Succinate thiokinase

Succinyl-CoA is converted to succinate by the enzyme succinate thiokinase (succinyl-CoA synthetase). This is the only example in the citric acid cycle of substrate-level phosphorylation. Tissues in which glu-coneogenesis occurs (the hver and kidney) contain two isoenzymes of succinate thiokinase, one specific for GDP and the other for ADP. The GTP formed is used for the decarboxylation of oxaloacetate to phos-phoenolpymvate in gluconeogenesis and provides a regulatory hnk between citric acid cycle activity and the withdrawal of oxaloacetate for gluconeogenesis. Nongluconeogenic tissues have only the isoenzyme that uses ADP. 

As a result of oxidations catalyzed by the dehydrogenases of the citric acid cycle, three molecules of NADH and one of FADHj are produced for each molecule of acetyl-CoA catabohzed in one mrn of the cycle. These reducing equivalents are transferred to the respiratory chain (Figure 16-2), where reoxidation of each NADH results in formation of 3 ATP and reoxidation of FADHj in formation of 2 ATP. In addition, 1 ATP (or GTP) is formed by substrate-level phosphorylation catalyzed by succinate thiokinase. 

Citric acid cycle Succinate thiokinase Phosphorylation at substrate level 2... 

Succinyl CoA synthetase (succinate thiokinase) catalyzes a substrate-level phosphorylation of GDP to GTP. 

The subsequent cleavage of the thio-ester succinylCoA into succinate and coenzyme A by succinic acid-CoA ligase (succinyl CoA synthetase, succinic thiokinase) is strongly exergonic and is used to synthesize a phosphoric acid anhydride bond ( substrate level phosphorylation , see p. 124). However, it is not ATP that is produced here as is otherwise usually the case, but instead guanosine triphosphate (CTP). However, GTP can be converted into ATP by a nucleoside diphosphate kinase (not shown). 

Succinyl-CoA synthetase (SCS), also known as succinate thiokinase (STK) or succinate CoA ligase ( 6.2.1.4-5), is so far the only known hydrogenosomal enzyme directly involved in energy conservation. The protein catalyzes the reversible, substrate-level phosphorylation of ADP or GDP to the respective triphosphate at the expense of the high-energy thioester bond of succinyl-CoA. Succinate and CoA are released in the reaction. The I vaginalis enzyme... 

Jenkins TM, Gorrell , Muller M, Weitzman PD (1991) Hydrogenosomal succinate thiokinase in Tritrichomonas foetus and Trichomonas vaginalis. Biochem Biophys Res Commun 179 892-896... 

Fig. 8 SDS-PAGE (a) and Western blot (b) analysis of the purified hydrogenosomal fractions isolated from the metronidazole-susceptible T. vaginalis strain TV 10-02 (P) and its metronidazole-resistant derivatives MR-3, MR-5, MR-30, MR-50, and MR-100 displaying the aerobic (3), early anaerobic (5), advanced anaerobic (30, 50), and fully developed anaerobic resistance (100) to metronidazole. Numbers in the designation of MR strains indicate the concentrations of metronidazole in ixg/ml at which the organisms multiply in culture. About 10 pg protein was loaded per line. PFOR pyruvate ferredoxin oxidoreduc-tase, a-STK a subunit of succinate thiokinase (hydrogenosomal enzyme not involved in metronidazole resistance used as control), Fdx ferredoxin. From Rasoloson et al. (2002) by courtesy of the Society of General Microbiology...

Fig. 1 Enzymes localized to B. hominis mitochondrial-like organelle. The enzymes are 1 malic enzyme, 2 pyruvate NADP oxidoreductase, 3 acetate succinate CoA transferase, 4 succinate thiokinase, 5 a-ketoglutarate dehydrogenase, 6 isocitrate dehydrogenase, and 7 aconitase...

Succinate thiokinase (also called succinyl CoA synthetase) cleaves the high-energy thioester bond of succinyl CoA (see Figure 9.6). This reaction is coupled to phosphorylation of GDP to GTP. GTP and ATP are energetically interconvertible by the nucleoside diphos phate kinase reaction ... 

The generation of GTP by succinate thiokinase is another example of substrate-level phosphorylation (see p. 100). [Note Succinyl CoA is also produced from propionyl CoA derived from the metabolism of fatty acids with an odd number of carbon atoms (see p. 191), and from metabolism of several amino acids (see p. 264). 

Succinyl CoA is cleaved by succinate thiokinase (also called succinyl CoA synthetase), producing succinate and ATP (or GTP). This is an example of substrate-level phosphory lation. Succinate is oxidized to fumarate by succinate dehydrogenase, producing FADH2. The enzyme is inhibited by oxaloacetate. Fumarate is hydrated to malate by fumarase (fumarate hydratase), and malate is oxidized to oxaloacetate by malate dehy drogenase, producing NADH. 

Fig. 5.22. Oxidation of acetyl-CoA via the tricarboxylic acid (TCA) cycle. Individual enzymes of the pathway are marked. 1, citrate synthase 2 and 3, cis-aconitate hydratase 4 and 3, isocitrate dehydrogenase 6, a-oxo glutarate dehydrogenas 7, succinate thiokinase 8, succinate...

Succinate Thiokinase Couples the Conversion of Succinyl-CoA to Succinate with the Synthesis of GTP... 

The reaction is complex and involves an intermediate in which a phosphate is attached to a histidine residue of the succinate thiokinase enzyme. Probably CoA is first displaced by inorganic phosphate, forming succinyl phosphate. A nitrogen atom of a specific histidine residue then attacks phosphorus, displacing succinate and forming an A-phos-phoryl derivative. In the final step GDP attacks the phosphorus atom of that derivative forming GTP. The role of GTP in this reaction is played by ATP in some organisms. 

Notice the intermediate in the reaction of citrate synthase (fig. 13.7). Do you think at some time in the future evolution will produce a variety of citrate synthase that recovers the energy in the thioester, analogous to the production of GTP (ATP) by succinate thiokinase (page 291) Would this energy recovery have any effect on the thermodynamics of the tricarboxylic acid cycle ... 

Consider the glyceraldehyde-3-phosphate dehydro-genase-phosphoglycerokinase enzymes of glycolysis and the succinate thiokinase of the TCA cycle. Compare the mechanisms of incorporation of inorganic phosphate into the respective nucleoside diphosphates. 

This complete oxidative cycle is found in a number of archaebacteria. Halophiles can fulfil their energy requirements by metabolism of amino acids and other nitrogenous compounds, and therefore it is probable that they possess an oxidative citric acid cycle. Aitken and Brown [45] have reported the presence of the cycle s enzymes in Halobacterium halobium and we have found the key enzymes, citrate synthase and succinate thiokinase, in a range of classical and alkaliphilic halophiles [46], Thus, it is probable that the cycle is generally present in this group of archaebacteria, but exhaustive studies have not been carried out. 

In mitochondria, more than 90 % of the respiratory phosphorylation is catalyzed by the H -ATP-synthase, an enzyme converting the respiratory chain-produced electrochemical potential difference (ApH+) into ATP [1-4]. Very small (but sometimes essential) portion of the respiratory energy is converted to GTP by succinate thiokinase [4]. Both respiratory chain enzymes (Complexes I, III and IV), catalyzing electron transfer from NAD(P)H to O2, and H -ATP-synthase are localized in the inner mitochondrial membrane. The great majority of the formed ATP molecules is exported from mitochondria by the ATP/ADP antiporter in exchange for extramitochondrial ADP (eqs. 1-3). 

In this complex reaction catalyzed by succinyl-CoA synthase (succinate thiokinase), the energy-rich thioester linkage of succinyl-CoA is hydrolyzed with release of free energy that is conserved in the substrate phosphorylation of GDP with phosphate to form GTP ... 

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