Peroxidase and peroxidase-oxidase activities of isolated human myeloperoxidases.
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ABSTRACT:
SUBMITTER: Svensson BE
PROVIDER: S-EPMC1147764 | biostudies-other | 1987 Mar
REPOSITORIES: biostudies-other
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Project description:Pyranose 2-oxidase (POx) has long been accredited a physiological role in lignin degradation, but evidence to provide insights into the biochemical mechanisms and interactions is insufficient. There are ample data in the literature on the oxidase and dehydrogenase activities of POx, yet the biological relevance of this duality could not be established conclusively. Here we present a comprehensive biochemical and phylogenetic characterization of a novel pyranose 2-oxidase from the actinomycetous bacterium Kitasatospora aureofaciens (KaPOx) as well as a possible biomolecular synergism of this enzyme with peroxidases using phenolic model substrates in vitro A phylogenetic analysis of both fungal and bacterial putative POx-encoding sequences revealed their close evolutionary relationship and supports a late horizontal gene transfer of ancestral POx sequences. We successfully expressed and characterized a novel bacterial POx gene from K. aureofaciens, one of the putative POx genes closely related to well-known fungal POx genes. Its biochemical characteristics comply with most of the classical hallmarks of known fungal pyranose 2-oxidases, i.e., reactivity with a range of different monosaccharides as electron donors as well as activity with oxygen, various quinones, and complexed metal ions as electron acceptors. Thus, KaPOx shows the pronounced duality of oxidase and dehydrogenase similar to that of fungal POx. We further performed efficient redox cycling of aromatic lignin model compounds between KaPOx and manganese peroxidase (MnP). In addition, we found a Mn(III) reduction activity in KaPOx, which, in combination with its ability to provide H2O2, implies this and potentially other POx as complementary enzymatic tools for oxidative lignin degradation by specialized peroxidases.IMPORTANCE Establishment of a mechanistic synergism between pyranose oxidase and (manganese) peroxidases represents a vital step in the course of elucidating microbial lignin degradation. Here, the comprehensive characterization of a bacterial pyranose 2-oxidase from Kitasatospora aureofaciens is of particular interest for several reasons. First, the phylogenetic analysis of putative pyranose oxidase genes reveals a widespread occurrence of highly similar enzymes in bacteria. Still, there is only a single report on a bacterial pyranose oxidase, stressing the need of closing this gap in the scientific literature. In addition, the relatively small K. aureofaciens proteome supposedly supplies a limited set of enzymatic functions to realize lignocellulosic biomass degradation. Both enzyme and organism therefore present a viable model to study the mechanisms of bacterial lignin decomposition, elucidate physiologically relevant interactions with specialized peroxidases, and potentially realize biotechnological applications.
| S-EPMC6581175 | biostudies-literature
Project description:Background: Methylxanthines including caffeine and theobromine are widely consumed compounds and were recently shown to interact with bovine copper-containing amine oxidase. To the best of our knowledge, no direct demonstration of any interplay between these phytochemicals and human primary amine oxidase (PrAO) has been reported to date. We took advantage of the coexistence of PrAO and monoamine oxidase (MAO) activities in human subcutaneous adipose tissue (hScAT) to test the interaction between several methylxanthines and these enzymes, which are involved in many key pathophysiological processes. Methods: Benzylamine, methylamine, and tyramine were used as substrates for PrAO and MAO in homogenates of subcutaneous adipose depots obtained from overweight women undergoing plastic surgery. Methylxanthines were tested as substrates or inhibitors by fluorimetric determination of hydrogen peroxide, an end-product of amine oxidation. Results: Semicarbazide-sensitive PrAO activity was inhibited by theobromine, caffeine, and isobutylmethylxanthine (IBMX) while theophylline, paraxanthine, and 7-methylxanthine had little effect. Theobromine inhibited PrAO activity by 54% at 2.5 mM. Overall, the relationship between methylxanthine structure and the degree of inhibition was similar to that seen with bovine PrAO, although higher concentrations (mM) were required for inhibition. Theobromine also inhibited oxidation of tyramine by MAO, at the limits of its solubility in a DMSO vehicle. At doses higher than 12 % v/v, DMSO impaired MAO activity. MAO was also inhibited by millimolar doses of IBMX, caffeine and by other methylxanthines to a lesser extent. Conclusions: This preclinical study extrapolates previous findings with bovine PrAO to human tissues. Given that PrAO is a potential target for anti-inflammatory drugs, it indicates that alongside phosphodiesterase inhibition and adenosine receptor antagonism, PrAO and MAO inhibition could contribute to the health benefits of methylxanthines, especially their anti-inflammatory effects.
| S-EPMC7235778 | biostudies-literature
Project description:Mammalian thioredoxin reductase (TrxR) is an NADPH-dependent homodimer with three redox-active centers per subunit: a FAD, an N-terminal domain dithiol (Cys(59)/Cys(64)), and a C-terminal cysteine/selenocysteine motif (Cys(497)/Sec(498)). TrxR has multiple roles in antioxidant defense. Opposing these functions, it may also assume a pro-oxidant role under some conditions. In the absence of its main electron-accepting substrates (e.g. thioredoxin), wild-type TrxR generates superoxide (O ), which was here detected and quantified by ESR spin trapping with 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO). The peroxidase activity of wild-type TrxR efficiently converted the O adduct (DEPMPO/HOO(*)) to the hydroxyl radical adduct (DEPMPO/HO(*)). This peroxidase activity was Sec-dependent, although multiple mutants lacking Sec could still generate O . Variants of TrxR with C59S and/or C64S mutations displayed markedly reduced inherent NADPH oxidase activity, suggesting that the Cys(59)/Cys(64) dithiol is required for O generation and that O is not derived directly from the FAD. Mutations in the Cys(59)/Cys(64) dithiol also blocked the peroxidase and disulfide reductase activities presumably because of an inability to reduce the Cys(497)/Sec(498) active site. Although the bulk of the DEPMPO/HO(*) signal generated by wild-type TrxR was due to its combined NADPH oxidase and Sec-dependent peroxidase activities, additional experiments showed that some free HO(*) could be generated by the enzyme in an H(2)O(2)-dependent and Sec-independent manner. The direct NADPH oxidase and peroxidase activities of TrxR characterized here give insights into the full catalytic potential of this enzyme and may have biological consequences beyond those solely related to its reduction of thioredoxin.
| S-EPMC2898413 | biostudies-literature
Project description:We have carried out an analysis of whether blood IgG antibodies can protect humans from oxidative stress by oxidizing different harmful compounds. A somewhat unexpected result was obtained. We show here for the first time that healthy human sera IgGs with the peroxidase (in the presence H2O2) efficiently oxidize different compounds: 3,3'-diaminobenzidine (1; DAB), 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (2; ATBS), o-phenylenediamine (3; OPD), homovanillic acid (4; HVA), ?-naphthol (5), 5-aminosalicylic acid (6; 5-ASA) and 3-amino-9-ethylcarbazole (7; AEC), but seven of nine IgG preparations from different volunteers cannot oxidize p-hydroquinone (8: pHQ). The average apparent kcat values in the H2O2-dependent oxidation by human IgGs decreased in the following order (min-1): ATBS (73.7)???DAB (66.3)?>?AEC (38.0)???HVA (19.8)????-naphthol (8.6)?>?OPD (0.62)???5-ASA (0.48)?>?pHQ (0.24). In the absence of H2O2 (oxidoreductase activity), the relative average kcat values decreased in the following order (min-1): DAB (52.1)???ATBS (50.5)?>?OPD (0.25). The peroxidase average activity of human IgGs was higher than the oxidoreductase one: 1.2-, 1.5- and 2.5-fold for DAB, ATBS and OPD, respectively. It should be assumed that antibodies can oxidize in addition to the large number of other different compounds analysed by us. As a whole, the specific wide repertoire of polyclonal human IgGs oxidizing various compounds could play an important role in protecting humans from oxidative stress and serve as an additional natural system destroying H2O2 and different toxic mutagenic and carcinogenic compounds.
| S-EPMC5792901 | biostudies-literature
Project description:The essential role of human dual oxidase 2 (hDUOX2) in thyroid hormone biosynthesis defines this member of the NOX/DUOX family, whose absence due to mutation has been directly related to disease, specifically hypothyroidism. Both human DUOX isoforms, hDUOX1 and hDUOX2, are expressed in thyroid tissue; however, hDUOX1 cannot compensate for inactivation of hDUOX2, suggesting that each enzyme is differentially regulated and/or functions in a unique manner. In efforts to uncover relevant structural and functional differences we have expressed and purified the peroxidase domain of hDUOX2(1-599) for direct comparison with the previously studied hDUOX1(1-593). As was shown for hDUOX1, the truncated hDUOX2 domain purifies without a bound heme co-factor and displays no peroxidase activity. However, hDUOX2(1-599) displays greater stability than hDUOX1(1-593). Surprisingly, upon titration with heme, both isoforms bind heme with a low micromolar affinity, demonstrating that they retain a heme binding site. A conformational difference in the full-length protein and/or a protein-protein interaction may be required to increase the heme binding affinity.
| S-EPMC3139011 | biostudies-literature
Project description:In this work, we investigated the oxidative modification of histidine residues induced by peroxidase and thiol oxidase activities of bovine copper-zinc superoxide dismutase (Cu-ZnSOD) using NMR and pulse EPR spectroscopy. 1D NMR and 2D-NOESY were used to determine the oxidative damage at the Zn(II) and Cu(II) active sites as well as at distant histidines. Results indicate that during treatment of SOD with hydrogen peroxide (H(2)O(2)) or cysteine in the absence of bicarbonate anion (HCO(3)(-)), both exchangeable and nonexchangeable protons were affected. Both His-44 and His-46 in the Cu(II) active site were oxidized based on the disappearance of NOESY cross-peaks between CH and NH resonances of the imidazole rings. In the Zn(II) site, only His-69, which is closer to His-44, was oxidatively modified. However, addition of HCO(3)(-) protected the active site His residues. Instead, resonances assigned to the His-41 residue, 11 Å away from the Cu(II) site, were completely abolished during both HCO(3)(-)-stimulated peroxidase activity and thiol oxidase activity in the presence of HCO(3)(-) . Additionally, ESEEM/HYSCORE and ENDOR studies of SOD treated with peroxide/Cys in the absence of HCO(3)(-) revealed that hyperfine couplings to the distal and directly coordinated nitrogens of the His-44 and His-46 ligands at the Cu(II) active site were modified. In the presence of HCO(3)(-), these modifications were absent. HCO(3)(-)-mediated, selective oxidative modification of histidines in SOD may be relevant to understanding the molecular mechanism of SOD peroxidase and thiol oxidase activities.
| S-EPMC3501376 | biostudies-literature
Project description:Among 276 herbal extracts, a methanol extract of Castanopsis cuspidata var. sieboldii stems was selected as an experimental source for novel acetylcholinesterase (AChE) inhibitors. Five compounds were isolated from the extract by activity-guided screening, and their inhibitory activities against butyrylcholinesterase (BChE), monoamine oxidases (MAOs), and β-site amyloid precursor protein cleaving enzyme 1 (BACE-1) were also evaluated. Of these compounds, 4'-O-(α-L-rhamnopyranosyl)-3,3',4-tri-O-methylellagic acid (3) and 3,3',4-tri-O-methylellagic acid (4) effectively inhibited AChE with IC50 values of 10.1 and 10.7 µM, respectively. Ellagic acid (5) inhibited AChE (IC50 = 41.7 µM) less than 3 and 4. In addition, 3 effectively inhibited MAO-B (IC50 = 7.27 µM) followed by 5 (IC50 = 9.21 µM). All five compounds weakly inhibited BChE and BACE-1. Compounds 3, 4, and 5 reversibly and competitively inhibited AChE, and were slightly or non-toxic to MDCK cells. The binding energies of 3 and 4 (- 8.5 and - 9.2 kcal/mol, respectively) for AChE were greater than that of 5 (- 8.3 kcal/mol), and 3 and 4 formed a hydrogen bond with Tyr124 in AChE. These results suggest 3 is a dual-targeting inhibitor of AChE and MAO-B, and that these compounds should be viewed as potential therapeutics for the treatment of Alzheimer's disease.
| S-EPMC8260592 | biostudies-literature
Project description:The seven members of the NOX/DUOX family are responsible for generation of the superoxide and H(2)O(2) required for a variety of host defense and cell signaling functions in nonphagocytic cells. Two members, the dual oxidase isozymes DUOX1 and DUOX2, share a structurally unique feature: an N-terminal peroxidase-like domain. Despite sequence similarity to the mammalian peroxidases, the absence of key active site residues makes their binding of heme and their catalytic function uncertain. To explore this domain we have expressed in a baculovirus system and purified the Caenorhabditis elegans (CeDUOX1(1-589)) and human (hDUOX1(1-593)) DUOX1 "peroxidase" domains. Evaluation of these proteins demonstrated that the isolated hDUOX1(1-593) does not bind heme and has no intrinsic peroxidase activity. In contrast, CeDUOX1(1-589) binds heme covalently, exhibits a modest peroxidase activity, but does not oxidize bromide ion. Surprisingly, the heme appears to have two covalent links to the protein despite the absence of a second conserved carboxyl group in the active site. Although the N-terminal dual oxidase motif has been proposed to directly convert superoxide to H(2)O(2), neither DUOX1 domain demonstrated significant superoxide dismutase activity. These results strengthen the in vivo conclusion that the CeDUOX1 protein supports controlled peroxidative polymerization of tyrosine residues and indicate that the hDUOX1 protein either has a unique function or must interact with other protein factors to express its catalytic activity.
| S-EPMC2707201 | biostudies-literature
Project description:A set of bifunctional oxidase-peroxidases has been prepared by fusing four distinct oxidases to a peroxidase. Although such fusion enzymes have not been observed in nature, they could be expressed and purified in good yields. Characterization revealed that the artificial enzymes retained the capability to bind the two required cofactors and were catalytically active as oxidase and peroxidase. Peroxidase fusions of alditol oxidase and chitooligosaccharide oxidase could be used for the selective detection of xylitol and cellobiose with a detection limit in the low-micromolar range. The peroxidase fusions of eugenol oxidase and 5-hydroxymethylfurfural oxidase could be used for dioxygen-driven, one-pot, two-step cascade reactions to convert vanillyl alcohol into divanillin and eugenol into lignin oligomers. The designed oxidase-peroxidase fusions represent attractive biocatalysts that allow efficient biocatalytic cascade oxidations that only require molecular oxygen as an oxidant.
| S-EPMC5708271 | biostudies-literature
Project description:Extracellular matrices in diverse biological systems are cross-linked by dityrosine covalent bonds catalyzed by the peroxidase/oxidase system. We show that a peroxidase, secreted by the Anopheles gambiae midgut, and dual oxidase form a dityrosine network that decreases gut permeability to immune elicitors. This network protects the microbiota by preventing activation of epithelial immunity. It also provides a suitable environment for malaria parasites to develop within the midgut lumen without inducing nitric oxide synthase expression. Disruption of this barrier results in strong and effective pathogen-specific immune responses.
| S-EPMC3510679 | biostudies-literature