Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:1.6.5.3 (complex I)
8,901 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bovine mitochondrial complex I (NADH: ubiquinone oxidoreductase) is composed of 3 structural domains, designated FP (flavoprotein, 3 subunits), IP (iron-sulfur protein, 7-8 subunits) and HP (hydrophobic protein, > 30 subunits). IP intervenes between FP and HP, and in complex I its 75 kDa subunit appears to interact with the 51 kDa subunit of FP. In this study, we show by the technique of ligand blotting that isolated IP binds (a) only to the 51 kDa subunit of FP, and (b) to the 42, 39, 23, 20 and 16 kDa subunits of HP. Because a 23 kDa and a 20 kDa subunit of complex I are potential iron-sulfur proteins, these and our previous results are consistent with the following possible path of electrons in complex I: NADH-->51 and 24 kDa subunit of FP-->75 kDa subunit of IP-->23 and 20 kDa subunits of HP-->ubiquinone.
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PMID:Intersubunit interactions in the bovine mitochondrial complex I as revealed by ligand blotting. 885 15

Cytosolic NADPH-dependent ubiquinone reductase (NADPH-UQ reductase) accounted for about 68% of the total ubiquinone (UQ) reductase activity in rat liver homogenate [Takahashi, T. et al. (1995) Biochem. J. 309, 883-890]. We investigated the effects of various factors on this enzyme activity in rat liver cytosol with the aim of elucidating its physiological roles. The NADPH-UQ reductase in rat liver cytosol catalyzed the reduction of UQ to UQH2 with concomitant oxidation of equimolar NADPH. The optimal pH was around 7.4, and the optimal temperatures were about 28 degrees C for NADH and about 37 degrees C for NADPH. NADH, deamino NADH, and deamino NADPH were much less active hydrogen donors than NADPH, whereas reduced nicotinamide mononucleotide, ascorbate, erythorbate, reduced glutathione, and cysteine were inactive. As the hydrogen acceptor, UQ-9 had the highest Vmax/Km among the long-chain UQ homologues tested. FAD and FMN stimulated the activity. Anionic detergents, Mg2+ and Sr2+ also enhanced the activity. Rotenone, malonic acid, antimycin A, and KCN, which inhibit mitochondrial and microsomal electron transfer enzymes, superoxide dismutase, and acetylated cytochrome c had no effect on the NADPH-UQ reductase activity. These results indicated that the NADPH-UQ reductase in rat liver cytosol is a flavoprotein that reduces UQ-10 by a two-electron reduction mechanism and is distinguishable from known microsomal and mitochondrial enzymes, as well as DT-diaphorase [EC 1.6.99.2].
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PMID:Characterization of NADPH-dependent ubiquinone reductase activity in rat liver cytosol: effect of various factors on ubiquinone-reducing activity and discrimination from other quinone reductases. 888 15

Defects in electron transfer flavoprotein (ETF) or its electron acceptor, electron transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO), cause the human inherited metabolic disease glutaric acidemia type II. In this disease, electron transfer from nine primary flavoprotein dehydrogenases to the main respiratory chain is impaired. Among these dehydrogenases are the four chain length-specific flavoprotein dehydrogenases of fatty acid beta-oxidation. In this investigation, two mutations in the alpha subunit that have been identified in patients were expressed in Escherichia coli. Of the two mutant alleles, alphaT266M and alphaG116R, the former is the most frequent mutation found in patients with ETF deficiency. The crystal structure of human ETF shows that alphaG116 lies in a hydrophobic pocket, under a contact residue of the alpha/beta subunit interface, and that the hydroxyl hydrogen of alphaT266 is hydrogen-bonded to N(5) of the FAD; the amide backbone hydrogen of alphaT266 is hydrogen-bonded to C(4)-O of the flavin prosthetic group (Roberts, D. L., Frerman, F. E. and Kim, J-J. P. (1996) Proc. Natl. Acad. Sci. U. S. A. 93, 14355-14360). Stable expression of the alphaG116R ETF required coexpression of the chaperonins, GroEL and GroES. alphaG116R ETF folds into a conformation different from the wild type, and is catalytically inactive in crude extracts. It is unstable and could not be extensively purified. The alphaT266M ETF was purified and characterized after stabilization to proteolysis in crude extracts. Although the global structure of this mutant protein is unchanged, its flavin environment is altered as indicated by absorption and circular dichroism spectroscopy and the kinetics of flavin release from the oxidized and reduced protein. The loss of the hydrogen bond at N(5) of the flavin and the altered flavin binding increase the thermodynamic stability of the flavin semiquinone by 10-fold relative to the semiquinone of wild type ETF. The mutation has relatively little effect on the reductive half-reaction of ETF catalyzed by sarcosine and medium chain acyl-CoA dehydrogenases which reduce the flavin to the semiquinone. However, kcat/Km of ETF-QO in a coupled acyl-CoA:ubiquinone reductase assay with oxidized alphaT266M ETF as substrate is reduced 33-fold; this decrease is due in largest part to a decrease in the rate of disproportionation of the alphaT266M ETF semiquinone catalyzed by ETF-QO.
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PMID:Expression and characterization of two pathogenic mutations in human electron transfer flavoprotein. 933 18

The human gene for the 10-kDa flavoprotein subunit of the mitochondrial NADH:ubiquinone oxidoreductase (Complex I) was completely cloned and sequenced. The so-called NDUFV3 gene contains three exons, spanning 20 kb. The open reading frame contains a 34-codon import sequence and a 74-codon mature protein sequence. A database search revealed close homology to bovine and rat protein sequence but not to any other known protein. Northern blot analysis showed that the NDUFV3 gene is ubiquitously expressed. The NDUFV3 gene was assigned by FISH to a single location on chromosome 21q22.3 and might contribute to the Down syndrome phenotype.
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PMID:Molecular cloning and characterization of the human mitochondrial NADH:oxidoreductase 10-kDa gene (NDUFV3). 934 73

NADH:ubiquinone oxidoreductase (complex I) of the mitochondrial respiratory chain can be fragmented in a flavoprotein (FP), iron-sulfur protein (IP), and hydrophobic protein (HP) subfraction. The IP subfraction is hypothesized to be significant, since it contains important prosthetic groups highly conserved among species. We cloned the cDNA of three remaining human NADH:ubiquinone oxidoreductase subunits of this IP fraction: the NDUFS2 (49 kDa), NDUFS3 (30 kDa), and NDUFS6 (13 kDa) subunits. All presented cDNAs include the complete open reading frame (ORF), which consist of 1392, 795, and 375 base pairs, coding for 463, 264, and 124 amino acids, respectively. The latter show 96, 90, and 83% homology with the corresponding bovine translation products. The 3' untranslated regions (UTR) are complete in all three cDNAs. Polymerase chain reaction performed with DNA isolated from somatic human-rodent cell hybrids containing defined human chromosomes as template gave a human-specific signal which mapped the NDUFS2 and NDUFS3 subunits to chromosomes 1 and 11, respectively. In the case of the NDUFS6 subunit a pseudogene may be present since signals were seen in the lanes containing chromosomes 5 and 6. The NDUFS2 contains a highly conserved protein kinase C phosphorylation site and the NDUFS3 subunit contains a highly conserved casein kinase II phosphorylation site which make them strong candidates for future mutation detection studies in enzymatic complex I-deficient patients.
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PMID:cDNA sequence and chromosomal localization of the remaining three human nuclear encoded iron sulphur protein (IP) subunits of complex I: the human IP fraction is completed. 964 66

The genes that encode the two different subunits of the novel electron-transferring flavoprotein (ETF) from Megasphaera elsdenii were identified by screening a partial genomic DNA library with a probe that was generated by amplification of genomic sequences using the polymerase chain reaction. The cloned genes are arranged in tandem with the coding sequence for the beta-subunit in the position 5' to the alpha-subunit coding sequence. Amino acid sequence analysis of the two subunits revealed that there are two possible dinucleotide-binding sites on the alpha-subunit and one on the beta-subunit. Comparison of M. elsdenii ETF amino acid sequence to other ETFs and ETF-like proteins indicates that while homology occurs with the mitochondrial ETF and bacterial ETFs, the greatest similarity is with the putative ETFs from clostridia and with fixAB gene products from nitrogen-fixing bacteria. The recombinant ETF was isolated from extracts of Escherichia coli. It is a heterodimer with subunits identical in size to the native protein. The isolated enzyme contains approximately 1 mol of FAD, but like the native protein it binds additional flavin to give a total of about 2 mol of FAD/dimer. It serves as an electron donor to butyryl-CoA dehydrogenase, and it also has NADH dehydrogenase activity.
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PMID:Cloning and analysis of the genes for a novel electron-transferring flavoprotein from Megasphaera elsdenii. Expression and characterization of the recombinant protein. 969 53

The fitness of organisms depends upon the rate at which they generate superoxide (O-2) and hydrogen peroxide (H2O2) as toxic by-products of aerobic metabolism. In Escherichia coli these oxidants arise primarily from the autoxidation of components of its respiratory chain. Inverted vesicles that were incubated with NADH generated O-2 and H2O2 at accelerated rates either when treated with cyanide or when devoid of quinones, implicating an NADH dehydrogenase as their source. Null mutations in the gene encoding NADH dehydrogenase II averted autoxidation of vesicles, and its overproduction accelerated it. Thus NADH dehydrogenase II but not NADH dehydrogenase I, respiratory quinones, or cytochrome oxidases formed substantial O-2 and H2O2. NADH dehydrogenase II that was purified from both wild-type and quinone-deficient cells generated approximately 130 H2O2 and 15 O-2 min-1 by autoxidation of its reduced FAD cofactor. Sulfite reductase is a second autoxidizable electron transport chain of E. coli, containing FAD, FMN, [4Fe-4S], and siroheme moieties. Purified flavoprotein that contained only the FAD and FMN cofactors had about the same oxidation turnover number as did the holoenzyme, 7 min-1 FAD-1. Oxidase activity was largely lost upon FMN removal. Thus the autoxidation of sulfite reductase, like that of the respiratory chain, occurs primarily by autoxidation of an exposed flavin cofactor. Great variability in the oxidation turnover numbers of these and other flavoproteins suggests that endogenous oxidants will be predominantly formed by only a few oxidizable enzymes. Thus the degree of oxidative stress in a cell may depend upon the titer of such enzymes and accordingly may vary with growth conditions and among different cell types. Furthermore, the chemical nature of these reactions was manifested by their acceleration at high temperatures and oxygen concentrations. Thus these environmental parameters may also directly affect the O-2 and H2O2 loads that organisms must bear.
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PMID:The identification of primary sites of superoxide and hydrogen peroxide formation in the aerobic respiratory chain and sulfite reductase complex of Escherichia coli. 1018 94

NADH-quinone oxidoreductase is classified into two groups, NADH dehydrogenase-1 (NDH-1) and NADH dehydrogenase-2 (NDH-2). Animal mitochondrial complex I is an NDH-1 type enzyme. Previously, we isolated potent inhibitors from plants to both NDH-1 and NDH-2. We have now examined detailed inhibitory effects of three tannins (pentagalloylglucose, sanguiin H-11, and oolonghomobisflavan A) on NDH-1 using bovine heart mitochondrial complex I and a subcomplex flavoprotein (containing 3 subunits) derived from complex I. Although many specific inhibitors of NDH-1 (e.g. rotenone and piericidin A) have been reported, the reactive sites are at or near to, the ubiquinone-binding site. NADH-ubiquinone-1 oxidoreductase activity of complex I was inhibited by the three tannins, among which sanguiin H-11 was the most potent inhibitor. NADH-menadione oxidoreductase activity of complex I was susceptible to the three tannins, but completely resistant to rotenone. The inhibitory effects of tannins were all noncompetitive with respect to NADH, ubiquinone-1, and menadione. The NADH-menadione oxidoreductase of flavoprotein was also inhibited by the three tannins, but not by rotenone, which is consistent with the fact that flavoprotein does not contain a native ubiquinone-binding site. The study of the NADH reduced-minus-oxidized difference spectrum of flavoprotein under steady-state conditions indicated that the inhibitory sites of sanguiin H-11 and oolonghomobisflavan A exist between the NADH binding site and the FMN site, and that for pentagalloylglucose exists between FMN and an artificial electron acceptor-binding site. These results suggest that the tannins are potent inhibitors of NADH dehydrogenases, and that the inhibitory mechanisms are novel.
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PMID:Inhibitory effects of tannins on the NADH dehydrogenase activity of bovine heart mitochondrial complex I. 1022 Feb 77

Oxygen sensing was investigated in rat pheochromocytoma PC12 cells. They respond to hypoxia with an increased intracellular generation of reactive oxygen species (ROS), measured by oxidation of dihydrorhodamine 123. This increase is abolished by intracellular superoxide scavenging by Mn(III)-tetrakis(1-methyl-4-pyridyl)-porphyrin, and reduced or absent in the presence of the flavoprotein/complex I inhibitors, diphenyl-eneiodonium and rotenone. The same inhibitors, but neither intra- nor extracellular (superoxide dismutase) superoxide scavenging, abolish the hypoxia-induced increase in tyrosine hydroxylase (TH) gene expression. Thus, ROS production increases in PC12 cells during hypoxia, but this is not the cause of hypoxic TH mRNA upregulation that involves a flavoprotein.
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PMID:Hypoxic upregulation of tyrosine hydroxylase gene expression is paralleled, but not induced, by increased generation of reactive oxygen species in PC12 cells. 1048 62

Some sterically hindered N-substituted derivatives of daunorubicin are known to be poor substrates for NADH dehydrogenase, NADPH cytochrome P450 reductase and xanthine oxidase. In consequence, poor oxygen radical generation by these compounds is observed. In this study we examined a new family of sugar-N-substituted derivatives of daunorubicin bearing a bulky substituent introduced on the nitrogen atom through the amidine spacer. These compounds were found to be very active in radical formation catalyzed by all three studied enzymes. Thus, the introduction of a heterocyclic ring, even if it is bulky but flexible, on the nitrogen atom of daunosamine moiety through the one-atom spacer (amidine group), does not induce the steric hindrance effect on the interaction of daunorubicin derivatives with these flavoprotein enzymes.
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PMID:The ability of new formamidine sugar-modified derivatives of daunorubicin to stimulate free radical formation in three enzymatic systems: NADH dehydrogenase, NADPH cytochrome P450 reductase and xanthine oxidase. 1096 87


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