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:1.12.7.2 (hydrogenase)
3,522 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Hydrogenase activity and the H(2)-fumarate electron transport system in a carbohydrate-fermenting obligate anaerobe, Bacteroides fragilis, were investigated. In both whole cells and cell extracts, hydrogenase activity was demonstrated with methylene blue, benzyl viologen, flavin mononucleotide, or flavin adenine dinucleotide as the electron acceptor. A catalytic quantity of benzyl viologen or ferredoxin from Clostridium pasteurianum was required to reduce nicotinamide adenine dinucleotide or nicotinamide adenine dinucleotide phosphate with H(2). Much of the hydrogenase activity appeared to be associated with the soluble fraction of the cell. Fumarate reduction to succinate by H(2) was demonstrable in cell extracts only in the presence of a catalytic quantity of benzyl viologen, flavin mononucleotide, flavin adenine dinucleotide, or ferredoxin from C. pasteurianum. Sulfhydryl compounds were not required for fumarate reduction by H(2), but mercaptoethanol and dithiothreitol appeared to stimulate this activity by 59 and 61%, respectively. Inhibition of fumarate reduction by acriflavin, rotenone, 2-heptyl-4-hydroxyquinoline-N-oxide, and antimycin A suggest the involvement of a flavoprotein, a quinone, and cytochrome b in the reduction of fumarate to succinate. The involvement of a quinone in fumarate reduction is also apparent from the inhibition of fumarate reduction by H(2) when cell extracts were irradiated with ultraviolet light. Based on the evidence obtained, a possible scheme for the flow of electrons from H(2) to fumarate in B. fragilis is proposed.
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PMID:Hydrogenase activity and the H2-fumarate electron transport system in Bacteroides fragilis. 89 48

Bovine mitochondrial NADH-ubiquinone reductase (complex I), the first enzyme in the electron-transport chain, is a membrane-bound assembly of more than 30 different proteins, and the flavoprotein (FP) fraction, a water-soluble assembly of the 51-, 24-, and 10-kDa subunits, retains some of the catalytic properties of the enzyme. The 51-kDa subunit binds the substrate NAD(H) and probably contains both the cofactor, FMN, and also a tetranuclear iron-sulfur center, while a binuclear iron-sulfur center is located in the 24- or 10-kDa proteins. The 75-kDa subunit is the largest of the six proteins in the iron-sulfur protein (IP) fraction, and its sequence indicates that it too contains iron-sulfur clusters. Partial protein sequences have been determined at the N-terminus and at internal sites in the 51-kDa subunit, and the corresponding cDNA encoding a precursor of the protein has been isolated by using a novel strategy based on the polymerase chain reaction. The mature protein is 444 amino acids long. Its sequence, and those of the 24- and 75-kDa subunits, shows that mitochondrial complex I is related to a soluble NAD-reducing hydrogenase from the facultative chemolithotroph Alcaligenes eutrophus H16. This enzyme has four subunits, alpha, beta, gamma, and delta, and the alpha gamma dimer is an NADH oxidoreductase that contains FMN. The gamma-subunit is related to residues 1-240 of the 75-kDa subunit of complex I, and the alpha-subunit sequence is a fusion of homologues of the 24- and 51-kDa subunits, in the order N- to C-terminal. The most highly conserved regions are in the 51-kDa subunit and probably form parts of nucleotide binding sites for NAD(H) and FMN. Another conserved region surrounds the sequence motif CysXXCysXXCys, which is likely to provide three of the four ligands of a 4Fe-4S center, possibly that known as N-3. Characteristic ligands for a second 4Fe-4S center are conserved in the 75-kDa and gamma-subunits. This relationship with the bacterial enzyme implies that the 24- and 51-kDa subunits, together with part of the 75-kDa subunit, constitute a structural unit in mitochondrial complex I that is concerned with the first steps of electron transport.
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PMID:Relationship between mitochondrial NADH-ubiquinone reductase and a bacterial NAD-reducing hydrogenase. 190 Jan 94

A brown carbon monoxide dehydrogenase from CO-autotrophically grown cells of Acinetobacter sp. strain JC1, which is unstable outside the cells, was purified 80-fold in seven steps to better than 95% homogeneity, with a yield of 44% in the presence of the stabilizing agents iodoacetamide (1 mM) and ammonium sulfate (100 mM). The final specific activity was 474 mumol of acceptor reduced per min per mg of protein as determined by an assay based on the CO-dependent reduction of thionin. Methyl viologen, NAD(P), flavin mononucleotide, flavin adenine dinucleotide, and ferricyanide were not reduced by the enzyme, but methylene blue, thionin, and dichlorophenolindophenol were reduced. The molecular weight of the native enzyme was determined to be 380,000. Sodium dodecyl sulfate-gel electrophoresis revealed at least three nonidentical subunits of molecular weights 16,000 (alpha), 34,000 (beta), and 85,000 (gamma). The purified enzyme contained particulate hydrogenase-like activity. Selenium did not stimulate carbon monoxide dehydrogenase activity. The isoelectic point of the native enzyme was found to be 5.8; the Km of CO was 150 microM. The enzyme was rapidly inactivated by methanol. One mole of native enzyme was found to contain 2 mol of each of flavin adenine dinucleotide and molybdenum and 8 mol each of nonheme iron and labile sulfide, which indicated that the enzyme was a molybdenum-containing iron-sulfur flavoprotein. The ratio of densities of each subunit after electrophoresis (alpha:beta:gamma = 1:2:6) and the number of each cofactor in the native enzyme suggest a alpha 2 beta 2 gamma 2 structure of the enzyme. The carbon monoxide dehydrogenase of Acinetobacter sp. strain JC1 was found to have no immunological relationship with enzymes of Pseudomonas carboxydohydrogena and Pseudomonas carboxydovorans.
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PMID:Purification and some properties of carbon monoxide dehydrogenase from Acinetobacter sp. strain JC1 DSM 3803. 253 87

The method of purification up to electrophoretical homogeneity of cytochrome c552 from the phototrophic bacterium Thiocapsa roseopersicina, strain BBS is described. For the cytochrome absorption spectrum the maxima at 417, 523 and 552 nm are characteristic for the reduced state and at 409 nm--for the oxidized state. The molecular weight is equal to 62000. The cytochrome contains two hemes per molecule and consists of two subunits. pI is 4.1; E0' is about 10 mV. Cytochrome c552 is a flavoprotein according to its fluorescence spectrum and subunit structure. T. roseopersicina cytochrome c552 is able to be reduced with sulphide, cysteine and ascorbate as well as with H2 in the presence of hydrogenase from the same bacterium. These data suggest that cytochrome c552 from T. roseopersicina functions in vivo at the initial stage of electron transport from hydrogen and sulphide.
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PMID:[Purification and properties of cytochrome c552 from purple sulfur bacterium Thiocapsa roseopersicina]. 626 68

A new FMN-containing flavoprotein isolated from Desulfovibrio gigas provided maximum coupling efficiency for the reduction of bisulfite from molecular H2. This protein, which is distinct from flavodoxin and for which the name flavoredoxin is proposed, is required for reconstitution of an electron transfer chain between hydrogenase and bisulfite reductase. A Ca(2+)-binding protein functions as a modulator in the presence of Ca2+ in the process. The finding of a membrane-bound cytochrome c with a molecular weight of 104,000 Da that is also active in this electron transfer chain provides an explanation for the energetic linkage between periplasmic and cytoplasmic proteins in this sulfate-reducing bacterium.
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PMID:Isolation and characterization of flavoredoxin, a new flavoprotein that permits in vitro reconstitution of an electron transfer chain from molecular hydrogen to sulfite reduction in the bacterium Desulfovibrio gigas. 838 52

A monomeric flavoprotein (18.8 kDa) was isolated from the soluble cell fraction of Wolinella succinogenes and was identified as a flavodoxin based on its N-terminal sequence, FMN content, and redox properties. The midpoint potentials of the flavodoxin (Fld) at pH 7. 5 were measured as -95 mV (Fldox/Flds) and -450 mV (Flds/Fldred) relative to the standard hydrogen electrode. The cellular flavodoxin content [0.3 micromol (g protein)-1] was the same in bacteria grown with fumarate or with polysulfide as the terminal acceptor of electron transport. The flavodoxin did not accept electrons from hydrogenase or formate dehydrogenase, the donor enzymes of electron transport to fumarate or polysulfide. Pyruvate:flavodoxin oxidoreductase activity [180 U (g cellular protein)-1] was detected in the soluble cell fraction of W. succinogenes grown with fumarate or polysulfide. The enzyme was equally active with Fldox or Flds at high concentrations. The Km for Flds (80 microM) was larger than that for Fldox and for the ferredoxin isolated from W. succinogenes (15 microM). We conclude that flavodoxin serves anabolic rather than catabolic functions in W. succinogenes.
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PMID:Flavodoxin from Wolinella succinogenes. 877 74

LANGUAGE="EN">Summary In most methanogenic archaea, two hydrogenase systems that can catalyze the reduction of coenzyme F420 (F420) with H2 are present: (1) the F420-reducing hydrogenase, which is a nickel iron-sulfur flavoprotein composed of three different subunits, and (2) the N5, N10-methylenetetrahydromethanopterin dehydrogenase system, which is composed of H2-forming methylenetetrahydromethanopterin dehydrogenase and F420-dependent methylenetetrahydromethanopterin dehydrogenase, both metal-free proteins without an apparent prosthetic group. We report here that in nickel-limited chemostat cultures of Methanobacterium thermoautotrophicum, the specific activity of the F420-reducing Ni/Fe-hydrogenase was essentially zero, whereas that of the H2-forming methylenetetrahydromethanopterin dehydrogenase was six times higher, and that of the F420-dependent methylenetetrahydromethanopterin dehydrogenase was four times higher than in cells grown under non-nickel-limited conditions. This evidence supports the hypothesis that when M. thermoautotrophicum grows under conditions of nickel limitation, the reduction of F420 with H2 is catalyzed by the metal-free methylenetetrahydromethanopterin dehydrogenase system.
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PMID:Function of H2-forming methylenetetrahydromethanopterin dehydrogenase from methanobacterium thermoautotrophicum in coenzyme F420 reduction with H2 947 54

The hyperthermophilic bacterium, Thermotoga maritima, grows up to 90 degrees C by fermenting carbohydrates and it disposes of excess reductant by H(2) production. The H(2)-evolving cytoplasmic hydrogenase of this organism was shown to consist of three different subunits of masses 73 (alpha), 68 (beta) and 19 (gamma) kDa and to contain iron as the only metal. The genes encoding the subunits were clustered in a single operon in the order hydC (gamma), hydB (beta), and hydA (alpha). Sequence analyses indicated that: (a) the enzyme is an Fe-S-cluster-containing flavoprotein which uses NADH as an electron donor; and (b) the catalytic Fe-S cluster resides within the alpha-subunit, which is equivalent to the single subunit that constitutes most mesophilic Fe-hydrogenases. The alpha- and beta-subunits of the purified enzyme were separated by chromatography in the presence of 4 M urea. As predicted, the H(2)-dependent methyl viologen reduction activity of the holoenzyme (45-70 U mg(-1)) was retained in the alpha-subunit (130-160 U mg(-1)) after subunit separation. However, the holoenzyme did not contain flavin and neither it nor the alpha-subunit used NAD(P)(H) or T. maritima ferredoxin as an electron carrier. The holoenzyme, but not the alpha-subunit, reduced anthraquinone-2,6-disulfonate (apparent K(m), 690 microM) with H(2). The EPR properties of the reduced holoenzyme, when compared with those of the separated and reduced subunits, indicate the presence of a catalytic 'H-cluster' and three [4Fe-4S] and one [2Fe-2S] cluster in the alpha-subunit, together with one [4Fe-4S] and two [2Fe-2S] clusters in the beta-subunit. Sequence analyses predict that the alpha-subunit should contain an additional [2Fe-2S] cluster, while the beta-subunit should contain one [2Fe-2S] and three [4Fe-4S] clusters. The latter cluster contents are consistent with the measured Fe contents of about 32, 20 and 14 Fe mol(-1) for the holoenzyme and the alpha- and beta-subunits, respectively. The T. maritima enzyme is the first 'complex' Fe-hydrogenase to be purified and characterized, although the reason for its complexity remains unclear.
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PMID:The hyperthermophilic bacterium, Thermotoga maritima, contains an unusually complex iron-hydrogenase: amino acid sequence analyses versus biochemical characterization. 1048 84

DNA microarrays were constructed by using 271 open reading frame (ORFs) from the genome of the archaeon Pyrococcus furiosus. They were used to investigate the effects of elemental sulfur (S(primary)) on the levels of gene expression in cells grown at 95 degrees C with maltose as the carbon source. The ORFs included those that are proposed to encode proteins mainly involved in the pathways of sugar and peptide catabolism, in the metabolism of metals, and in the biosynthesis of various cofactors, amino acids, and nucleotides. The expression of 21 ORFs decreased by more than fivefold when cells were grown with S(primary) and, of these, 18 encode subunits associated with three different hydrogenase systems. The remaining three ORFs encode homologs of ornithine carbamoyltransferase and HypF, both of which appear to be involved in hydrogenase biosynthesis, as well as a conserved hypothetical protein. The expression of two previously uncharacterized ORFs increased by more than 25-fold when cells were grown with S(primary). Their products, termed SipA and SipB (for sulfur-induced proteins), are proposed to be part of a novel S(primary)-reducing, membrane-associated, iron-sulfur cluster-containing complex. Two other previously uncharacterized ORFs encoding a putative flavoprotein and a second FeS protein were upregulated more than sixfold in S(primary)-grown cells, and these are also thought be involved in S(primary) reduction. Four ORFs that encode homologs of proteins involved in amino acid metabolism were similarly upregulated in S(primary)-grown cells, a finding consistent with the fact that growth on peptides is a S(primary)-dependent process. An ORF encoding a homolog of the eukaryotic rRNA processing protein, fibrillarin, was also upregulated sixfold in the presence of S(primary), although the reason for this is as yet unknown. Of the 20 S(primary)-independent ORFs that are the most highly expressed (at more than 20 times the detection limit), 12 of them represent enzymes purified from P. furiosus, but none of the products of the 34 S(primary)-independent ORFs that are not expressed above the detection limit have been characterized. These results represent the first derived from the application of DNA microarrays to either an archaeon or a hyperthermophile.
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PMID:DNA microarray analysis of the hyperthermophilic archaeon Pyrococcus furiosus: evidence for anNew type of sulfur-reducing enzyme complex. 1171 59

Entamoeba histolytica is a protozoan parasite that causes colitis and liver abscesses. Several Entamoeba species and strains with differing levels of virulence have been identified. E. histolytica HM-1:IMSS is a virulent strain, E. histolytica Rahman is a nonvirulent strain, and Entamoeba dispar is a nonvirulent species. We used an E. histolytica DNA microarray consisting of 2,110 genes to assess the transcriptional differences between these species/strains with the goal of identifying genes whose expression correlated with a virulence phenotype. We found 415 genes expressed at lower levels in E. dispar and 32 genes with lower expression in E. histolytica Rahman than in E. histolytica HM-1:IMSS. Overall, 29 genes had decreased expression in both the nonvirulent species/strains than the virulent E. histolytica HM-1:IMSS. Interestingly, a number of genes with potential roles in stress response and virulence had decreased expression in either one or both nonvirulent Entamoeba species/strains. These included genes encoding Fe hydrogenase (9.m00419), peroxiredoxin (176.m00112), type A flavoprotein (6.m00467), lysozyme (6.m00454), sphingomyelinase C (29.m00231), and a hypothetical protein with homology to both a Plasmodium sporozoite threonine-asparagine-rich protein (STARP) and a streptococcal hemagglutinin (238.m00054). The function of these genes in Entamoeba and their specific roles in parasite virulence need to be determined. We also found that a number of the non-long-terminal-repeat retrotransposons (EhLINEs and EhSINEs), which have been shown to modulate gene expression and genomic evolution, had lower expression in the nonvirulent species/strains than in E. histolytica HM-1:IMSS. Our results, identifying expression profiles and patterns indicative of a virulence phenotype, may be useful in characterizing the transcriptional framework of virulence.
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PMID:Identification of differentially expressed genes in virulent and nonvirulent Entamoeba species: potential implications for amebic pathogenesis. 1636 89


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