reaction catalyzed by creatine kinase is not considered as "substrate-level phosphorylation"). Fatal infantile lactic acidosis: Defective SCS has been implicated as a cause of fatal infantile lactic acidosis, which is a disease in infants that is characterized by the build-up of toxic levels of lactic acid. Euglena gracilis cells synthesize the key tetrapyrrole precursor, -aminolevulinic acid (ALA), by two routes: plastid ALA is formed from glutamate via the transfer RNA-dependent five-carbon route, and ALA that serves as the precursor to mitochondrial hemes is formed by ALA synthase-catalyzed condensation of succinyl-coenzyme A and glycine. The essential role of sucCD in the activation of 3SP in vivo was proved by genetic complementation. A succinyl coenzyme A synthetase preparation from Escherichia coli has the following characteristics: apparent chromatographic, sedimentation, and electrophoretic homogeneity; molecular weight of about 141,000; A 280 (1 mg per ml) = 0.511; higher specific activity than previous preparations; and identity by several criteria with a phosphorylated protein (E-P) obtained upon exposure to Mg . Bethesda, MD 20894, Web Policies Department of Biochemistry and Molecular Biology, St. Louis University School of Medicine, Missouri 63104, USA. The .gov means its official. * denotes M14, which is mutated in the patient. PMC The following tuning parameters were retained for optimum detection of CoA-thioesters: capillary temperature, 300C; sheath gas flow, 12 liters/h; auxiliary gas flow, 6 liters/h; and sweep gas flow, 1 liter/h. Desalted extract of Euglena cells catalyzed ALA formation in a reaction that required coenzyme A and GTP but did not require exogenous succinyl-coenzyme A synthetase. 1986 Dec;82(4):1096-101 This may not be the complete list of references from this article. 1) could not be utilized by any of the investigated SucCD enzymes. Petrcek M, Petrckov K, Havlcek L, Felsberg J. J Bacteriol. [9], Johnson et al. ITMS + c ESI Full ms, ion trap mass spectrometry, positive mode with electrospray ionization (full MS spectrum recorded); cid, collision-induced dissociation (typically with an energy of 30.00 eV [electron volts]). eCollection 2015. The Km values for these substrates were in the same range as those for 3SP obtained in this study. [6][7] As can be seen in Image 2, the two subunits (pink and green) reside on opposite sides of the structure and the two subunits (yellow and blue) interact in the middle region of the protein. The standard in vitro activity of succinate-CoA ligase in the direction of ADP formation was assayed by a continuous spectrophotometric assay according to the method of Cha and Parks (50). After expression of sucCDAboHis in E. coli BL21(DE3)/pLysS, cell harvest, and cell disruption, the soluble fraction was applied to an Ni-nitrilotriacetic acid (NTA) Sepharose column (1 ml; HisTrap HP; GE Healthcare, Munich, Germany). The enzyme from Escherichia coli is an alpha 2 beta 2 tetramer (mol. Mitochondrion 9: 438442, Ostergaard E, Christensen E, Kristensen E, Mogensen B, Duno M, Shoubridge EA, Wibrand F 2007 Deficiency of the -subunit of succinate-coenzyme A ligase causes infantile lactic acidosis with mitochondrial DNA depletion. Succinyl-Coenzyme A Synthetase and its Role in delta - PubMed Succinyl-CoA synthetase structure-function relationships and other considerations. The SucC subunit coeluted from the column matrix. aInstitut fr Molekulare Mikrobiologie und Biotechnologie, Westflische Wilhelms-Universitt Mnster, Mnster, Germany, bEnvironmental Sciences Department, King Abdulaziz University, Jeddah, Saudi Arabia. 8600 Rockville Pike For elution purposes, a gradient of imidazole was applied. -, FEBS Lett. Careers, Unable to load your collection due to an error. Succinyl Coenzyme A - an overview | ScienceDirect Topics However, the SucCD activity obtained with the last three compounds was below 1% of that obtained with succinate; therefore, it is assumed that these activities do not have any relevance in vivo. Schlegel HG, Kaltwasser H, Gottschalk G. 1990 Nov;94(3):1365-75 -ketoglutarate is later converted to succinate. In conclusion, patients with succinyl-CoA ligase deficiency present with a progressive hepatoencephalomypathy. Fraser ME, James MN, Bridger WA, Wolodko WT. strain MA (ATCC 23819) and which is also a member of the group of microorganisms able to assimilate C1 compounds, was biochemically characterized in the past (53,56). Inclusion in an NLM database does not imply endorsement of, or agreement with, In this study, we purified three homologous SucCD enzymes and characterized these enzymes with regard to their ability to convert different carbon acid substrates as analogues of succinate to their corresponding CoA-thioesters in vitro. The NIST mass spectral search program, Windows software version 1.6d. Bioinformatic analysis using the MitoProt II 1.0a4 software (16) indicates that the M14L substitution does not significantly alter the probability of the amino terminus to function as a mitochondrial targeting sequence (0.3309 for M14 and 0.3366 for L14). Succinyl CoA synthetase (EC 6.2.1.5) stimulates the hydrolysis of succinyl CoA into succinate and ATP and thus represents a remarkable point of substrate level phosphorylation [2,18]. The https:// ensures that you are connecting to the the contents by NLM or the National Institutes of Health. Supplemental material for this article may be found at http://dx.doi.org/10.1128/AEM.03075-13. Degradation of nucleic acids is a catabolic reaction and the resulting parts of the nucleotides or nucleobases can be salvaged to recreate new nucleotides. The ability to convert both enantiomers of malate, with a slight preference for l-malate, was a general feature of the SucCD enzymes investigated in this study. Black error bars, standard deviations. The https:// ensures that you are connecting to the The reaction mechanism as determined by initial rate studies, Malate adenosine triphosphate lyase. Liver has primarily GDP-linked activity with the SUCLG2 encoded beta-subunit, and heart and kidney have fairly equal amounts of ADP and GDP-linked activities (24). mass. Fatal infantile lactic acidosis: Defective SCS has been implicated as a cause of fatal infantile lactic acidosis, which is a disease in infants that is characterized by the build-up of toxic levels of lactic acid. Shimizu S, Inoue K, Tani Y, Yamada H. Methods Cell Biol 80: 437463, Saada A 2004 Deoxyribonucleotides and disorders of mitochondrial DNA integrity. This patient had a normal birth weight and was described as normal for the first 3 months; then, developed failure to thrive, hypotonia and motor delay, progressive muscle atrophy and absent tendon reflexes, and severe hearing loss. 20C [11], SCS facilitates the flux of molecules into other metabolic pathways by controlling the interconversion between succinyl CoA and succinate. In this case, the vector-specific Shine-Dalgarno sequence was used. Although enzyme activities were very low for the additional substrates investigated, liquid chromatography/electrospray ionization-mass spectrometry analyses proved the ability of SucCD enzymes to form CoA-thioesters of adipate, glutarate, and fumarate. We identified the genes encoding these enzymes, and they were homologs of SCACT and SCS genes found in other bacteria. Nucleoside-diphosphate kinases are enzymes that catalyze the exchange of terminal phosphate between different nucleoside diphosphates (NDP) and triphosphates (NTP) in a reversible manner to produce nucleotide triphosphates. Learn About Succinyl Coa | Chegg.com The Preparation, Properties, and Reactions of Succinyl Coenzyme A Ramping was performed as follows: equilibration with 90% eluent A for 2 min before injection and 90 to 45% eluent A for 20 min, followed by holding for 2 min and then a return to 90% eluent A within 5 min after injection. 5). [9], Johnson et al. Unable to load your collection due to an error, Unable to load your delegates due to an error. The latter was used for homo- and heterologous expression of genes in E. coli under the control of the lac promoter and T7 promoter, respectively. 242, No. The LC/ESI-MS device used in this study was provided by funds of the DFG (Deutsche Forschungsgemeinschaft, grant no. 2017 Jan 24;56(3):534-542. doi: 10.1021/acs.biochem.6b00098. 1961. As mentioned, the enzyme facilitates coupling of the conversion of succinyl CoA to succinate with the formation of NTP from NDP and Pi. studies) suggest that co-operative interactions between alternatingly functional active sites on the two halves of the E. coli enzyme contribute to its catalytic efficacy. contributed equally to this study. Disclaimer. An Ultracel regenerated cellulose membrane with a nominal molecular mass limit of 10 kDa was used for protein concentration and buffer exchange to the binding conditions for chromatography. A submersion method for culture of hydrogen-oxidizing bacteria: growth physiological studies. Euglena gracilis cells synthesize the key tetrapyrrole precursor, delta-aminolevulinic acid (ALA), by two routes: plastid ALA is formed from glutamate via the transfer RNA-dependent five-carbon route, and ALA that serves as the precursor to mitochondrial hemes is formed by ALA synthase-catalyzed condensation of succinyl-coenzyme A and glycine. The stimulatory effect of ADP is not altered by sample dilution, is retained upon partial purification of the enzyme, and reflects the binding of ADP to a site other than the catalytic site. The genes encoding SucCDAboHis were also expressed in one bicistronic operon. The latter is an intermediate of the degradation pathway of 3,3-dithiodipropionate (DTDP), a precursor for the biotechnical production of polythioesters (PTEs) in bacteria. Bridger WA, Wolodko WT, Henning W, Upton C, Majumdar R, Williams SP. -ketoglutarate dehydrogenase enzyme helps in the synthesis of -ketoglutarate by the process of decarboxylation. Succinyl-CoA Ligase Deficiency: A Mitochondrial - Nature Succinyl coenzyme A synthetase - Wikipedia 1984;26(1):1-17. doi: 10.1002/jcb.240260102. Succinyl coenzyme A synthetase - WikiMili Succinyl coenzyme A synthetase (SCS, also known as succinyl-CoA synthetase or succinate thiokinase or succinate-CoA ligase) is an enzyme that catalyzes the reversible reaction of succinyl-CoA to succinate. However, this grasp domain is located about 35 away from the phosphorylated histidine residue. Succinyl coenzyme A synthetase (SCS, also known as succinyl-CoA synthetase or succinate thiokinase or succinate-CoA ligase) is an enzyme that catalyzes the reversible reaction of succinyl-CoA to succinate. J Mol Biol. official website and that any information you provide is encrypted In enzymology, a succinateCoA ligase (GDP-forming) ( EC 6.2.1.4) is an enzyme that catalyzes the chemical reaction. The .gov means its official. After amplification, a ligation reaction was performed in the buffer used for PCR, and the sample was used for transformation of CaCl2-competent cells of E. coli Top10. Chronic liver disease has not yet been reported in this condition. [16] It has been demonstrated that the gene for SCS (sucCD) is transcribed along with the gene for -ketoglutarate dehydrogenase (sucAB) under the control of a promoter called sdhC, which is part of the succinate dehydrogenase operon. 5), it appeared that the affected methionine is more likely to serve as the translation initiator methionine. CoA is removed from succinyl-CoA, producing free succinate; this is coupled with substrate-level . Substrate synergism and phosphoenzyme formation in catalysis by succinyl coenzyme A synthetase. Identification and characterisation of the alpha and beta subunits of succinyl CoA ligase of tomato. Succinyl-CoA synthetase (SCS; succinyl-CoA ligase; succinate thiokinase; EC 6.2.1.5), also known as succinyl-CoA synthetase ligase/succinate thiokinase, is a mitochondrial enzyme. This is true when either ATP or succinyl-CoA and Pi are used to phosphorylate the enzyme. Succinyl coenzyme A synthetase from Escherichia coli. (26). Cloning, characterization, and expression of the beta subunit of pig heart succinyl-CoA synthetase. Kinetic studies: initial velocity, product inhibition, and effect of arsenate. Enriched SucCDAm in the Q-Sepharose eluate was further purified to electrophoretic homogeneity by DEAE-Sepharose anion-exchange chromatography and by a final polishing step using modified EAH-Sepharose 4B chromatography. HHS Vulnerability Disclosure, Help Strains of E. coli were cultivated in lysogeny broth (LB) medium (36) or ZYP-5052 complex medium for autoinduction according to the method of Studier (37). This patient's biochemical phenotype was more striking and included strongly elevated glutamate and alpha-ketoglutarate, in addition to the small but persistent elevations of methylmalonate and propionylcarnitine. The matrix was equilibrated with 50 mM Tris-HCl (pH 7.4) and 0 mM NaCl at a flow rate of 4 ml/min. SucCDBl21 eluted after 101 to 141 min according to the purification protocol. Even though it is branded as a 'cycle', it is not necessary for metabolites to follow only one specific route; at least three alternative segments of the citric acid cycle have been recognized. Accessibility Previous studies have documented decreased mtDNA in muscle in patients with mutations in SUCLA2 (79) and in SUCLG1 (11,12). Succinyl coenzyme A-stimulated nucleoside diphosphate kinase activity of the enzyme, Fatty acids bound to unilamellar lipid vesicles as substrates for microsomal acyl-CoA ligase, AMP-forming acetyl-CoA synthetase from the extremely halophilic archaeon Haloarcula marismortui: purification, identification and expression of the encoding gene, and phylogenetic affiliation, Purification and characterization of two reversible and ADP-dependent acetyl coenzyme A synthetases from the hyperthermophilic archaeon Pyrococcus furiosus, Propionyl-coenzyme A synthetases of Ralstonia solanacearum and Salmonella choleraesuis display atypical kinetics. In the liver of a SUCLG1 patient, mtDNA was reduced to 65% of normal (12) and in our patient to 50% of normal. Other processes regenerate ATP. CHEMISTRY Vol. [16] It has been demonstrated that the gene for SCS (sucCD) is transcribed along with the gene for -ketoglutarate dehydrogenase (sucAB) under the control of a promoter called sdhC, which is part of the succinate dehydrogenase operon. A novel ADP-forming succinyl-CoA synthetase in Thermococcus kodakaraensis structurally related to the archaeal nucleoside diphosphate-forming acetyl-CoA synthetases, The metabolism of itaconic acid by liver mitochondria, The utilization of aconate and itaconate by Micrococcus sp, Studies on itaconate metabolism, I. Itaconyl-CoA synthesizing reaction in cell-free extract, Novel precursor substrates for polythioesters (PTE) and limits of PTE biosynthesis in Ralstonia eutropha. From: Plant Physiology and Biochemistry, 2021 View all Topics Add to Mendeley About this page Mitochondrial Disorders Later, this reaction was proved for the SucCD from Micrococcus sp. According to Shikata et al., activity levels for different substrates were obtained and normalized to the activity with the substrate succinate at a final concentration of 10 mM (21) (Fig. Accessibility In addition to that, the authors observed the formation of 3SP-CoA by a crude extract of the expression strain Escherichia coli BL21(DE3)/pLysS not harboring genes for SucCDAm (26). 1972. The synthesis and purity of the substance were confirmed by gas chromatography (GC) and GC/mass spectrometry (MS). In the mitochondrion, the matrix is the space within the inner membrane. DTDP of high-purity grade was purchased from Sigma-Aldrich (Steinheim, Germany). An interesting study using pigeon cells showed that GTP specific SCSs were located in pigeon liver cells, and ATP specific SCSs were located in the pigeon breast muscle cells. These results complete the findings of Schrmann et al., who described the essential role of SucCD in A. mimigardefordensis DPN7T in the degradation of DTDP (26). [10], SCS is the only enzyme in the citric acid cycle that catalyzes a reaction in which a nucleotide triphosphate (GTP or ATP) is formed by substrate-level phosphorylation. With regard to chain length, succinate (C4) was found to be the best substrate, whereas CoA-thioesters of glutarate (C5) and adipate (C6) were formed in only trace amounts; malonate (C3) was not activated at all. Pediatr Res 68, 159164 (2010). This child's phenotype was milder and similar to that of patients with a defect in the SUCLA2 gene. Novel mechanisms of Escherichia coli succinyl-coenzyme A synthetase Some of these biosynthetic pathways are located within a single cellular organelle, while others involve enzymes that are located within multiple cellular organelles. Examples of these biosynthetic pathways include the production of lipid membrane components and nucleotides. [11] Kinetics analysis of ATPSCS from the breast muscle of pigeons and GTPSCS from pigeon liver showed that their apparent Michaelis constants were similar for CoA, but different for the nucleotides, phosphate, and succinate. The crystal structure of Succinyl-CoA synthetase alpha subunit (succinyl-CoA-binding isoform) was determined by Joyce et al. SuccinateCoA ligase (GDP-forming) - Wikipedia In mammals, the enzyme is a heterodimer of an - and a -subunit. PCR products were isolated from agarose gels using a peqGOLD gel extraction kit (Peqlab Biotechnologie GmbH, Erlangen, Germany), digested with the appropriate restriction enzymes provided in the primer name, and ligated with digested pET-23a(+) (Novagen, Madison, WI) or pBluescriptSK() (Stratagene, San Diego, CA), yielding pBluescriptSK()::sucCDAm, pBluescriptSK()::sucCDBL21, and pET-23a(+)::sucCDAbo. Hence, the vector pET-23a(+)::sucCDAboHis, encoding a C-terminal hexahistidine tag on the SucD subunit, was generated (Table 1). [7] This histidine residue becomes phosphorylated during the succinate forming step in the reaction mechanism. Comparably high enzyme activity was determined for itaconate, l-malate, and d-malate (Fig. Kinetic parameters determined for SucCDBL21, SucCDAm, and SucCDAboHisa. The biosynthetic source of succinyl-coenzyme A in Euglena is of interest because this species has been reported not to contain -ketoglutarate dehydrogenase and not to use succinyl-coenzyme A as a tricarboxylic acid cycle intermediate. Another fate of succinyl-CoA is porphyrin synthesis, where succinyl-CoA and glycine are combined by ALA synthase to form -aminolevulinic acid (dALA). Can J Biochem Cell Biol. Novel reaction of succinyl coenzyme A (Succinyl-CoA) synthetase Consequently, translation initiation may occur at the next downstream AUG (i.e. Succinyl-coenzyme A synthetase activity was detected in extracts of dark- and light-grown wild-type and nongreening mutant cells. Association of guanine nucleotide-linked succinate thiokinase with ketone body utilization. The temperatures of the injector and detector were 250 and 240C, respectively. Succinyl-coenzyme A, abbreviated as succinyl-CoA or SucCoA, is a thioester of succinic acid and coenzyme A. Association of guanine nucleotide-linked succinate thiokinase with ketone body utilization", "Deficiency of the alpha subunit of succinate-coenzyme A ligase causes fatal infantile lactic acidosis with mitochondrial DNA depletion", https://en.wikipedia.org/w/index.php?title=Succinyl_coenzyme_A_synthetase&oldid=1159137411, This page was last edited on 8 June 2023, at 13:59. Four new derivatives of the broad-host-range cloning vector pBBR1MCS, carrying different antibiotic-resistance cassettes, Colloidal dispersion of chloroplast material, Succinyl coenzyme A synthetase from Escherichia coli. 1993 Aug;2(8):1255-62. doi: 10.1002/pro.5560020808. A protein solution enriched SucCDBL21 after Q-Sepharose chromatography (see Purification of homo- and heterologously expressed sucCD genes in native state above) was applied to a Cibacron Blue F3GA column (50 ml; GE Healthcare, Munich, Germany) equilibrated to the binding conditions (50 mM Tris-HCl [pH 7.4], 0 mM NaCl) at a flow rate of 3 ml/min. Kinetic parameters for all three SucCD enzymes indicate a preference for the physiological substrates CoA, ATP, and succinate; therefore, this clearly allocated this enzyme to the citric acid cycle. 2007. Liu M, Han X, Xian M, Ding Y, Liu H, Zhao G. Bioengineered. The enzyme consists of two different subunits forming a heterodimer or a heterotetramer structure (3, 4). Succinyl Coenzyme A Synthetase also known as succinyl coenzyme A synthetase, or succinate thiokinase) is an essential enzyme in tricarboxylic acid cycle catalysing the reversible conversion of succinyl-CoA to succinate.