Subunits succinate dehydrogenase

Complex II is perhaps better known by its other name—succinate dehydrogenase, the only TCA cycle enzyme that is an integral membrane protein in the inner mitochondrial membrane. This enzyme has a mass of approximately 100 to 140 kD and is composed of four subunits two Fe-S proteins of masses 70 kD and 27 kD, and two other peptides of masses 15 kD and 13 kD. Also known as flavoprotein 2 (FP2), it contains an FAD covalently bound to a histidine residue (see Figure 20.15), and three Fe-S centers a 4Fe-4S cluster, a 3Fe-4S cluster, and a 2Fe-2S cluster. When succinate is converted to fumarate in the TCA cycle, concomitant reduction of bound FAD to FADHg occurs in succinate dehydrogenase. This FADHg transfers its electrons immediately to Fe-S centers, which pass them on to UQ. Electron flow from succinate to UQ,... 

Complex II contains four peptides, the two largest form succinate dehydrogenase, the largest has covalently boiuid flavin adenine dinucleotide (FAD) which reacts with succinate, and the other has three iron-sulphur centers. Smaller subunits anchor the two larger subunits to the membrane and form the UQ binding site. Ubiquinone is the electron acceptor but complex II does not pump protons (see below). 

This complex consists of four subunits, all of which are encoded on nuclear DNA, synthesized on cytosolic ribosomes, and transported into mitochondria. The succinate dehydrogenase (SDH) component of the complex oxidizes succinate to fumarate with transfer of electrons via its prosthetic group, FAD, to ubiquinone. It is unique in that it participates both in the respiratory chain and in the tricarboxylic acid (TC A) cycle. Defects of complex II are rare and only about 10 cases have been reported to date. Clinical syndromes include myopathy, but the major presenting features are often encephalopathy, with seizures and psychomotor retardation. Succinate oxidation is severely impaired (Figure 11). 

Kuramochi, T., Hirawake, H., Kojima, S., Takamiya, S., Furashima, R., Aoki, T., Komuniecki, R.W. and Kita, K. (1994) Sequence comparison between the flavoprotein subunit of the fumarate reductase (complex II) of the anaerobic parasitic nematode, Ascaris suum, and the succinate dehydrogenase of the aerobic, free-living nematode, Caenorhabditis elegans. Molecular and Biochemical Parasitolog 68, 177-187. 

Tielens, A.G. and Roos, M.H. (1994) Differential expression of two succinate dehydrogenase subunit-(3 genes and a transition in energy metabolism during the development of the parasitic nematode Haemonchus contortus. Molecular and Biochemical Parasitology 66, 273-281. 

The activity of complex II (succinate dehydrogenase) is measured as the succinate-dependent reduction of decylubiquinone, which is in turn recorded spectro-photometrically through the reduction of dichlorophenol indophenol at 600 nm (e 19,100-M -cm Fig. 3.8.5). In order to ensure a linear rate for the activity, the medium is added with rotenone, ATP, and a high concentration of succinate. As noticed previously for complex I, decylubiquinone is not a perfect acceptor for electrons from the membrane-inserted complex II [70]. Malonate, a competitive inhibitor of the enzyme, is used to inhibit it. Rather than decylubiquinone, phenazine methosulfate can be utilized, which diverts the electrons from the complex before they are conveyed through subunits C and D, therefore allowing measurement of the activity of subunits A and B. 

Complex II The Succinate Dehydrogenase Complex. Succinate dehydrogenase is the only enzyme of the TCA cycle that is embedded in the inner membrane. Its four subunits include two iron-sulfur proteins, one of which also has a covalently attached FAD. As in NADH dehydrogenase, the substrate-oxidation site is on the matrix side of the membrane (fig. 14.10). 

Table 29 Subunit Structures and Other Properties of Fumarate Reductase and Succinate Dehydrogenase from...

Fumarate is able to serve as an electron acceptor in anaerobic respiration, as it may be reduced reversibly to succinate in a two-electron process. The succinate-fumarate couple may therefore be utilized as an oxidant or reductant in the respiratory chain, and so differs from the other examples given in this section. These two reactions are catalyzed by succinate dehydrogenase and fumarate reductase, which have many similarities in subunit structure. These are shown in Table 29. Although they are different enzymes, the fumarate reductase can substitute for succinate dehydrogenase under certain conditions. The synthesis of succinate dehydrogenase is induced... 

ESR studies on fumarate reductase in situ have suggested1436 the presence of two [2Fe-2S] clusters and a HiPIP cluster. Examination of the occurrence of cysteine residues in the two peptides and a comparison with succinate dehydrogenase suggests that the two-iron centres are in the smaller iron-sulfur protein, while the [4Fe-4S] centre may be in either subunit. 

Complex II consists of four polypeptide subunits (70,000, 27,000, 13,500, and 7,000 daltons). The two larger subunits are components of succinate dehydrogenase and this enzyme or the entire complex II was directly incorporated into phospholipid vesicles at the high protein lipid ratio of 1 2 (w/w). The phospholipid (egg yolk lecithin) contained a small amount of lipid (d) (Table 6.2 I molecule in 1000) or lipid (e) (Table 6.2 1 in 400). After irradiation the labeling pattern was analyzed directly by SDS-polyacrylamide gel electrophoresis. Presumably the lipids that did not become attached to polypeptides ran at the dye front, although this was not demonstrated. 

A number of interesting observations were made. When the reconstituted succinate dehydrogenase was labeled in the absence of the smallest subunits the lipid (e) labeled both the 70,000 dalton and the 27,000 dalton subunits, while lipid (d) labeled only the smaller subunit strongly. The authors concluded that the 70,000 dalton subunit was peripheral and the 27,000 dalton subunit integral. The small amount of label from (d) that was... 

Succinate dehydrogenase contains iron-sulfur centers and covalently bound FAD (both on the 70,000-dalton subunit). Iron-sulfur centers are also present on the 27,000-dalton subunit. The 15,500-dalton subunit of complex II is cytochrome bw Electrons from FADH2 are channeled to UQ via cytochrome b. ... 

The subunits separated by the above technique indicated a distribution of iron and labile sulfide as shown in Table X. Thus the larger subunit contained flavin, iron, and labile sulfide in the approximate ratio of 1 4 4, while the smaller subunit appeared to have the characteristics of a soluble iron-sulfur protein. The absorption spectra of succinate dehydrogenase and its two subunits analyzed to show the contributions of flavin and the iron-sulfur chromophore in each preparation are given in Fig. 27. 

From the small subunit obtained by chaotrope-induced resolution of succinate dehydrogenase (143). The small subunit obtained by resolution and column chromatography in presence of SDS is completely free of flavin. 

Fia. 27. Absorption spectra of succinate dehydrogenase (A), its larger subunit (B), and its smaller subunit (C). (A) Trace 1, oxidized succinate dehydrogenase trace 2, flavin contribution to the spectrum of oxidized enzyme trace 5, iron-sulfur contribution to the spectrum of oxidized enzyme trace 4, oxidized enzyme treated with dithionite trace 3, oxidized enzyme treated with sodium mersalyl and dithio-nite. Protein = 1.48 mg/ml. (B) Trace 1, oxidized trace 2, flavin contribution to trace 1 trace 3, after treatment of 1 with sodium mersalyl and dithionite. Protein = 3.0 mg/ml. (C) Trace 1, oxidized trace 2, after treatment with mersalyl trace 3, after treatment of 2 with dithionite. Protein = OA mg/ml. From Davis and Hatefi U43). 

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