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:1.6.99.3 (diaphorase)
5,903 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Antibodies have been raised against synthetic peptides corresponding to several computer-predicted epitopes of three mtDNA-encoded subunits, ND4, ND5 and ND6, of the human respiratory chain NADH dehydrogenase (Complex I). Antibodies were characterized by a sensitive immunoblotting assay using proteins from human skeletal muscle mitochondria and by immunoprecipitation of radio-labeled HeLa cell mitochondrial translation products. Only antibodies against two of six selected peptides of the ND4 subunit, i.e., the C-terminal peptide and an internal peptide close to the C-terminus, reacted in both assays with the subunit. Antibodies raised against an internal peptide close to the N-terminus of the ND5 subunit and antibodies raised against an internal epitope of the ND6 subunit also reacted in both the immunoblotting and immunoprecipitation assays. The antibodies described above and other Complex I subunit- or holoenzyme-specific antibodies were used to investigate the subunit deficiencies of the respiratory NADH dehydrogenase in the skeletal muscle of patients affected by mitochondrial myopathies associated with Complex I defects. The reduction in enzyme activity correlated in an immunoblot assay with a decrease of four mtDNA-encoded subunits of the enzyme, as well as with a decrease of other subunits of Complex I encoded in the nDNA. The present work provides the first evidence of a decrease in NADH dehydrogenase subunits encoded in the mitochondrial genome in myopathy patients.
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PMID:Multiple deficiencies of mitochondrial DNA- and nuclear-encoded subunits of respiratory NADH dehydrogenase detected with peptide- and subunit-specific antibodies in mitochondrial myopathies. 753 43

The mitochondrial NADH dehydrogenase (complex I) in mammalian cells is a multimeric enzyme consisting of approximately 40 subunits, 7 of which are encoded in mitochondrial DNA (mtDNA). Very little is known about the function of these mtDNA-encoded subunits. In this paper, we describe the efficient isolation from a human cell line of mutants affected in any of these subunits. In the course of analysis of eight mutants of the human cell line VA2B selected for their resistance to high concentrations of the complex I inhibitor rotenone, seven were found to be respiration deficient, and among these, six exhibited a specific defect of complex I. Transfer of mitochondria from these six mutants into human mtDNA-less cells revealed, surprisingly, in all cases a cotransfer of the complex I defect but not of the rotenone resistance. This result indicated that the rotenone resistance resulted from a nuclear mutation, while the respiration defect was produced by an mtDNA mutation. A detailed molecular analysis of the six complex I-deficient mutants revealed that two of them exhibited a frameshift mutation in the ND4 gene, in homoplasmic or in heteroplasmic form, resulting in the complete or partial loss, respectively, of the ND4 subunit; two other mutants exhibited a frameshift mutation in the ND5 gene, in near-homoplasmic or heteroplasmic form, resulting in the ND5 subunit being undetectable or strongly decreased, respectively. It was previously reported (G. Hofhaus and G. Attardi, EMBO J. 12:3043-3048, 1993) that the mutant completely lacking the ND4 subunit exhibited a total loss of NADH:Q1 oxidoreductase activity and a lack of assembly of the mtDNA-encoded subunits of complex I. By contrast, in the mutant characterized in this study in which the ND5 subunit was not detectable and which was nearly totally deficient in complex I activity, the capacity to assemble the mtDNA-encoded subunits of the enzyme was preserved, although with a decreased efficiency or a reduced stability of the assembled complex. The two remaining complex I-deficient mutants exhibited a normal rate of synthesis and assembly of the mtDNA-encoded subunits of the enzyme, and the mtDNA mutation(s) responsible for their NADH dehydrogenase defect remains to be identified. The selection scheme used in this work has proven to be very valuable for the isolation of mutants from the VA2B cell line which are affected in different mtDNA-encoded subunits of complex I and may be applicable to other cell lines.
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PMID:Efficient selection and characterization of mutants of a human cell line which are defective in mitochondrial DNA-encoded subunits of respiratory NADH dehydrogenase. 782 60

Analyses of the Trypanosoma equiperdum (ATCC 30019) maxicircle reveals deletions, duplications and rearrangement compared to T. brucei. The genes for 9S rRNA and 12 proteins are absent. The 12S rRNA and cytochrome oxidase subunit I (COI) genes lack their 3' ends and are adjacent indicating deletion of intervening genes. The remaining two NADH dehydrogenase subunit genes (ND4 and ND5), the ribosomal protein RPS12 gene and the CR5 gene are duplicated and rearranged. ND4, RPS12 and the CR4 transcripts are abundant in steady state RNA while 12S rRNA and COI transcripts are not detected. Full length ND5 transcripts are rare, if present, but chimeric ND5/ND4 transcripts are abundant. The CR4 and RPS12 transcripts are the size of unedited RNAs suggesting that they are processed. However, they are not edited normally, presumably due to the absence of minicircle gRNA genes.
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PMID:Mitochondrial transcripts are processed but are not edited normally in Trypanosoma equiperdum (ATCC 30019) which has kDNA sequence deletion and duplication. 820 73

In most eukaryotic cells, the respiratory chain NADH dehydrogenase (Complex I) is a multimeric enzyme under dual (nuclear and mitochondrial) genetic control. Several genes encoding subunits of this enzyme have been identified in the mitochondrial genome from various organisms, but the functions of these subunits are in most part unknown. We describe here a human cell line in which the enzyme lacks the mtDNA-encoded subunit ND4 due to a frameshift mutation in the gene. In this cell line, the other mtDNA-encoded subunits fail to assemble, while at least some of the nuclear-encoded subunits involved in the redox reactions appear to be assembled normally. In fact, while there is a complete loss of NADH:Q1 oxidoreductase activity, the NADH:Fe(CN)6 oxidoreductase activity is normal. These observations provide the first clear evidence that the ND4 gene product is essential for Complex I activity and give some insights into the function and the structural relationship of this polypeptide to the rest of the enzyme. They are also significant for understanding the pathogenetic mechanism of the ND4 gene mutation associated with Leber's hereditary optic neuropathy.
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PMID:Lack of assembly of mitochondrial DNA-encoded subunits of respiratory NADH dehydrogenase and loss of enzyme activity in a human cell mutant lacking the mitochondrial ND4 gene product. 834 46

Mitochondrial DNA from two genetically unrelated patients carrying the mutation at position 11778 that causes Leber's hereditary optic neuropathy has been transferred with mitochondria into human mtDNA-less rho0206 cells. As analyzed in several transmitochondrial cell lines thus obtained, the mutation, which is in the gene encoding subunit ND4 of the respiratory chain NADH dehydrogenase (ND), did not affect the synthesis, size, or stability of ND4, nor its incorporation into the enzyme complex. However, NADH dehydrogenase-dependent respiration, as measured in digitonin-permeabilized cells, was specifically decreased by approximately 40% in cells carrying the mutation. This decrease, which was significant at the 99.99% confidence level, was correlated with a significantly reduced ability of the mutant cells to grow in a medium containing galactose instead of glucose, indicating a clear impairment in their oxidative phosphorylation capacity. On the contrary, no decrease in rotenone-sensitive NADH dehydrogenase activity, using a water-soluble ubiquinone analogue as electron acceptor, was detected in disrupted mitochondrial membranes. This is the first cellular model exhibiting in a foreign nuclear background mitochondrial DNA-linked biochemical defects underlying the optic neuropathy phenotype.
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PMID:Respiration and growth defects in transmitochondrial cell lines carrying the 11778 mutation associated with Leber's hereditary optic neuropathy. 866 57

ND4 is a mitochondrially encoded component of NADH dehydrogenase (Complex I) of the respiratory chain. A cDNA encoding a fusion protein comprised of the N-terminus of GAL10 of Saccharomyces cerevisiae and an N-terminally truncated form of human ND4 was isolated by its ability to functionally complement the thymidine transport deficiency of S. cerevisiae grown under conditions of thymidylate starvation. Epitope-tagged GAL10-ND4 was shown by immunological methods to be present in the plasma membranes of yeast expressing the GAL10-ND4 encoding cDNA. The ability of the GAL10-ND4 fusion protein to induce uptake of thymidine raises the possibility that native ND4, which is predicted to have 12 transmembrane domains, may function as a transporter or channel in the mitochondrial inner membrane.
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PMID:Functional complementation of a membrane transport deficiency in Saccharomyces cerevisiae by recombinant ND4 fusion protein. 932 73

The nucleotide sequences of two segments of 6,737 ntp and 258 nto of the 18.4-kb circular mitochondrial (mt) DNA molecule of the soft coral Sarcophyton glaucum (phylum Cnidaria, class Anthozoa, subclass Octocorallia, order Alcyonacea) have been determined. The larger segment contains the 3' 191 ntp of the gene for subunit 1 of the respiratory chain NADH dehydrogenase (ND1), complete genes for cytochrome b (Cyt b), ND6, ND3, ND4L, and a bacterial MutS homologue (MSH), and the 5' terminal 1,124 ntp of the gene for the large subunit rRNA (1-rRNA). These genes are arranged in the order given and all are transcribed from the same strand of the molecule. The smaller segment contains the 3' terminal 134 ntp of the ND4 gene and a complete tRNA(f-Met) gene, and these genes are transcribed in opposite directions. As in the hexacorallian anthozoan, Metridium senile, the mt-genetic code of S. glaucum is near standard: that is, in contrast to the situation in mt-genetic codes of other invertebrate phyla, AGA and AGG specify arginine, and ATA specifies isoleucine. However, as appears to be universal for metazoan mt-genetic codes, TGA specifies tryptophan rather than termination. Also, as in M. senile the mt-tRNA(f-Met) gene has primary and secondary structural features resembling those of Escherichia coli initiator tRNA, including standard dihydrouridine and T psi C loop sequences, and a mismatched nucleotide pair at the top of the amino-acyl stem. The presence of a mutS gene homologue, which has not been reported to occur in any other known mtDNA, suggests that there is mismatch repair activity in S. glaucum mitochondria. In support of this, phylogenetic analysis of MutS family protein sequences indicates that the S. glaucum mtMSH protein is more closely related to the nuclear DNA-encoded mitochondrial mismatch repair protein (MSH1) of the yeast Saccharomyces cerevisiae than to eukaryotic homologues involved in nuclear function, or to bacterial homologues. Regarding the possible origin of the S. glaucum mtMSH gene, the phylogenetic analysis results, together with comparative base composition considerations, and the absence of an MSH gene in any other known mtDNA best support the hypothesis that S. glaucum mtDNA acquired the mtMSH gene from nuclear DNA early in the evolution of octocorals. The presence of mismatch repair activity in S. glaucum mitochondria might be expected to influence the rate of evolution of this organism's mtDNA.
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PMID:Mitochondrial DNA of the coral Sarcophyton glaucum contains a gene for a homologue of bacterial MutS: a possible case of gene transfer from the nucleus to the mitochondrion. 954 36

The nucleotide sequence of a 9240-base pair DNA fragment of the mitochondrial (mt) genome of a squid, Loligo bleekeri, was determined, in which 8 protein and 14 tRNA genes were identified. The gene organization of the mt-genome exhibits a greater resemblance to the gene organization of arthropods and a chiton, Katharina tunicata, than to those of a mussel, Mytilus edulis, and land snails. A cloverleaf-like structure was observed between the genes for subunits 4 and 5 of NADH dehydrogenase (ND4 and -5), which is considered to have originated from histidine tRNA. It is presumed that this structure functions as a transcriptional punctuation signal for the maturation of the ND4 and ND5 mRNAs.
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PMID:Gene contents and organization of a mitochondrial DNA segment of the squid Loligo bleekeri. 1022 73

The activities and mRNA abundances of enzymes that regulate the rate of electron flow through the electron transport chain (ETC), including NADH dehydrogenase, succinate dehydrogenase, and cytochrome c oxidase, were examined in young and senescent fetal lung fibroblasts (WI-38). We also determined the activities and mRNA abundances of antioxidant defenses including superoxide dismutase, catalase, and glutathione peroxidase. We confirmed our previous report of a senescence-related increase in the abundance of ND4, a mitochondrially encoded subunit of NADH dehydrogenase. The activities of cytochrome c oxidase and NADH dehydrogenase were also elevated in senescent cultures. No differences were observed in the mRNA abundances of COX-1, a mitochondrially encoded subunit of cytochrome c oxidase or of nuclearly encoded subunits of various electron transport components (SD, COX-4, and ND 51). Lucigenin-detected chemiluminescence and H2O2 generation were both elevated in senescent cells. Catalase activity was also elevated in senescent fibroblasts. However, no differences in catalase mRNA abundance were observed. A small decrease in GSH peroxidase (GPx) mRNA abundance was observed in senescent cells. No other changes in the activities or mRNA abundances of any of the antioxidant defenses were observed in early and late passage cultures. The relationships between oxidant generation, mitochondrial enzyme activities, and antioxidant defense observed during proliferative senescence are dissimilar to those detected between fetal and postnatal fibroblasts as well as those found between fibroblast lines obtained from young and old individuals. The relevance of the differences between these models is discussed.
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PMID:Differences in electron transport potential, antioxidant defenses, and oxidant generation in young and senescent fetal lung fibroblasts (WI-38). 1036 24

In this study, changes of the expression of two mitochondrial and two nuclear genes encoding the subunits of cytochrome c oxidase (CO) and NADH dehydrogenase (ND) were studied in the hippocampus, inferior parietal lobule, and cerebellum of 10 Alzheimer's disease (AD) and 10 age-matched control subjects. The altered proportion between CO II and CO IV mRNAs was observed in the AD brain. Changes of the proportion between CO II and CO IV transcripts may contribute to the kinetic perturbation of CO documented in AD. A coordinated decrease of ND4 and ND15 mRNAs was found in the AD hippocampus and inferior parietal lobule, but not in cerebellum. The decrease of ND4 gene expression may lead to the inhibition of normal ubiquinone oxidoreductase activity of ND. This study suggests that changes of the expression of mitochondrial and nuclear genes, encoding parts of ND and CO enzyme complexes, may contribute to alterations of oxidative metabolism in AD.
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PMID:The expression of several mitochondrial and nuclear genes encoding the subunits of electron transport chain enzyme complexes, cytochrome c oxidase, and NADH dehydrogenase, in different brain regions in Alzheimer's disease. 1044 60


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