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)

By crossed immunoelectrophoresis with antibodies against the NAD-linked hydrogenase the presence of three hydrogenase protein species was demonstrated in crude extracts of Alcaligenes eutrophus H16. Protein 1 (antigen 1) exhibited NAD-reducing activity and was shown to be identical with the native heterotetrameric enzyme. Protein 2 (antigen 2) was catalytically inactive in the antibody-precipitated form and corresponded to the beta subunit (56 kDa) of the holoenzyme. Protein 3 (antigen 3) was serologically distinct from antigen 2 and catalyzed NADH-oxidizing (diaphorase) activity, suggesting that it either consists of the alpha peptide or of the alpha and gamma subunits of the diaphorase dimer. Tandem immunoelectrophoresis revealed that antigen 2 was the predominant protein species in cells cultivated under nickel deficiency. Low concentrations of the diaphorase-active antigen 3 were also detected under these conditions. Extracts from mutants defective in the catalytic activity of NAD-reducing hydrogenase still contained the four polypeptides. This was shown by immunodiffusion and immunoblotting with antibodies raised against the individual subunits. However, as observed with nickel-deficient cells, no complete tetrameric protein could be identified, and the dominant subunit species (70-80%) was the beta peptide.
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PMID:Identification of distinct NAD-linked hydrogenase protein species in mutants and nickel-deficient wild-type cells of Alcaligenes eutrophus H16. 211 62

Highly-purified bidirectional hydrogenase (hydrogenase 1) of Clostridium pasteurianum could rapidly reduce several 2-, 4- and 5-nitroimidazole compounds via an electron carrier-coupled mechanism. Hydrogenase 1 was also shown to reduce a 2-nitroimidazole (misonidazole) and a 4-nitroimidazole in the presence of its required electron carriers including ferredoxin, the flavin coenzymes FAD and FMN, and the low potential electron carrier dyes methyl- and benzyl-viologen. No drug reduction by hydrogenase 1 occurred when any one of these electron carriers was replaced by nicotinamide electron carriers (NAD and NADP), or was omitted from the reaction mixture. The rates of reduction of the nitroimidazole compounds correlated with their one electron reduction potentials at pH 7(E7(1)); the higher the drug's E7(1), the faster its rate of reduction by the enzyme. Reduction rates for the drugs did not correlate with the antibacterial activity of these compounds against C. pasteurianum, suggesting that other factors are also important in determining the antimicrobial potencies of nitroimidazoles.
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PMID:Reduction of 2-, 4- and 5-nitroimidazole drugs by hydrogenase 1 in Clostridium pasteurianum. 218 Aug 90

The genes hoxF, -U, -Y, and -H which encode the four subunit polypeptides alpha, gamma, delta, and beta of the NAD-reducing hydrogenase (HoxS) of Alcaligenes eutrophus H16, were cloned, expressed in Pseudomonas facilis, and sequenced. On the basis of the nucleotide sequence, the predicted amino acid sequences, and the N-terminal amino acid sequences, it was concluded that the structural genes are tightly linked and presumably organized as an operon, denoted hoxS. Two pairs of -24 and -12 consensus sequences resembling RpoN-activatable promoters lie upstream of hoxF, the first of the four genes. Primer extension experiments indicate that the second promoter is responsible for hoxS transcription. hoxF and hoxU code for the flavin-containing dimer (alpha and gamma subunits) of HoxS which exhibits NADH:oxidoreductase activity. A putative flavin-binding region is discussed. The 26.0-kilodalton (kDa) gamma subunit contains two cysteine clusters which may participate in the coordination of two [4F3-4S]centers. The genes hoxY and hoxH code for the small 22.9-kDa delta subunit and the nickel-containing 54.8-kDa beta subunit, respectively, of the hydrogenase dimer of HoxS. The latter dimer exhibits several conserved regions found in all nickel-containing hydrogenases. The roles of these regions in coordinating iron and nickel are discussed. Although the deduced amino acid sequences of the delta and beta subunits share some conserved regions with the corresponding polypeptides of other [NiFe] hydrogenases, the overall amino acid homology is marginal. Nevertheless, significant sequence homology (35%) to the corresponding polypeptides of the soluble methylviologen-reducing hydrogenase of Methanobacterium thermoautotrophicum was found. Unlike the small subunits of the membrane-bound and soluble periplasmic hydrogenases, the HoxS protein does not appear to be synthesized with an N-terminal leader peptide.
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PMID:Cloning and nucleotide sequences of the genes for the subunits of NAD-reducing hydrogenase of Alcaligenes eutrophus H16. 218 45

The regeneration of nicotinamide-adenine dinucleotide (reduced form, NADH) by the reaction of NAD with hydrogen gas was carried out in the presence of the hydrogenase from Alcaligenes eutrophus. And the formations of alcohol, CO2, and 6-phospho-gluconate were observed by a combination of the above system and corresponding dehydrogenases. NADH was regenerated by hydrogen gas with the hydrogenase and recycled in these reactions.
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PMID:Regeneration of NADH and ketone hydrogenation by hydrogen with the combination of hydrogenase and alcohol dehydrogenase. Scientific note. 219 25

Polyclonal, monospecific antibodies were produced against the two subunits (Mr 62,000, and Mr 31,000), isolated from the membrane-bound hydrogenase of Alcaligenes eutrophus H16. The antibodies (IgG fractions) were purified from crude sera by Protein A-Sepharose CL-4B chromatography. By double immunodiffusion assays and tandem-crossed immunoelectrophoresis the large and the small subunit were demonstrated not to be immunologically related. Immunological comparison of these subunits with the four non-identical subunits (Mr 63,000, 56,000, 30,000 and 26,000) of the NAD-linked, soluble hydrogenase from A. eutrophus H16 showed that the subunits of the membrane-bound hydrogenase did not cross-react with any of the antibodies raised against the four subunits of the NAD-linked enzyme and that, vice versa, none of these four subunits cross-reacted with antibodies raised against the two subunits of the membrane-bound hydrogenase. This means that A. eutrophus H16 contains altogether six non-identical immunologically unrelated hydrogenase polypeptides. The membrane-bound hydrogenases were isolated and purified from various aerobic H2-oxidizing bacteria: A. eutrophus H16, A. eutrophus type strain, A. eutrophus CH34, A. eutrophus Z1, A. hydrogenophilus, Paracoccus denitrificans and strain Cd2/01. All these proteins resembled each other and each consisted of two non-identical polypeptides. A complete separation of these subunits was achieved at high-yield by preparative FPLC gel filtration on three Superose 12 columns connected in series, using SDS and DTT-containing sodium phosphate buffer (pH 7.0). The small subunits of these enzymes turned out to be immunologically closely related to each other; they were either identical or almost identical. The large subunits were also related, but less pronounced. Only the large subunits from Z1 and type strain reacted fully identical with the H16 subunit. Of the two isolated, homogeneous subunits of the membrane-bound hydrogenase from A. eutrophus H16, the amino acid compositions and the NH2-terminal sequences have been determined. The results confirmed the diversity of the large and the small subunit. Furthermore, for comparison also the NH2-terminal sequences of the two subunits from the hydrogenase of A. eutrophus CH34 have been analysed.
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PMID:Immunological comparison of subunits isolated from various hydrogenases of aerobic hydrogen bacteria. 249 16

The cytoplasmic, NAD-linked hydrogenase of the Gram-positive hydrogen-oxidizing bacterium Nocardia opaca 1b was compared with the analogous enzyme isolated from the Gram-negative bacterium Alcaligenes eutrophus H16. The hydrogenase of N. opaca 1b was purified by a new procedure applying chromatography on phenyl-Sepharose and DEAE-Sephacel with two columns in series. A homogeneous enzyme preparation with a specific activity of 74 mumol H2 oxidized.min-1.mg protein-1 and a yield of 32% was isolated. The A. eutrophus enzyme was purified as previously published. Both enzymes are tetrameric proteins composed of four non-identical subunits (alpha, beta, gamma, delta). The four subunits of both of these enzymes were separated and isolated as single polypeptides by preparative polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Immunological comparison of the four subunits of the Nocardia hydrogenase with those of the Alcaligenes enzyme showed that the alpha, beta, gamma, and delta subunits of one organism were serologically related to the analogous subunits of the other organism. Among themselves, the four subunits do not have any serological relationship. The eight individual polypeptides were also compared with respect to the NH2-terminal amino acid sequences determined by automated Edman degradation and to the amino acid compositions. Strong sequence similarities exist between the analogous subunits isolated from the two bacteria. Within the established N-terminal sequences the similarities between both alpha, beta, gamma and delta subunits amount to 63%, 79%, 80% and 65%, respectively. No similarities exist between the different, non-analogous subunits alpha, beta, gamma and delta.
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PMID:Comparison of the NH2-terminal amino acid sequences of the four non-identical subunits of the NAD-linked hydrogenases from Nocardia opaca 1b and Alcaligenes eutrophus H16. 249 82

The effect of redox potential on the catalytic activities of the soluble hydrogenase from the hydrogen bacterium Alcaligenes eutrophus Z1 was studied. Several transitions were observed on the enzyme catalytic activity vs potential profiles. The coenzyme-dependent activities of the hydrogenase, its diaphorase activity and activity toward NAD, are controlled by the Em -300 mV, while the process of hydrogen evolution from reduced methyl viologen is governed by the midpoint redox potential of -435 mV. This value of Em was independent of pH in the range 5 to 8. The redox potential of the medium appears to be one of the major factors determining the hydrogenase activation, inactivation, and catalytic properties. It is suggested that a change in the redox state of the enzyme electron transport chain is followed by structural rearrangements within the protein affecting both the hydrogenase catalytic activity and stability. The probable mechanism of enzyme activity regulation is discussed.
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PMID:Effect of redox potential on the catalytic properties of the NAD-dependent hydrogenase from Alcaligenes eutrophus Z1. 253 63

A formal kinetic treatment of the autocatalytic activation cycle of the NAD-dependent hydrogenase from Alcaligenes eutrophus Z1 is presented. The value for the enzyme first-order activation rate constant is estimated to be (2.0 +/- 0.6) s-1 (pH 7.8, 25 degrees C). The effect of the redox potential on the activation properties of the NAD-dependent hydrogenase is studied. Hydrogenase activation is controlled by a midpoint redox potential of approximately -100 mV (pH 7.8). Once activated the enzyme is not immediately transformed back into an inactive state on rapid reoxidation and is able to preserve its catalytic properties for at least 3-4 h of intense oxigenation. Several lines of evidence show that the reductive activation of the NAD-dependent hydrogenase is accompanied by a structural reorganization of the protein. A possible origin of the -100 mV transition is discussed. A model for the activation process of the NAD-dependent hydrogenase is suggested.
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PMID:Effect of redox potential on the activation of the NAD-dependent hydrogenase from Alcaligenes eutrophus Z1. 264 85

A novel inactivation mechanism of the NAD-dependent hydrogenase from Alcaligenes eutrophus Z1 comprising redox-dependent steps is described. The model of the hydrogenase inactivation process is proposed which implies that the enzyme may exist in several forms which differ in their stability and spectral properties. One of these forms, existing within a limited (approximately -200 +/- 30 mV) potential range, undergoes a rapid and irreversible inactivation. The dissociation of the FMN prosthetic group from the apohydrogenase appears to be the main reason for the enzyme inactivation. The rationale for the enzyme stabilization under real operational conditions based on the chemical modification of the hydrogenase molecule is suggested.
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PMID:Redox-dependent inactivation of the NAD-dependent hydrogenase from Alcaligenes eutrophus Z1. 264 86

Evidence is presented for the existence of a relatively high-potential regulatory centre in the NAD-dependent hydrogenase from the hydrogen oxidizing bacterium Alcaligenes eutrophus Z1. Reduction of the hydrogenase to the redox potentials lower than -100 mV converts the enzyme into a catalytically active state that is remarkably stable to oxidants. Once activated, the enzyme does not loose its activity on intensive oxygenation for at least 3 hours. A novel hydrogenase ESR signal with a wide temperature optimum and a approximately -100 mV midpoint redox potential was detected. We suggest that the reduction of this redox centre trigger conformational changes in the inactive oxidized enzyme molecule, thus reorganizing the latter into the active one.
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PMID:NAD-dependent hydrogenase from Alcaligenes eutrophus Z1: does it have a regulatory centre? 302 9


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