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)

We characterized the genes in the regions of large inverted repeats (IRA and IRB, 10,058 base-pairs each) and a small single copy (SSC 19,813 bp) of chloroplast DNA from Marchantia polymorpha. The inverted repeat (IR) regions contain genes for four ribosomal RNAs (16 S, 23 S, 4.5 S and 5 S rRNAs) and five transfer RNAs (valine tRNA(GAC), isoleucine tRNA(GAU), alanine tRNA(UGC), arginine tRNA(ACG) and asparagine tRNA(GUU)). The gene organization of the IR regions in the liverwort chloroplast genome is conserved, although the IR regions are smaller (10,058 base-pairs) than any reported in higher plant chloroplasts. The small single-copy region (19,813 base-pairs) encoded genes for 17 open reading frames, a leucine tRNA(UAG) and a proline tRNA(GGG)-like sequence. We identified 12 open reading frames by homology of their coding sequences to a 4Fe-4S-type ferredoxin protein, a bacterial nitrogenase reductase component (Fe-protein), five human mitochondrial components of NADH dehydrogenase (ND1, ND4, ND4L, ND5 and ND6), two Escherichia coli ribosomal proteins (S15 and L21), two putative proteins encoded in the kinetoplast maxicircle DNA of Leishmania tarentolae (LtORF 3 and LtORF 4), and a bacterial permease inner membrane component (encoded by malF in E. coli or hisQ in Salmonella typhimurium).
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PMID:Structure and organization of Marchantia polymorpha chloroplast genome. IV. Inverted repeat and small single copy regions. 319 37

The nucleotide sequence of a 3849-bp fragment of starfish mitochondrial genome was determined. The genes for NADH dehydrogenase subunits 3, 4, 5, and COIII, and three kinds of (tRNA(UCNSer), tRNA(His), and tRNA(AGYSer) were identified by comparing with the genes of other animal mitochondria so far elucidated. The gene arrangement of starfish mitochondrial genome was different from those of vertebrate and insect mitochondrial genomes. Comparison of the protein-encoding nucleotide sequences of starfish mitochondria with those of other animal mitochondria suggested a unique genetic code in starfish mitochondrial genome; both AGA and AGG (arginine in the universal code) code for serine, AUA (isoleucine in the universal code but methionine in most mitochondrial systems) for isoleucine, and AAA (lysine) for asparagine. It was also inferred that these AGA and AGG codons are decoded by serine tRNA(AGYSer) originally corresponding to AGC and AGU codons. This situation is similar to the case of Drosophila mitochondrial genome. Variations in the use of AGA and AGG codons were discussed on the basis of the evolution of animals and decoding capacity of various tRNA(AGYSer) species possessing different sizes of the dihydrouridine (D) arm.
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PMID:Unusual genetic codes and a novel gene structure for tRNA(AGYSer) in starfish mitochondrial DNA. 367 36

Site-directed mutagenesis of the acidic clusters 207Asp-Asp-Asp209 and 213Glu-Glu-Asp215 of NADPH-cytochrome P450 oxidoreductase demonstrates that both cytochrome c and cytochrome P450 interact with this region; however, the sites and mechanisms of interaction of the two substrates are clearly distinct. Substitutions in the first acidic cluster did not affect cytochrome c or ferricyanide reductase activity, but substitution of asparagine for aspartate at position 208 reduced cytochrome P450-dependent benzphetamine N-demethylase activity by 63% with no effect on KP450m or KNADPHm. Substitutions in the second acidic cluster affected cytochrome c reduction but not benzphetamine N-demethylase or ferricyanide reductase activity. The E213Q enzyme exhibited a 59% reduction in cytochrome c reductase activity and a 47% reduction in KCyt cm under standard conditions (x0.27 M potassium phosphate, pH 7.7), as well as a decreased KCyt cm at every ionic strength and a shift of the salt dependence of cytochrome c reductase activity toward lower ionic strengths. The E214Q substitution did not affect cytochrome c reductase activity under standard conditions, but shifted the salt dependence of cytochrome c reductase activity toward higher ionic strengths. Measurements of the effect of ionic strength on steady-state kinetic properties indicated that increasing ionic strength destabilized the reductase-cytochrome c3+ ground state and reductase-cytochrome c transition state complexes for the wild-type, E213Q, and E214Q enzymes, suggesting the presence of electrostatic interactions involving Glu213 and Glu214 as well as additional residues outside this region. The ionic strength dependence of kcat/KCyt cm for the wild-type and E214Q enzymes is consistent with the presence of charge-pairing interactions in the transition state and removal of a weak ionic interaction in the reductase-cytochrome c transition-state complex by the E214Q substitution. The ionic strength dependence of the E213Q enzyme, however, is not consistent with a simple electrostatic model. Effects of ionic strength on kinetic properties of E213Q suggest that substitution of glutamine stabilizes the reductase-cytochrome c3+ ground-state complex, leading to a net increase in activation energy and decrease in kcat. Glu213 is also involved in a repulsive interaction with cytochrome c3+. Cytochrome c2+ Ki for the wild-type enzyme was 82.4 microM at 118 mM ionic strength and 10.8 microM at 749 mM ionic strength; similar values were observed for the E214Q enzyme. Cytochrome c Ki for the E213Q enzyme was 17.6 microM at 118 mM and 15.7 microM at 749 mM ionic strength, consistent with removal of an electrostatic repulsion between the reductase and cytochrome c2+.
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PMID:Role of acidic residues in the interaction of NADPH-cytochrome P450 oxidoreductase with cytochrome P450 and cytochrome c. 749 4

In yeast iso-1-cytochrome c, the side chain of histidine 26 (His26) attaches omega loop A to the main body of the protein by forming a hydrogen bond to the backbone atom carbonyl of glutamic acid 44. The His26 side chain also forms a stabilizing intra-loop interaction through a hydrogen bond to the backbone amide of asparagine 31. To investigate the importance of loop-protein attachment and intra-loop interactions to the structure and function of this protein, a series of site-directed and random-directed mutations were produced at His26. Yeast strains expressing these variant proteins were analyzed for their ability to grow on non-fermentable carbon sources and for their intracellular production of cytochrome c. While the data show that mutations at His26 lead to slightly decreased intracellular amounts of cytochrome c, the level of cytochrome c function is decreased more. The data suggest that cytochrome c reductase binding is affected more than cytochrome c oxidase or lactate dehydrogenase binding. We propose that mutations at this residue increase loop mobility, which, in turn, decreases the protein's ability to bind redox partners.
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PMID:Mutagenesis of histidine 26 demonstrates the importance of loop-loop and loop-protein interactions for the function of iso-1-cytochrome c. 956 6

Site-directed mutagenesis has been employed to study the mechanism of hydride transfer from NADPH to NADPH-cytochrome P450 oxidoreductase. Specifically, Ser457, Asp675, and Cys630 have been selected because of their proximity to the isoalloxazine ring of FAD. Substitution of Asp675 with asparagine or valine decreased cytochrome c reductase activities 17- and 677-fold, respectively, while the C630A substitution decreased enzymatic activity 49-fold. Earlier studies had shown that the S457A mutation decreased cytochrome c reductase activity 90-fold and also lowered the redox potential of the FAD semiquinone (Shen, A., and Kasper, C. B. (1996) Biochemistry 35, 9451-9459). The S457A/D675N and S457A/D675N/C630A mutants produced roughly multiplicative decreases in cytochrome c reductase activity (774- and 22000-fold, respectively) with corresponding decreases in the rates of flavin reduction. For each mutation, increases were observed in the magnitudes of the primary deuterium isotope effects with NADPD, consistent with decreased rates of hydride transfer from NADPH to FAD and an increase in the relative rate limitation of hydride transfer. Asp675 substitutions lowered the redox potential of the FAD semiquinone. In addition, the C630A substitution shifted the pKa of an ionizable group previously identified as necessary for catalysis (Sem, D. S., and Kasper, C. B. (1993) Biochemistry 32, 11539-11547) from 6.9 to 7.8. These results are consistent with a model in which Ser457, Asp675, and Cys630 stabilize the transition state for hydride transfer. Ser457 and Asp675 interact to stabilize both the transition state and the FAD semiquinone, while Cys630 interacts with the nicotinamide ring and the fully reduced FAD, functioning as a proton donor/acceptor to FAD.
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PMID:Mechanistic studies on the reductive half-reaction of NADPH-cytochrome P450 oxidoreductase. 1002 49

Phylogenetic relationships of a subset of Aphanius fish comprising central Anatolia, Turkey, are investigated to test the hypothesis of geographic speciation driven by early Pliocene orogenic events in spite of morphological similarity. We use 3434 aligned base pairs of mitochondrial DNA from 42 samples representing 36 populations of three species and six outgroup species to test this hypothesis. Genes analyzed include those encoding the 12S and 16S ribosomal RNAs; transfer RNAs coding for valine, leucine, isoleucine, glutamine, methionine, tryptophan, alanine, asparagine, cysteine, and tyrosine; and complete NADH dehydrogenase subunits I and II. Distance based minimum evolution and maximum-likelihood analyses identify six well-supported clades consisting of Aphanius danfordii, Aphanius sp. aff danfordii, and four clades of Aphanius anatoliae. Parsimony analysis results in 462 equally parsimonious trees, all of which contain the six well supported clades identified in the other analyses. Our phylogenetic results are supported by hybridization studies (Villwock, 1964), and by the geological history of Anatolia. Phylogenetic relationships among the six clades are only weakly supported, however, and differ among analytical methods. We therefore test and subsequently reject the hypothesis of simultaneous diversification among the six central Anatolian clades. However, our analyses do not identify any internodes that are significantly better supported than expected by chance alone. Therefore, although bifurcating branching order is hypothesized to underlie this radiation, the exact branching order is difficult to estimate with confidence.
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PMID:Molecular phylogeny and historical biogeography of the Aphanius (Pisces, Cyprinodontiformes) species complex of central Anatolia, Turkey. 1238 56

Two conserved charged amino acids, aspartate-186 and arginine-190, localized in the aqueous head region of the iron-sulfur protein of the cytochrome bc(1) complex of yeast mitochondria, were mutated to alanine, glutamate, or asparagine and isoleucine, respectively. The R190I mutation resulted in the complete loss of antimycin- and myxothiazol-sensitive cytochrome c reductase activity due to loss of more than 60% of the iron-sulfur protein in the complex. Mitochondria isolated from the D186A mutant had a 50% decrease in cytochrome c reductase activity but no loss of the iron-sulfur protein or the [2Fe-2S] cluster. The midpoint potential of the [2Fe-2S] cluster of the D186A mutant was decreased from 281 to 178 mV. The D186E and D186N mutations did not result in a loss of cytochrome c reductase activity or content of iron-sulfur protein; however, the redox potential of the [2Fe-2S] cluster of D186N was decreased from 281 to 241 mV. Molecular modeling/dynamics studies predicted that substituting an alanine for Asp-186 causes global structural changes in the head group of the iron-sulfur protein resulting in changes in the orientation of the [2Fe-2S] cluster and consequently a lowered redox potential. The rate of electrogenic proton pumping in the bc(1) complex isolated from mutant D186A reconstituted into proteoliposomes decreased 64%; however, the H(+)/2e(-) ratio of 1.9 was identical in the mutant and the wild-type complexes. The carboxyl binding reagent, N-(ethoxycarbonyl)-2-ethoxyl-1,2-dihydroquinoline (EEDQ) blocked electrogenic proton pumping in the bc(1) complex reconstituted into proteoliposomes without affecting electron transfer resulting in a decrease in the H(+)/2e(-) ratio to 1.2 and 1.1, respectively. EEDQ was bound to the iron-sulfur protein and core protein II in both the wild type and the D186A mutant, indicating that Asp-186 of the iron-sulfur protein is not required for proton translocation in the bc(1) complex.
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PMID:Aspartate-186 in the head group of the yeast iron-sulfur protein of the cytochrome bc1 complex contributes to the protein conformation required for efficient electron transfer. 1467 May 97

BphA3 from Pseudomonas sp. KKS102 is a Rieske-type [2Fe-2S] ferredoxin that transfers electrons from an NADH-dependent oxidoreductase, BphA4, to a biphenyl dioxygenase complex. A high-level expression and purification system for the recombinant BphA3 in Escherichia coli was constructed. Two histidine ligands of the Rieske-type cluster in BphA3, were each replaced with serine, cysteine, asparagine and tyrosine. The single mutants, in which either His44 or His65 was replaced with a cysteine residue (CH and HC mutants respectively), and the double mutant, in which both histidine residues were replaced with cysteine residue (CC mutant), accumulated to high levels in the E. coli cells, while the other single mutants did not. The purified WT (wild-type) protein showed characteristic near-UV and visible absorption and CD spectra of Rieske-type clusters. The X-ray absorption spectra were suggestive of the existence of [2Fe-2S] clusters, with one histidine and three cysteine ligands in the CH and HC mutants, and an [2Fe-2S] cluster with four cysteine ligands in the CC mutant. The BphA4-dependent cytochrome c reductase activities of the mutants were less than 0.3% of that of the WT protein. The redox potential of the WT protein determined by cyclic voltammetry was -180+/-5 mV compared with the standard hydrogen electrode, and that of the CH mutant was approx. 175 mV lower. The changes in the near-UV and visible absorption spectra of the mutants showed that the reduced iron-sulphur clusters in the mutants were unstable. His44 and His65 in BphA3 can be replaced with cysteine residues, but are required for the stabilization of the reduced form of the cluster.
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PMID:Tolerance of the Rieske-type [2Fe-2S] cluster in recombinant ferredoxin BphA3 from Pseudomonas sp. KKS102 to histidine ligand mutations. 1573 56

Phylogenetic relationships of members of the subfamily Poeciliinae (Cyprinodontiformes) are investigated to test alternate hypotheses of diversification resulting from the assembly of the Central America and the Caribbean from the Cretaceous period onwards. We use 4333 aligned base pairs of mitochondrial DNA and 1549 aligned base pairs of nuclear DNA from 55 samples representing 48 ingroup and seven outgroup species to test this hypothesis. Mitochondrial genes analyzed include those encoding the 12S and 16S ribosomal RNAs; transfer RNAs coding for valine, leucine, isoleucine, glutamine, methionine, tryptophan, alanine, asparagine, cysteine and tyrosine; and complete cytochrome b and NADH dehydrogenase subunit I and II; nuclear gene analyzed included the third exon of the recombination activation gene 1 (RAG1). Analyses of combined mtDNA and nuclear DNA data sets result in a well-supported phylogenetic hypothesis. This hypothesis is in conflict with the classical taxonomic assignment of genera into tribes and phylogenetic hypotheses based on the taxonomy; however, the molecular hypothesis defines nine clades that are geographically restricted and consistent with the geological evolution of Central America and the Caribbean. Our analyses support multiple colonization events of Middle America followed by a mix of vicariance and dispersal events.
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PMID:A phylogenetic and biogeographic perspective on the evolution of poeciliid fishes. 1718 5

Leigh syndrome can be caused by defects in both nuclear and mitochondrial genes involved in energy metabolism. Recently, an increasing number of mutations in mitochondrial DNA encoding regions, especially in NADH dehydrogenase (respiratory chain complex I) subunits, have been reported as causative of early onset Leigh syndrome. We describe a patient whose fetal brain ultrasound demonstrated periventricular pseudocyst suggestive of a possible mitochondrial disorder who presented postnatally with Leigh syndrome. A muscle biopsy demonstrated a partial decrease in complex I and pyruvate dehydrogenase (PDH-E1 alpha) activity. Sequencing of the PDH-E1 alpha gene did not reveal any mutation. Sequencing of the mtDNA revealed a novel heteroplasmic G10254A (D66N) mutation in the ND3 gene. This change results in a substitution of aspartic acid to asparagine in a highly conserved domain of the ND3 subunit. The mutation could not be detected in the mother's blood or urine sediment. Blue native gel electrophoresis of muscle mitochondria revealed a normal size, albeit a decreased level of complex I. The G10254A substitution in the mtDNA-ND3 gene is another cause of maternally inherited Leigh syndrome. This case demonstrates that periventricular pseudocysts may be the initial in utero presentation in patients with mitochondrial disorders. We emphasize the importance of screening the mtDNA in pediatric patients as the first step in molecular diagnosis of Leigh syndrome.
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PMID:Leigh disease presenting in utero due to a novel missense mutation in the mitochondrial DNA-ND3. 2020 74


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