Identification of ubiquinone-binding proteins in yeast mitochondrial ubiquinol-cytochrome c reductase using an azido-ubiquinone derivative (original) (raw)
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Journal of Biological Chemistry
The amino acid sequence of the ubiquinone binding protein (QP-C) in the cytochrome b c l region of the mitochondrial electron transfer chain was determined by analysis of peptides obtained by cyanogen bromide cleavage and staphylococcal protease digestion of succinylated derivatives. It was found to consist of 110 amino acid residues and its amino terminus to be blocked by an acetyl group, as determined by mass spectrometry of the amino-terminal peptide and a comparison with peptides chemically synthesized on highperformance liquid chromatography. The molecular weight of this ubiquinone binding protein including the acetyl group was calculated to be 13,389. The predicted secondary structure of QP-C has a-helical content of about 50% and QP-C was classified as an "all-a" or + 8" protein. This is the first report describing the amino acid sequence of the ubiquinone binding protein. A comparison of this sequence with that of the 14-kDa subunit of the yeast ubiquinolcytochrome c reductase complex from the nucleotide sequence showed these two sequences to be quite similar.
Biochemical Society Transactions, 1987
Structurc of' uhiquinol : cytochrome c reductase Ubiquinol : cytochrome c reductase (cytochrome reductase) from Neurospora crassa mitochondria is a dimer and the monomeric unit consists of nine different subunits. The subunits I and I I are known as core-proteins, III, IV and V are the cytochromes h, c, and the iron-sulphur protein, and VI to IX are small proteins without prosthetic groups (Weiss & Kolb, 1979). We have recently determined the low-resolution three-dimensional structure of cytochrome reductase and of a bc,-subcomplex of the enzyme by means of electron microscopy of membrane crystals (Leonard ct a/.. 1981; Karlsson et al., 1983). The structure of the dimeric cytochrome reductase shows that the monomeric units are related by a twofold axis running perpendicular to the membrane. They are elongated, protrude 7 nm into the matrix space of mitochondria and 3 nm into the intermembrane space and contact each other partly in the membrane, partly in the matrix space (Figs. 1 and 2). In the structure of the hc,-subcomplex, which is a dimer like the whole enzyme but lacks the core-protein and iron-sulphur protein, the large peripheral section and an outward-facing part of the small peripheral section are missing. Taking into consideration secondary structures of subunits predicted from Abbreviations used: Q. ubiquinone-10; QH2. reduced ubiquinone-10.
The Journal of biological chemistry, 1979
Although the energy conserving membranes of the photosynthetic bacterium Rhodopseudomonas sphaeroides contain a 25 (+/- 3)-fold molar excess of ubiquinone over the photochemical reaction center, the activity of the ubiquinone-cytochrome b-c2 oxidoreductase is unaffected by quinone extraction until only 3, or at most 4, ubiquinones remain; only then does further extraction prevent the function of the oxidoreductase. Since 2 of these last ubiquinones are integral parts of the photochemical reaction center, we conclude that the ubiquinone-cytochrome b-c2 oxidoreductase requires only 1, or at most 2, molecules of ubiquinone-10 for its function. Earlier kinetic data identified a major electron donor to ferricytochrome c2 as a single molecule (known as Z) which requires 2 electrons and 2 protons for its equilibrium reduction. Hence, we identify a single molecule of quinone, probably ubiquinone-10 in a special environment, as a major electron donor to ferricytochrome c2 in the ubiquinone c...
A New Ubiquinone Metabolite and Its Activity at the Mitochondrial bc 1 Complex
Chemical Research in Toxicology, 2007
Ubichromanol, a reductive cyclization product of ubiquinone, acts as radical scavenging antioxidant and is similarly effective as R-tocopherol. However, nothing is known so far on the two-electron oxidation product of this antioxidant and its bioactivity. This study demonstrates that ubichromanol yields a ubiquinone-like compound with a hydroxyl-substituted side chain (UQOH) on oxidation. HPLC/MS and HPLC/ECD measurements revealed its natural presence in bovine liver mitochondria. The bioactivity of this formerly unknown compound as substrate for mitochondrial complex III was tested by measurements of the quinol:cytochrome c oxidoreductase activity in bovine submitochondrial particles and isolated mitochondrial bc 1 complex. Consistently in both model systems, reduced UQOH exhibited substrate efficiencies below that of native ubiquinone but a significantly higher efficiency than R-tocopheryl quinone. Model calculations revealed that on binding of reduced UQOH to the bc 1 complex the polar hydroxyl group was located close to hydrophobic amino acid residues. This fact could in part explain the lower efficiency of reduced UQOH in comparison to ubiquinone as a substrate for the mitochondrial bc 1 complex. Therefore, the hydroxylation of the aliphatic or isoprenoid side chains of bioquinones, which is typical for quinoid oxidation products of chromanols, such as R-tocopherol and ubichromanol, disturbs substrate binding at the mitochondrial electron-transfer complexes, which usually interact with ubiquinone.
The biomolecule ubiquinone exerts a variety of biological functions
BioFactors, 2003
The chemistry of ubiquinone allows reversible addition of single electrons and protons. This unique property is used in nature for aerobic energy gain, for unilateral proton accumulation, for the generation of reactive oxygen species involved in physiological signaling and a variety of pathophysiological events. Since several years ubiquinone is also considered to play a major role in the control of lipid peroxidation, since this lipophilic biomolecule was recognized to recycle α-tocopherol radicals back to the chain-breaking form, vitamin E. Ubiquinone is therefore a biomolecule which has increasingly focused the interest of many research groups due to its alternative pro-and antioxidant activity. We have intensively investigated the role of ubiquinone as prooxidant in mitochondria and will present experimental evidences on conditions required for this function, we will also show that lysosomal ubiquinone has a double function as proton translocator and radical source under certain metabolic conditions. Furthermore, we have addressed the antioxidant role of ubiquinone and found that the efficiency of this activity is widely dependent on the type of biomembrane where ubiquinone exerts its chain-breaking activity.
Archives of Biochemistry and Biophysics, 2000
Determination of the number of ubiquinone-and inhibitor-binding sites in the mitochondrial complex I (NADH:ubiquinone oxidoreductase) is a controversial question with a direct implication for elaborating a suitable model to explain the bioenergetic mechanism of this complicated enzyme. We have used combinations of both selective inhibitors and common ubiquinone-like substrates to demonstrate the multiplicity of the reaction centers in the complex I in contrast with competition studies that have suggested the existence of a unique binding site for ubiquinone. Our results provide new evidence for the existence of at least two freely exchangeable ubiquinone-binding sites with different specificity for substrates, as well as for a different kinetic interaction of inhibitors with the enzyme.
The specificity of mitochondrial complex I for ubiquinones
Biochemical Journal, 1996
We report the first detailed study on the ubiquinone (coenzyme Q; abbreviated to Q) analogue specificity of mitochondrial complex I, NADH: Q reductase, in intact submitochondrial particles. The enzymic function of complex I has been investigated using a series of analogues of Q as electron acceptor substrates for both electron transport activity and the associated generation of membrane potential.