Myeloperoxidase amplified high glucose-induced endothelial dysfunction in vasculature: Role of NADPH oxidase and hypochlorous acid.

Science.gov (United States)

Tian, Rong; Ding, Yun; Peng, Yi-Yuan; Lu, Naihao

2017-03-11

Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-derived reactive oxygen species (ROS) such as superoxide and hydrogen peroxide (H 2 O 2 ), have emerged as important molecules in the pathogenesis of diabetic endothelial dysfunction. Additionally, neutrophils-derived myeloperoxidase (MPO) and MPO-catalyzed hypochlorous acid (HOCl) play important roles in the vascular injury. However, it is unknown whether MPO can use vascular-derived ROS to induce diabetic endothelial dysfunction. In the present study, we demonstrated that NADPH oxidase was the main source of ROS formation in high glucose-cultured human umbilical vein endothelial cells (HUVECs), and played a critical role in high glucose-induced endothelial dysfunction such as cell apoptosis, loss of cell viability and reduction of nitric oxide (NO). However, the addition of MPO could amplify the high glucose-induced endothelial dysfunction which was inhibited by the presence of apocynin (NADPH oxidase inhibitor), catalase (H 2 O 2 scavenger), or methionine (HOCl scavenger), demonstrating the contribution of NADPH oxidase-H 2 O 2 -MPO-HOCl pathway in the MPO/high glucose-induced vascular injury. In high glucose-incubated rat aortas, MPO also exacerbated the NADPH oxidase-induced impairment of endothelium-dependent relaxation. Consistent with these in vitro data, in diabetic rat aortas, both MPO expresion and NADPH oxidase activity were increased while the endothelial function was simultaneously impaired. The results suggested that vascular-bound MPO could amplify high glucose-induced vascular injury in diabetes. MPO-NADPH oxidase-HOCl may represent an important pathogenic pathway in diabetic vascular diseases. Copyright © 2017 Elsevier Inc. All rights reserved.

Towards printed perovskite solar cells with cuprous oxide hole transporting layers

DEFF Research Database (Denmark)

Wang, Yan; Xia, Zhonggao; Liang, Jun

2015-01-01

Solution-processed p-type metal oxide materials have shown great promise in improving the stability of perovskite-based solar cells and offering the feasibility for a low cost printing fabrication process. Herein, we performed a device modeling study on planar perovskite solar cells with cuprous...... oxide (Cu2O) hole transporting layers (HTLs) by using a solar cell simulation program, wxAMPS. The performance of a Cu2O/perovskite solar cell was correlated to the material properties of the Cu2O HTL, such as thickness, carrier mobility, mid-gap defect, and doping...

Light-Induced Reduction of Cuprous Oxide in an Environmental Transmission Electron Microscope

DEFF Research Database (Denmark)

Cavalca, Filippo Carlo; Laursen, Anders Bo; Wagner, Jakob Birkedal

2013-01-01

Photocatalysts for solar fuel production are subject to intensive investigation as they constitute one viable route for solar energy harvesting. Cuprous oxide (Cu2O) is a working photocatalyst for hydrogen evolution but it photocorrodes upon light illumination in an aqueous environment....... Environmental transmission electron microscopy (ETEM) makes it possible to obtain insight into the local structure, composition and reactivity of catalysts in their working environment, which is of fundamental interest for sustainable energy research and is essential for further material optimization. Herein...

Copper-doped silica cuprous sulfate: A highly efficient heterogeneous nano-catalyst for one-pot three-component synthesis of 1-H-2-substituted benzimidazoles from 2-bromoanilines, aldehydes, and [bmim]N3

Directory of Open Access Journals (Sweden)

Somayeh Behrouz

2018-03-01

Full Text Available A facile and highly efficient one-pot three-component synthesis of 1-H-2-substituted benzimidazole derivatives from readily available substrates catalyzed by copper-doped silica cuprous sulfate (CDSCS is described. In this method, treatment of diverse 2-bromoanilines, aldehydes, and [bmim]N3 in DMF at 110 °C in the presence of CDSCS as a highly efficient heterogeneous nano-catalyst affords the corresponding 1-H-2-substituted benzimidazoles in good to excellent yields. The CDSCS is an inexpensive and stable nano-catalyst that could be simply prepared, recovered and reused for many consecutive reaction runs without significant loss of its activity.

Preparing cuprous oxide nanomaterials by electrochemical method for non-enzymatic glucose biosensor

Science.gov (United States)

Nguyen, Thu-Thuy; Huy, Bui The; Hwang, Seo-Young; Vuong, Nguyen Minh; Pham, Quoc-Thai; Nghia, Nguyen Ngoc; Kirtland, Aaron; Lee, Yong-Ill

2018-05-01

Cuprous oxide (Cu2O) nanostructure has been synthesized using an electrochemical method with a two-electrode system. Cu foils were used as electrodes and NH2(OH) was utilized as the reducing agent. The effects of pH and applied voltages on the morphology of the product were investigated. The morphology and optical properties of Cu2O particles were characterized using scanning electron microscopy, x-ray diffraction, and diffuse reflectance spectra. The synthesized Cu2O nanostructures that formed in the vicinity of the anode at 2 V and pH = 11 showed high uniform distribution, small size, and good electrochemical sensing. These Cu2O nanoparticles were coated on an Indium tin oxide substrate and applied to detect non-enzyme glucose as excellent biosensors. The non-enzyme glucose biosensors exhibited good performance with high response, good selectivity, wide linear detection range, and a low detection limit at 0.4 μM. Synthesized Cu2O nanostructures are potential materials for a non-enzyme glucose biosensor.

Microwave—enhanced Mannich Condensation of Terminal Alkynes,Primary Amines with Paraformaldehyde on cuprous Iodide Doped Alumina under Solvent Free Conditions

Institute of Scientific and Technical Information of China (English)

王磊; 李品华

2003-01-01

A microwave-enhanced,solventless Mannich condensation of terminal alkynes,primary amines with paraformaldehyde on cuprous iodide doped alumina has been investigated.The structures of products depend on the ratio of alkyne to amine and paraformaldehyde.

Cuprous oxide nanoparticles selectively induce apoptosis of tumor cells

Science.gov (United States)

Wang, Ye; Zi, Xiao-Yuan; Su, Juan; Zhang, Hong-Xia; Zhang, Xin-Rong; Zhu, Hai-Ying; Li, Jian-Xiu; Yin, Meng; Yang, Feng; Hu, Yi-Ping

2012-01-01

In the rapid development of nanoscience and nanotechnology, many researchers have discovered that metal oxide nanoparticles have very useful pharmacological effects. Cuprous oxide nanoparticles (CONPs) can selectively induce apoptosis and suppress the proliferation of tumor cells, showing great potential as a clinical cancer therapy. Treatment with CONPs caused a G1/G0 cell cycle arrest in tumor cells. Furthermore, CONPs enclosed in vesicles entered, or were taken up by mitochondria, which damaged their membranes, thereby inducing apoptosis. CONPs can also produce reactive oxygen species (ROS) and initiate lipid peroxidation of the liposomal membrane, thereby regulating many signaling pathways and influencing the vital movements of cells. Our results demonstrate that CONPs have selective cytotoxicity towards tumor cells, and indicate that CONPs might be a potential nanomedicine for cancer therapy. PMID:22679374

Cuprous oxide nanoparticles dispersed on reduced graphene oxide as an efficient electrocatalyst for oxygen reduction reaction.

Science.gov (United States)

Yan, Xiao-Yan; Tong, Xi-Li; Zhang, Yue-Fei; Han, Xiao-Dong; Wang, Ying-Yong; Jin, Guo-Qiang; Qin, Yong; Guo, Xiang-Yun

2012-02-11

Cuprous oxide (Cu(2)O) nanoparticles dispersed on reduced graphene oxide (RGO) were prepared by reducing copper acetate supported on graphite oxide using diethylene glycol as both solvent and reducing agent. The Cu(2)O/RGO composite exhibits excellent catalytic activity and remarkable tolerance to methanol and CO in the oxygen reduction reaction. This journal is © The Royal Society of Chemistry 2012

Size-selective QD@MOF core-shell nanocomposites for the highly sensitive monitoring of oxidase activities.

Science.gov (United States)

Wang, Ke; Li, Nan; Zhang, Jing; Zhang, Zhiqi; Dang, Fuquan

2017-01-15

In this work, we proposed a novel and facile method to monitor oxidase activities based on size-selective fluorescent quantum dot (QD)@metal-organic framework (MOF) core-shell nanocomposites (CSNCPs). The CSNCPs were synthesized from ZIF-8 and CdTe QDs in aqueous solution in 40min at room temperature with stirring. The prepared CdTe@ZIF-8 CSNCPs , which have excellent water dispersibility and stability, displays distinct fluorescence responses to hole scavengers of different molecular sizes (e.g., H 2 O 2 , substrate, and oxidase) due to the aperture limitation of the ZIF-8 shell. H 2 O 2 can efficiently quench the fluorescence of CdTe@ZIF-8 CSNCPs over a linearity range of 1-100nM with a detection limit of 0.29nM, whereas large molecules such as substrate and oxidase have very little effect on its fluorescence. Therefore, the highly sensitive detection of oxidase activities was achieved by monitoring the fluorescence quenching of CdTe@ZIF-8 CSNCPs by H 2 O 2 produced in the presence of substrate and oxidase, which is proportional to the oxidase activities. The linearity ranges of the uricase and glucose oxidase activity are 0.1-50U/L and 1-100U/L, respectively, and their detection limits are 0.024U/L and 0.26U/L, respectively. Therefore, the current QD@MOF CSNCPs based sensing system is a promising, widely applicable means of monitoring oxidase activities in biochemical research. Copyright © 2016 Elsevier B.V. All rights reserved.

Multicomponent exciton gas in cuprous oxide: cooling behaviour and the role of Auger decay

Science.gov (United States)

Semkat, D.; Sobkowiak, S.; Schöne, F.; Stolz, H.; Koch, Th; Fehske, H.

2017-10-01

In this paper we present a hydrodynamic model to describe the dynamics of para- and orthoexcitons in cuprous oxide at ultralow temperatures inside a stress induced potential trap. We take into account the finite lifetime of the excitons, the excitation process and exciton-phonon as well as exciton-exciton interaction. Furthermore, we model the two-body loss mechanism assuming an Auger-like effect and compare it to an alternative explanation which relies on the formation of biexcitons. We discuss in detail the influence on the numerical results and compare the predictions to experimental data.

Exploring flavin-containing carbohydrate oxidases

NARCIS (Netherlands)

Ferrari, Alessandro Renato

2017-01-01

Oxidases are enzymes capable of removing one or more electrons from their substrate and transfer them to molecular oxygen, forming hydrogen peroxide. Due to their high regio- and enantioselectivity, their use is preferred over traditional organic chemistry methods. Among the oxidases, flavoprotein

Directing the Branching Growth of Cuprous Oxide by OH- Ions

Science.gov (United States)

Chen, Kunfeng; Si, Yunfei; Xue, Dongfeng

The effect of OH- ions on the branching growth of cuprous oxide microcrystals was systematically studied by a reduction route, where copper-citrate complexes were reduced by glucose under alkaline conditions. Different copper salts including Cu(NO3)2, CuCl2, CuSO4, and Cu(Ac)2 were used in this work. The results indicate that the Cu2O branching growth habit is closely correlated to the concentration of OH- ions, which plays an important role in directing the diffusion-limited branching growth of Cu2O and influencing the reduction power of glucose. A variety of Cu2O branching patterns including 6-pod, 8-pod and 24-pod branches, have been achieved without using template and surfactant. The current method can provide a good platform for studying the growth mechanism of microcrystal branching patterns.

A highly sensitive electrochemical glucose sensor structuring with nickel hydroxide and enzyme glucose oxidase

International Nuclear Information System (INIS)

Mathew, Manjusha; Sandhyarani, N.

2013-01-01

Graphical abstract: A combination of Ni 2+ /Ni 3+ redox couple and glucose oxidase has successfully been exploited for the realization of a highly sensitive glucose sensor for the first time. -- Highlights: • A multilayered glucose biosensor with enhanced sensitivity was fabricated. • Combination of Ni 2+ /Ni 3+ redox couple and glucose oxidase has been exploited for the first time. • Exhibits a lower detection limit of 100 nM with a high sensitivity of 16,840 μA mM −1 cm −2 . • The surface shows a low Michaelis–Menten constant value of 2.4 μM. • Detailed mechanism of sensing was proposed and justified. -- Abstract: A multilayered glucose biosensor with enhanced electron transport was fabricated via the sequential electrodeposition of chitosan gold nanocomposite (CGNC) and nickel hydroxide (Ni(OH) 2 ) on a bare gold electrode and subsequent immobilization of glucose oxidase. A thin film of Ni(OH) 2 deposited on CGNC modified gold electrode serves as an electrochemical redox probe as well as a matrix for the immobilization of glucose oxidase retaining its activity. Electron transport property of CGNC has been exploited to enhance the electron transport between the analyte and electrode. Electrochemical characteristics of the biosensor were studied by cyclic voltammetry and chronoamperometry. Under optimal conditions the biosensor exhibits a linear range from 1 μM to 100 μM with a limit of detection (lod) down to 100 nM. The sensor shows a low Michaelis-Menten constant value of 2.4 μM indicates the high affinity of enzyme to the analyte points to the retained activity of enzyme after immobilization. The present glucose sensor with the high selectivity, sensitivity and stability is promising for practical clinical applications

Optimization of Glucose oxidase towards oxygen independency and high mediator activity for amperometric glucose determination in diabetes analytics

OpenAIRE

Arango Gutierrez, Erik Uwe

2015-01-01

Glucose oxidase is an oxidoreductase exhibiting a high β-D-glucose specificity and high stability which renders glucose oxidase well-suited for applications in diabetes care. Nevertheless, GOx activity is highly oxygen dependent which can lead to inaccuracies in amperometric β-D-glucose determinations. Therefore a directed evolution campaign with two rounds of random mutagenesis (SeSaM followed by epPCR), site saturation mutagenesis studies, and one simultaneous site saturation library (OmniC...

Glucose Oxidase Directly Immobilized onto Highly Porous Gold Electrodes for Sensing and Fuel Cell applications

International Nuclear Information System (INIS)

Toit, Hendrik du; Di Lorenzo, Mirella

2014-01-01

Highlights: • Electrochemical adsorption of glucose oxidase (GOx) on highly porous gold (hPG); • Rapid one-step immobilisation protocol with no use of expensive and/or harsh reagents; • Linear response to glucose in the range 50 μM -10 mM; • Lower detection limit, stable over 5 days: 25 μM. • The use of the GOx-hPG in a fuel cell lead to the peak power density of 6 μW cm −2 . - Abstract: The successful implementation of redox-enzyme electrodes in biosensors and enzymatic biofuel cells has been the subject of extensive research. For high sensitivity and high energy-conversion efficiency, the effective electron transfer at the protein-electrode interface has a key role. This is difficult to achieve in the case of glucose oxidase, due to the fact that for this enzyme the redox centre is buried inside the structure, far from any feasible electrode binding sites. This study reports, a simple and rapid methodology for the direct immobilisation of glucose oxidase into highly porous gold electrodes. When the resulting electrode was tested as glucose sensor, a Michaelis-Menten kinetic trend was observed, with a detection limit of 25 μM. The bioelectrode sensitivity, calculated against the superficial surface area of the bioelectrode, was of 22.7 ± 0.1 μA mM −1 cm −2 . This glucose oxidase electrode was also tested as an anode in a glucose/O 2 enzymatic biofuel cell, leading to a peak power density of 6 μW cm −2 at a potential of 0.2 V

Resveratrol protects vascular endothelial cells from high glucose-induced apoptosis through inhibition of NADPH oxidase activation-driven oxidative stress.

Science.gov (United States)

Chen, Feng; Qian, Li-Hua; Deng, Bo; Liu, Zhi-Min; Zhao, Ying; Le, Ying-Ying

2013-09-01

Hyperglycemia-induced oxidative stress has been implicated in diabetic vascular complications in which NADPH oxidase is a major source of reactive oxygen species (ROS) generation. Resveratrol is a naturally occurring polyphenol, which has vasoprotective effects in diabetic animal models and inhibits high glucose (HG)-induced oxidative stress in endothelial cells. We aimed to examine whether HG-induced NADPH oxidase activation and ROS production contribute to glucotoxicity to endothelial cells and the effect of resveratrol on glucotoxicity. Using a murine brain microvascular endothelial cell line bEnd3, we found that NADPH oxidase inhibitor (apocynin) and resveratrol both inhibited HG-induced endothelial cell apoptosis. HG-induced elevation of NADPH oxidase activity and production of ROS were inhibited by apocynin, suggesting that HG induces endothelial cell apoptosis through NADPH oxidase-mediated ROS production. Mechanistic studies revealed that HG upregulated NADPH oxidase subunit Nox1 but not Nox2, Nox4, and p22(phox) expression through NF-κB activation, which resulted in elevation of NADPH oxidase activity and consequent ROS production. Resveratrol prevented HG-induced endothelial cell apoptosis through inhibiting HG-induced NF-κB activation, NADPH oxidase activity elevation, and ROS production. HG induces endothelial cell apoptosis through NF-κB/NADPH oxidase/ROS pathway, which was inhibited by resveratrol. Our findings provide new potential therapeutic targets against brain vascular complications of diabetes. © 2013 John Wiley & Sons Ltd.

The use of cuprous iodide as a precipitation matrix in the radiochemical determination of 131I in milk

International Nuclear Information System (INIS)

McCurdy, D.E.; Mellor, R.A.; Lambdin, R.W.; McLain, M.E. Jr.

1980-01-01

As a result of the implementation of the As Low As is Reasonably Achievable philosophy to the nuclear power industry, recent U.S. Nuclear Regulatory Commission requirements have prompted high sensitivity radiochemical analysis for the measurement of 131 I in milk. The most recognized and commonly employed technique incorporates costly palladium iodide as the final precipitate in the radiochemical purification of the iodine chemical species. The procedure presented in this paper outlines the many advantages of using cuprous iodide as the final precipitate. These include lower cost per analysis, consistent recoveries, better precipitate matrix and good self absorption characteristics. Typical lower limit of detection values and operating characteristics obtained for high sensitivity β-γ analysis as well as gas proportional counting and a comparison of radiochemical and Ge(Li) spectrometric results for environmental samples collected during a recent Chinese weapons fallout incident are presented. (author)

Covalently bound phosphate residues in bovine milk xanthine oxidase and in glucose oxidase from Aspergillus niger: A reevaluation

Energy Technology Data Exchange (ETDEWEB)

Johnson, J.L.; Rajagopalan, K.V. (Duke Univ. Medical Center, Durham, NC (USA)); London, R.E. (National Institute of Environmental Health Science, Research Triangle Park, NC (USA))

1989-09-01

The reported presence of covalently bound phosphate residues in flavoproteins has significant implications with regard to the catalytic mechanisms and structural stability of the specific enzymes themselves and in terms of general cellular metabolic regulation. These considerations have led to a reevaluation of the presence of covalently bound phosphorus in the flavoproteins xanthine oxidase and glucose oxidase. Milk xanthine oxidase purified by a procedure that includes anion-exchange chromatography is shown to contain three phosphate residues. All three are noncovalently associated with the protein, two with the FAD cofactor, and one with the molybdenum cofactor. Results of chemical analysis and {sup 31}P NMR spectroscopy indicate that enzyme purified by this method contains no phosphoserine residues. Xanthine oxidase preparations purified by chromatography on calcium phosphate gel in place of DEAE-Sephadex yielded higher phosphate-to-protein ratios, which could be reduced to the expected values by additional purification on a folate affinity column. Highly active, highly purified preparations of glucose oxidase are shown to contain only the two phosphate residues of the FAD cofactor. The covalently bound bridging phosphate reported by others may arise in aged or degraded preparations of the enzyme but appears not to be a constituent of functional glucose oxidase. These results suggest that the presence of covalent phosphate residues in other flavoproteins should be rigorously reevaluated as well.

Covalently bound phosphate residues in bovine milk xanthine oxidase and in glucose oxidase from Aspergillus niger: A reevaluation

International Nuclear Information System (INIS)

Johnson, J.L.; Rajagopalan, K.V.; London, R.E.

1989-01-01

The reported presence of covalently bound phosphate residues in flavoproteins has significant implications with regard to the catalytic mechanisms and structural stability of the specific enzymes themselves and in terms of general cellular metabolic regulation. These considerations have led to a reevaluation of the presence of covalently bound phosphorus in the flavoproteins xanthine oxidase and glucose oxidase. Milk xanthine oxidase purified by a procedure that includes anion-exchange chromatography is shown to contain three phosphate residues. All three are noncovalently associated with the protein, two with the FAD cofactor, and one with the molybdenum cofactor. Results of chemical analysis and 31 P NMR spectroscopy indicate that enzyme purified by this method contains no phosphoserine residues. Xanthine oxidase preparations purified by chromatography on calcium phosphate gel in place of DEAE-Sephadex yielded higher phosphate-to-protein ratios, which could be reduced to the expected values by additional purification on a folate affinity column. Highly active, highly purified preparations of glucose oxidase are shown to contain only the two phosphate residues of the FAD cofactor. The covalently bound bridging phosphate reported by others may arise in aged or degraded preparations of the enzyme but appears not to be a constituent of functional glucose oxidase. These results suggest that the presence of covalent phosphate residues in other flavoproteins should be rigorously reevaluated as well

Preparation of silver-cuprous oxide/stearic acid composite coating with superhydrophobicity on copper substrate and evaluation of its friction-reducing and anticorrosion abilities

Energy Technology Data Exchange (ETDEWEB)

Li, Peipei [Key Laboratory of Ministry of Education for Special Functional Materials, Henan University, Kaifeng 475004 (China); Chen, Xinhua [College of Chemistry and Chemical Engineering, Xuchang University, Xuchang 461000 (China); Yang, Guangbin; Yu, Laigui [Key Laboratory of Ministry of Education for Special Functional Materials, Henan University, Kaifeng 475004 (China); Zhang, Pingyu, E-mail: pingyu@henu.edu.cn [Key Laboratory of Ministry of Education for Special Functional Materials, Henan University, Kaifeng 475004 (China)

2014-01-15

A simple two-step solution immersion process was combined with surface-modification by stearic acid to prepare superhydrophobic coatings on copper substrates so as to reduce friction coefficient, increase wear resistance and improve the anticorrosion ability of copper. Briefly, cuprous oxide (Cu{sub 2}O) crystal coating with uniform and compact tetrahedron structure was firstly created by immersing copper substrate in 2 mol L{sup −1} NaOH solution. As-obtained Cu{sub 2}O coating was then immersed in 0.33 mmol L{sup −1} AgNO{sub 3} solution to incorporate silver nanoparticles, followed by modification with stearic acid (denoted as SA) coating to achieve hydrophobicity. The surface morphology and chemical composition of silver-cuprous oxide/stearic acid (denoted as Ag-Cu{sub 2}O/SA) composite coating were investigated using a scanning electron microscope and an X-ray photoelectron spectroscope (XPS); and its phase structure was examined with an X-ray diffractometer (XRD). Moreover, the contact angle of water on as-prepared Ag-Cu{sub 2}O/SA composite coating was measured, and its friction-reducing and anticorrosion abilities were evaluated. It was found that as-prepared Ag-Cu{sub 2}O/SA composite coating has a water contact angle of as high as 152.4{sup o} and can provide effective friction-reducing, wear protection and anticorrosion protection for copper substrate, showing great potential for surface-modification of copper.

Identification of aldehyde oxidase 1 and aldehyde oxidase homologue 1 as dioxin-inducible genes

International Nuclear Information System (INIS)

Rivera, Steven P.; Choi, Hyun Ho; Chapman, Brett; Whitekus, Michael J.; Terao, Mineko; Garattini, Enrico; Hankinson, Oliver

2005-01-01

Aldehyde oxidases are a family of highly related molybdo-flavoenzymes acting upon a variety of compounds of industrial and medical importance. We have identified aldehyde oxidase 1 (AOX1) as a 2,3,7,8-tetrachlorodibenzo-p-dioxin (dioxin) inducible gene in the mouse hepatoma cell line Hepa-1. AOX1 mRNA levels were not increased by dioxin in mutant derivatives of the Hepa-1 cell line lacking either functional aryl hydrocarbon receptor (AHR) or aryl hydrocarbon receptor nuclear translocator (ARNT) proteins, thus demonstrating that transcriptional induction of AOX1 in response to dioxin occurs through the AHR pathway. Dioxin induction of AOX1 mRNA was also observed in mouse liver. In addition, levels of AOX1 protein as well as those of aldehyde oxidase homologue 1 (AOH1), a recently identified homolog of AOX1, were elevated in mouse liver in response to dioxin. Employing an aldehyde oxidase specific substrate, AOX1/AOH1 activity was shown to be induced by dioxin in mouse liver. This activity was inhibited by a known inhibitor of aldehyde oxidases, and eliminated by including tungstate in the mouse diet, which is known to lead to inactivation of molybdoflavoenzymes, thus confirming that the enzymatic activity was attributable to AOX1/AOH1. Our observations thus identify two additional xenobiotic metabolizing enzymes induced by dioxin

Resveratrol prevents high glucose-induced epithelial-mesenchymal transition in renal tubular epithelial cells by inhibiting NADPH oxidase/ROS/ERK pathway.

Science.gov (United States)

He, Ting; Guan, Xu; Wang, Song; Xiao, Tangli; Yang, Ke; Xu, Xinli; Wang, Junping; Zhao, Jinghong

2015-02-15

Resveratrol (RSV) is reported to have renoprotective activity against diabetic nephropathy, while the mechanisms underlying its function have not been fully elucidated. In this study, we investigate the effect and related mechanism of RSV against high glucose-induced epithelial to mesenchymal transition (EMT) in human tubular epithelial cells (HK-2). A typical EMT is induced by high glucose in HK-2 cells, accompanied by increased levels of reactive oxygen species (ROS). RSV exhibits a strong ability to inhibit high glucose-induced EMT by decreasing intracellular ROS levels via down-regulation of NADPH oxidase subunits NOX1 and NOX4. The activation of extracellular signal-regulated kinase (ERK1/2) is found to be involved in high glucose-induced EMT in HK-2 cells. RSV, like NADPH oxidase inhibitor diphenyleneiodonium, can block ERK1/2 activation induced by high glucose. Our results demonstrate that RSV is a potent agent against high glucose-induced EMT in renal tubular cells via inhibition of NADPH oxidase/ROS/ERK1/2 pathway. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Polyphenol Oxidase Enzyme and Inactivation Methods

Directory of Open Access Journals (Sweden)

Leman Yılmaz

2018-03-01

Full Text Available Polyphenol oxidase enzyme is found in vegetables and fruits, as well as in some animal organs and microorganisms. Polyphenol oxidase enzyme responsible for enzymatic browning is a group of copper proteins that catalyses the oxidation of phenolic compounds to quinones, which produce brown pigments, commonly found in fruits and vegetables. During the industrial preparation of fruits and vegetables, results of catalytic effect of polyphenol oxidase causes enzymatic browning. Enzymatic browning impairs the appearance of products containing phenolic compounds along with undesirable colour, odor and taste formation and significant loss of nutritional value of the products. This affects the acceptability of the products by the consumers and causes economic losses. In this review, some characteristics of polyphenol oxidase enzyme in different fruits and vegetables have been reviewed and information about chemical antibrowning agents, thermal applications, irradiation applications and alternative methods such as high pressure processing, pulse electric field, supercritical carbon dioxide and ultrasound applications to inactivate this enzyme has been presented.

Cuprous oxide nanoparticles selectively induce apoptosis of tumor cells

Directory of Open Access Journals (Sweden)

Wang Y

2012-05-01

Full Text Available Ye Wang,1,2,* Xiao-Yuan Zi,1,* Juan Su,1 Hong-Xia Zhang,1 Xin-Rong Zhang,3 Hai-Ying Zhu,1 Jian-Xiu Li,1 Meng Yin,3 Feng Yang,3 Yi-Ping Hu,11Department of Cell Biology, 2School of Clinical Medicine, 3Department of Pharmaceuticals, Second Military Medical University, Shanghai, People's Republic of China*Authors contributed equally.Abstract: In the rapid development of nanoscience and nanotechnology, many researchers have discovered that metal oxide nanoparticles have very useful pharmacological effects. Cuprous oxide nanoparticles (CONPs can selectively induce apoptosis and suppress the proliferation of tumor cells, showing great potential as a clinical cancer therapy. Treatment with CONPs caused a G1/G0 cell cycle arrest in tumor cells. Furthermore, CONPs enclosed in vesicles entered, or were taken up by mitochondria, which damaged their membranes, thereby inducing apoptosis. CONPs can also produce reactive oxygen species (ROS and initiate lipid peroxidation of the liposomal membrane, thereby regulating many signaling pathways and influencing the vital movements of cells. Our results demonstrate that CONPs have selective cytotoxicity towards tumor cells, and indicate that CONPs might be a potential nanomedicine for cancer therapy.Keywords: nanomedicine, selective cytotoxicity, apoptosis, cell cycle arrest, mitochondrion-targeted nanomaterials

Metformin and liraglutide ameliorate high glucose-induced oxidative stress via inhibition of PKC-NAD(P)H oxidase pathway in human aortic endothelial cells.

Science.gov (United States)

Batchuluun, Battsetseg; Inoguchi, Toyoshi; Sonoda, Noriyuki; Sasaki, Shuji; Inoue, Tomoaki; Fujimura, Yoshinori; Miura, Daisuke; Takayanagi, Ryoichi

2014-01-01

Metformin and glucagon like peptide-1 (GLP-1) prevent diabetic cardiovascular complications and atherosclerosis. However, the direct effects on hyperglycemia-induced oxidative stress in endothelial cells are not fully understood. Thus, we aimed to evaluate the effects of metformin and a GLP-1 analog, liraglutide on high glucose-induced oxidative stress. Production of reactive oxygen species (ROS), activation of protein kinase C (PKC) and NAD(P)H oxidase, and changes in signaling molecules in response to high glucose exposure were evaluated in human aortic endothelial cells with and without treatment of metformin and liraglutide, alone or in combination. PKC-NAD(P)H oxidase pathway was assessed by translocation of GFP-fused PKCβ2 isoform and GFP-fused p47phox, a regulatory subunit of NAD(P)H oxidase, in addition to endogenous PKC phosphorylation and NAD(P)H oxidase activity. High glucose-induced ROS overproduction was blunted by metformin or liraglutide treatment, with a further decrease by a combination of these drugs. Exposure to high glucose caused PKCβ2 translocation and a time-dependent phosphorylation of endogenous PKC but failed to induce its translocation and phosphorylation in the cells treated with metformin and liraglutide. Furthermore, both drugs inhibited p47phox translocation and NAD(P)H oxidase activation, and prevented the high glucose-induced changes in intracellulalr diacylglycerol (DAG) level and phosphorylation of AMP-activated protein kinase (AMPK). A combination of these drugs further enhanced all of these effects. Metformin and liraglutide ameliorate high glucose-induced oxidative stress by inhibiting PKC-NAD(P)H oxidase pathway. A combination of these two drugs provides augmented protective effects, suggesting the clinical usefulness in prevention of diabetic vascular complications. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Electrical bistability and charge-transport mechanisms in cuprous sulfide nanosphere-poly(N-vinylcarbazole) composite films

International Nuclear Information System (INIS)

Tang Aiwei; Teng Feng; Liu Jie; Wang Yichao; Peng Hongshang; Hou Yanbing; Wang Yongsheng

2011-01-01

In this study, electrically bistable devices were fabricated by incorporating cuprous sulfide (Cu 2 S) nanospheres with mean size less than 10 nm into a poly(N-vinylcarbazole) (PVK) matrix. A remarkable electrical bistability was clearly observed in the current–voltage curves of the devices due to an electric-field-induced charge transfer between the dodecanethiol-capped Cu 2 S nanospheres and PVK. The maximum ON/OFF current ratio reached up to value as large as 10 4 , which was dependent on the mass ratios of Cu 2 S nanospheres to PVK, the amplitude of the scanning voltages, and the film thickness. The charge-transport mechanisms of the electrically bistable devices were described on the basis of the experimental results using different theoretical models of organic electronics.

Millimeter- and submillimeter-wave spectrum and molecular constants of cuprous iodide

International Nuclear Information System (INIS)

Manson, E.L.; De Lucia, F.C.; Gordy, W.

1975-01-01

Monomeric cuprous iodide (CuI) in the vapor state has been produced as an intermediate product of the reaction of copper with iodine vapor at 1100 degreeK in a quasi-free-space microwave absorption cell. Rotational transitions of 63 Cu 127 I and 65 Cu 127 I were measured for J as high as 87→88 and for ν as high as 11. Hyperfine splittings due to 127 I were observed, and eqQ ( 127 I) =-935(15) MHz was derived. Analysis of the rotational frequencies yields the following Dunham and equilibrium coefficients for 63 Cu 127 I: Y 01 =2197.10172(98) MHz, Y 11 =-8.51120(66) MHz, Y 21 =0.00934(14) MHz, Y 31 =-0.0375(88) kHz, Y 02 =-0.67269(10) kHz, Y 12 =0.031(34) Hz, Y 03 =-0.00006858(10) Hz, B/sube/=2197.102(2) MHz, ω/sube/=264.897(18) cm -1 , ω/sube/chi/sube/=0.715(2) cm -1 , r/sube/=2.33831686(104) A; for 65 Cu 127 I: Y 01 =2151.89989(94) MHz, Y 11 =-8.24990(66) MHz, Y 21 =0.00896(14) MHz, Y 31 =-0.0356(88) kHz, Y 02 =-0.64530(10) kHz, Y 12 =0.030(34) Hz, Y 03 =-0.00006443(10) Hz, B/sube/=2151.900(2) MHz, ω/sube/=262.157(18) cm -1 , ω/sube/chi/sube/=0.700(2) cm -1 , r/sube/=2.33831664(102) A. The Dunham potential constants for CuI are a 0 =239 366(34) cm -1 , a 1 =-3.33365(36), a 2 =7.391(16), and a 3 =-13.20(18)

A high-throughput colorimetric assay for glucose detection based on glucose oxidase-catalyzed enlargement of gold nanoparticles

Science.gov (United States)

Xiong, Yanmei; Zhang, Yuyan; Rong, Pengfei; Yang, Jie; Wang, Wei; Liu, Dingbin

2015-09-01

We developed a simple high-throughput colorimetric assay to detect glucose based on the glucose oxidase (GOx)-catalysed enlargement of gold nanoparticles (AuNPs). Compared with the currently available glucose kit method, the AuNP-based assay provides higher clinical sensitivity at lower cost, indicating its great potential to be a powerful tool for clinical screening of glucose.We developed a simple high-throughput colorimetric assay to detect glucose based on the glucose oxidase (GOx)-catalysed enlargement of gold nanoparticles (AuNPs). Compared with the currently available glucose kit method, the AuNP-based assay provides higher clinical sensitivity at lower cost, indicating its great potential to be a powerful tool for clinical screening of glucose. Electronic supplementary information (ESI) available: Experimental section and additional figures. See DOI: 10.1039/c5nr03758a

Hierarchical CNFs/MnCo2O4.5 nanofibers as a highly active oxidase mimetic and its application in biosensing

Science.gov (United States)

Gao, Mu; Lu, Xiaofeng; Nie, Guangdi; Chi, Maoqiang; Wang, Ce

2017-12-01

Recently, much attention has been paid on the nanomaterial-based artificial enzymes due to their tunable catalytic activity, high stability and low cost compared to the natural enzymes. Different from the peroxidase mimics which have been studied for several decades, nanomaterials with oxidase-like property are burgeoning in the recent years. In this paper, hierarchical carbon nanofibers (CNFs)/MnCo2O4.5 nanofibers as efficient oxidase mimics are reported. The products are synthesized by an electrospinning technique and an electrochemcial deposition process in which the CNFs are used as the working electrode where MnCo2O4.5 nanosheets deposit on. The resulting binary metal oxide-based nanocomposites exhibit a good oxidase-like activity toward the oxidations of 3,3‧,5,5‧tetramethylbenzi-dine (TMB), 2,2‧-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium (ABTS) salt and o-phenylenediamine (OPD) without exogenous addition of H2O2. The system of CNFs/MnCo2O4.5-TMB can be used as a candidate to detect sulfite and ascorbic acid via a colorimetric method with a high sensitivity. This work provides the efficient utilization and potential applications of binary metal oxide-based nanocomposites with oxidase activities in biosensors and other biotechnologies.

Activation of persulfate/copper by hydroxylamine via accelerating the cupric/cuprous redox couple.

Science.gov (United States)

Zhou, Peng; Zhang, Jing; Liang, Juan; Zhang, Yongli; Liu, Ya; Liu, Bei

2016-01-01

Cuprous copper [Cu(I)] reacts with sodium persulfate (PDS) to generate sulfate radical SO4(-)•, but it has been seldom investigated owing to its instability and difficulty in dissolving it. This study proposes a new method to regenerate Cu(I) from cupric copper [Cu(II)] by addition of hydroxylamine (HA) to induce the continuous production of radicals through active PDS, and investigates the resulting enhanced methyl orange (MO) degradation efficiency and mechanism in the new system. HA accelerated the degradation of MO markedly in the pH range from 6.0 to 8.0 in the HA/Cu(II)/PDS process. Both SO4(-)• and hydroxyl radicals (•OH) were considered as the primary reactive radicals in the process. The MO degradation in the HA/Cu(II)/PDS process can be divided into three stages: the fast stage, the transitory stage, and the low stage. MO degradation was enhanced with increased dosage of PDS. Although high dosage of HA could accelerate the transformation of the Cu(II)/Cu(I) cycle to produce more reactive radicals, excess HA can quench the reactive radicals. This study indicates that through a copper-redox cycling mechanism by HA, the production of SO4(-)• and •OH can be strongly enhanced, and the effective pH range can be expanded to neutral conditions.

Glucose oxidase variants with improved properities

OpenAIRE

Fischer, Rainer; Ostafe, Raluca; Prodanovic, Radivoje

2014-01-01

Source: WO14173822A3 [EN] The technology provided herein relates to novel variants of microbial glucose oxidase with improved properties, more specifically to polypeptides having glucose oxidase activity as their major enzymatic activity; to nucleic acid molecules encoding said glucose oxidases; vectors and host cells containing the nucleic acids and methods for producing the glucose oxidase; compositions comprising said glucose oxidase; methods for the preparation and production of such enzy...

Stability of sputter deposited cuprous oxide (Cu2O) subjected to ageing conditions for photovoltaic applications

Science.gov (United States)

Camacho-Espinosa, E.; Rimmaudo, I.; Riech, I.; Mis-Fernández, R.; Peña, J. L.

2018-02-01

Among various metal oxide p-type semiconductors, cuprous oxide (Cu2O) stands out as a nontoxic and abundant material, which also makes it a suitable candidate as a low-cost absorber for photovoltaic applications. However, the chemical stability of the absorber layer is critical for the solar cell lifetime, in particular, for Cu-based materials, concerning to its oxidation state changes. In this paper, we addressed the Cu2O stability depositing films of 170 nm by reactive radio frequency magnetron sputtering and subsequently ageing them in conditions similar to the typical accelerated life test for the solar module, in a period of time from one to five weeks. The stability of the optical, electrical, and structural properties of the Cu2O thin films was investigated using UV-VIS-near infrared transmittance, 4-probes electrical resistance characterization, high precision profilometry, X-ray photoelectron spectroscopy, and grazing incidence X-ray diffraction. Finally, we demonstrated that the aging tests affected only the surface of the films, while the bulk remained unaltered, making Cu2O a promising candidate for production of stable devices, including solar cells.

Evaluation of oxalate decarboxylase and oxalate oxidase for industrial applications.

Science.gov (United States)

Cassland, Pierre; Sjöde, Anders; Winestrand, Sandra; Jönsson, Leif J; Nilvebrant, Nils-Olof

2010-05-01

Increased recirculation of process water has given rise to problems with formation of calcium oxalate incrusts (scaling) in the pulp and paper industry and in forest biorefineries. The potential in using oxalate decarboxylase from Aspergillus niger for oxalic acid removal in industrial bleaching plant filtrates containing oxalic acid was examined and compared with barley oxalate oxidase. Ten different filtrates from chemical pulping were selected for the evaluation. Oxalate decarboxylase degraded oxalic acid faster than oxalate oxidase in eight of the filtrates, while oxalate oxidase performed better in one filtrate. One of the filtrates inhibited both enzymes. The potential inhibitory effect of selected compounds on the enzymatic activity was tested. Oxalate decarboxylase was more sensitive than oxalate oxidase to hydrogen peroxide. Oxalate decarboxylase was not as sensitive to chlorate and chlorite as oxalate oxidase. Up to 4 mM chlorate ions, the highest concentration tested, had no inhibitory effect on oxalate decarboxylase. Analysis of the filtrates suggests that high concentrations of chlorate present in some of the filtrates were responsible for the higher sensitivity of oxalate oxidase in these filtrates. Oxalate decarboxylase was thus a better choice than oxalate oxidase for treatment of filtrates from chlorine dioxide bleaching.

Tea polyphenols alleviate high fat and high glucose-induced endothelial hyperpermeability by attenuating ROS production via NADPH oxidase pathway.

Science.gov (United States)

Zuo, Xuezhi; Tian, Chong; Zhao, Nana; Ren, Weiye; Meng, Yi; Jin, Xin; Zhang, Ying; Ding, Shibin; Ying, Chenjiang; Ye, Xiaolei

2014-03-02

Hyperglycemia-induced endothelial hyperpermeability is crucial to cardiovascular disorders and macro-vascular complications in diabetes mellitus. The objective of this study is to investigate the effects of green tea polyphenols (GTPs) on endothelial hyperpermeability and the role of nicotinamide adenine dinucleotide phosphate (NADPH) pathway. Male Wistar rats fed on a high fat diet (HF) were treated with GTPs (0, 0.8, 1.6, 3.2 g/L in drinking water) for 26 weeks. Bovine aortic endothelial cells (BAECs) were treated with high glucose (HG, 33 mmol/L) and GTPs (0.0, 0.4, or 4 μg/mL) for 24 hours in vitro. The endothelial permeabilities in rat aorta and monolayer BAECs were measured by Evans blue injection method and efflux of fluorescein isothiocyanate (FITC)-dextran, respectively. The reactive oxygen species (ROS) levels in rat aorta and monolayer BAECs were measured by dihydroethidium (DHE) and 2', 7'-dichloro-fluorescein diacetate (DCFH-DA) fluorescent probe, respectively. Protein levels of NADPH oxidase subunits were determined by Western-blot. HF diet-fed increased the endothelial permeability and ROS levels in rat aorta while HG treatments increased the endothelial permeability and ROS levels in cultured BAECs. Co-treatment with GTPs alleviated those changes both in vivo and in vitro. In in vitro studies, GTPs treatments protected against the HG-induced over-expressions of p22phox and p67phox. Diphenylene iodonium chloride (DPI), an inhibitor of NADPH oxidase, alleviated the hyperpermeability induced by HG. GTPs could alleviate endothelial hyperpermeabilities in HF diet-fed rat aorta and in HG treated BAECs. The decrease of ROS production resulting from down-regulation of NADPH oxidase contributed to the alleviation of endothelial hyperpermeability.

Differences in Monoamine Oxidase Activity in the Brain of Wistar and August Rats with High and Low Locomotor Activity: A Cytochemical Study.

Science.gov (United States)

Sergutina, A V; Rakhmanova, V I

2016-06-01

Monoamine oxidase activity was quantitatively assessed by cytochemical method in brain structures (layers III and V of the sensorimotor cortex, caudate nucleus, nucleus accumbens, hippocampal CA3 field) of rats of August line and Wistar population with high and low locomotor activity in the open fi eld test. Monoamine oxidase activity (substrate tryptamine) predominated in the nucleus accumbens of Wistar rats with high motor activity in comparison with rats with low locomotor activity. In August rats, enzyme activity (substrates tryptamine and serotonin) predominated in the hippocampus of animals with high motor activity. Comparison of August rats with low locomotor activity and Wistar rats with high motor activity (i.e. animals demonstrating maximum differences in motor function) revealed significantly higher activity of the enzyme (substrates tryptamine and serotonin) in the hippocampus of Wistar rats. The study demonstrates clear-cut morphochemical specificity of monoaminergic metabolism based on the differences in the cytochemical parameter "monoamine oxidase activity", in the studied brain structures, responsible for the formation and realization of goal-directed behavior in Wistar and August rats.

Production of rabbit antibodies against purified Glucose oxidase

Directory of Open Access Journals (Sweden)

Muhammad Anjum Zia

2012-02-01

Full Text Available Glucose oxidase is an active oxygen species generating enzyme produced from Aspergillus niger grown in submerged fermentation. Disintegration of the mycelium resulted in high glucose oxidase activity that was subjected to ammonium sulfate precipitation at 60-85% saturation rates that resulted to 6.14 U mg -1 specific activity. Purification of enzyme by anion exchange column (DEAE-Cellulose resulted into 22.53 U mg-1 specific activity and 10.27 fold purification. This was applied to sephadex G-200 column for gel filtration chromatography. It was observed that enzyme achieved 59.37 U mg-1of specific activity with 27.08 fold purity and 64.36% recovery. Purified glucose oxidase was injected into rabbits through intravenous route, to raise the glucose oxidase antibodies. After 30 days incubation period, the rabbits were slaughtered and serum was separated from blood. The antibodies were isolated by ammonium sulfate precipitation and confirmed by agar gel precipitation test. This could be a convenient and low cost alternate assay for the estimation of glucose oxidase in biological fluids. Moreover, such antibodies against the said enzyme could be used in various therapeutic and diagnostic applications.

Evaluation of the Expression of Amine Oxidase Proteins in Breast Cancer

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Woo Young Sun

2017-12-01

Full Text Available We aimed to evaluate the expression of amine oxidase proteins in breast cancer and their clinical implications. We performed immunohistochemical staining of amine oxidase proteins (LOX, lysyl oxidase, AOC3, amine oxidase, MAOA, monoamine oxidase A, MAOB, monoamine oxidase B. Based on their hormone receptors, such as estrogen receptor (ER and progesterone receptor (PR, human epidermal growth factor receptor 2 (HER-2, and Ki-67 immunohistochemical staining, breast cancer was divided into four molecular subtypes: luminal A, luminal B, HER-2 type, and triple-negative breast cancer (TNBC. Luminal A was observed in 380 cases (49.4%, luminal B in 224 (29.1%, HER-2 type in 68 (8.8%, and TNBC in 98 (12.7%. Stromal AOC3, MAO-A, and MAO-B expression varied according to molecular subtypes. Stromal AOC3 expression was high in luminal B and HER-2 type and MAO-A expression was high in luminal A and luminal B (p < 0.001. MAO-B expression was higher in TNBC than in other subtypes (p = 0.020. LOX positivity was associated with high histological grade (p < 0.001 and high Ki-67 labeling index (LI (p = 0.009, and stromal AOC3 positivity was associated with high histological grade (p = 0.001, high Ki-67 LI (p < 0.001, and HER-2 positivity (p = 0.002. MAO-A positivity was related to low histological grade (p < 0.001, ER positivity, PR positivity (p < 0.001, and low Ki-67 LI (p < 0.001. In univariate analysis, MAO-A positivity was related to short disease-free survival in HER-2 type (p = 0.013, AOC3 negativity was related to short disease-free survival and overall survival in ER-positive breast cancer, PR-positive breast cancer, HER-2-negative breast cancer, and lymph node metastasis. In conclusion, the expression of amine oxidase proteins varies depending on the molecular subtype of breast cancer. Stromal AOC3 expression was high in luminal B and HER-2 type, and MAO-A expression was high in luminal A and luminal B.

Effect of high pressure on peanut allergens in the presence of polyphenol oxidase and caffeic acid

Science.gov (United States)

High pressure (HP) enhances enzymatic reactions. Because polyphenol oxidase (PPO) is an enzyme, and reduces IgE binding of peanut allergens in presence of caffeic acid (CA), we postulated that a further reduction in IgE binding can be achieved, using HP together with PPO and CA. Peanut extracts cont...

Platelet monoamine oxidase: specific activity and turnover number in headache

International Nuclear Information System (INIS)

Summers, K.M.; Brown, G.K.; Craig, I.W.; Peatfield, R.; Rose, F.C.

1982-01-01

Monoamine oxidase turnover numbers (molecules of substrate converted to product per minute per active site) have been calculated for the human platelet enzyme using [ 3 H]pargyline. Headache patients with high and low monoamine oxidase specific activities relative to controls were found to have turnover numbers very close to those for controls. This finding suggests that their specific activities vary because of differences in the concentration of active monoamine oxidase molecules, rather than differences in the ability of those enzyme molecules to catalyse the deamination reaction. (Auth.)

Peroxisomal Polyamine Oxidase and NADPH-Oxidase cross-talk for ROS homeostasis which affects respiration rate in Arabidopsis thaliana

Directory of Open Access Journals (Sweden)

Efthimios A. Andronis

2014-04-01

Full Text Available Homeostasis of reactive oxygen species (ROS in the intracellular compartments is of critical importance as ROS have been linked with nearly all cellular processes and more importantly with diseases and aging. PAs are nitrogenous molecules with an evolutionary conserved role in the regulation of metabolic and energetic status of cells. Recent evidence also suggests that polyamines (PA are major regulators of ROS homeostasis. In Arabidopsis the backconversion of the PAs spermidine (Spd and spermine (Spm to putrescine (Put and Spd, respectively is catalyzed by two peroxisomal PA oxidases (AtPAO. However, the physiological role of this pathway remains largely elusive. Here we explore the role of peroxisomal PA backconversion and in particular that catalyzed by the highly expressed AtPAO3 in the regulation of ROS homeostasis and mitochondrial respiratory burst. Exogenous PAs exert an NADPH-oxidase dependent stimulation of oxygen consumption, with Spd exerting the strongest effect. This increase is attenuated by treatment with the NADPH-oxidase blocker diphenyleneiodonium iodide (DPI. Loss-of-function of AtPAO3 gene results to increased NADPH-oxidase-dependent production of superoxide anions (O2.-, but not H2O2, which activate the mitochondrial alternative oxidase pathway (AOX. On the contrary, overexpression of AtPAO3 results to an increased but balanced production of both H2O2 and O2.-. These results suggest that the ratio of O2.-/H2O2 regulates respiratory chain in mitochondria, with PA-dependent production of O2.- by NADPH-oxidase tilting the balance of electron transfer chain in favor of the AOX pathway. In addition, AtPAO3 seems to be an important component in the regulating module of ROS homeostasis, while a conserved role for PA backconversion and ROS across kingdoms is discussed.

Production of rabbit antibodies against purified Glucose oxidase

OpenAIRE

Zia,Muhammad Anjum; Ain,Qurat-ul; Iftikhar,Tehreema; Abbas,Rao Zahid; Rahman,Khalil-ur

2012-01-01

Glucose oxidase is an active oxygen species generating enzyme produced from Aspergillus niger grown in submerged fermentation. Disintegration of the mycelium resulted in high glucose oxidase activity that was subjected to ammonium sulfate precipitation at 60-85% saturation rates that resulted to 6.14 U mg -1 specific activity. Purification of enzyme by anion exchange column (DEAE-Cellulose) resulted into 22.53 U mg-1 specific activity and 10.27 fold purification. This was applied to sephadex ...

Oxidase-based biocatalytic processes

DEFF Research Database (Denmark)

Ramesh, Hemalata; Woodley, John; Krühne, Ulrich

interestingbiocatalystsbecause they use a mild oxidant (oxygen) as a substrateas opposed to their chemical counterparts which use strong oxidants such as permanganates. A class of oxidases calledmonoamine oxidases has been used as the central case study for the thesis. The rationale for choosing thissystemis that it has been...

Putting together a plasma membrane NADH oxidase: a tale of three laboratories.

Science.gov (United States)

Löw, Hans; Crane, Frederick L; Morré, D James

2012-11-01

The observation that high cellular concentrations of NADH were associated with low adenylate cyclase activity led to a search for the mechanism of the effect. Since cyclase is in the plasma membrane, we considered the membrane might have a site for NADH action, and that NADH might be oxidized at that site. A test for NADH oxidase showed very low activity, which could be increased by adding growth factors. The plasma membrane oxidase was not inhibited by inhibitors of mitochondrial NADH oxidase such as cyanide, rotenone or antimycin. Stimulation of the plasma membrane oxidase by iso-proterenol or triiodothyronine was different from lack of stimulation in endoplasmic reticulum. After 25 years of research, three components of a trans membrane NADH oxidase have been discovered. Flavoprotein NADH coenzyme Q reductases (NADH cytochrome b reductase) on the inside, coenzyme Q in the middle, and a coenzyme Q oxidase on the outside as a terminal oxidase. The external oxidase segment is a copper protein with unique properties in timekeeping, protein disulfide isomerase and endogenous NADH oxidase activity, which affords a mechanism for control of cell growth by the overall NADH oxidase and the remarkable inhibition of oxidase activity and growth of cancer cells by a wide range of anti-tumor drugs. A second trans plasma membrane electron transport system has been found in voltage dependent anion channel (VDAC), which has NADH ferricyanide reductase activity. This activity must be considered in relation to ferricyanide stimulation of growth and increased VDAC antibodies in patients with autism. Copyright © 2012 Elsevier Ltd. All rights reserved.

NiCoBP-doped carbon nanotube hybrid: A novel oxidase mimetic system for highly efficient electrochemical immunoassay

Energy Technology Data Exchange (ETDEWEB)

Zhang, Bing; He, Yu; Liu, Bingqian; Tang, Dianping, E-mail: dianping.tang@fzu.edu.cn

2014-12-03

Highlights: • We report a new oxidase mimetic system for highly efficient electrochemical immunoassay. • NiCoBP-doped carbon nanotube hybrids were used as the nanocatalysts. • NiCoBP-doped carbon nanotube hybrids were used as the mimic oxidase. - Abstract: NiCoBP-doped multi-walled carbon nanotube (NiCoBP–MWCNT) was first synthesized by using induced electroless-plating method and functionalized with the biomolecules for highly efficient electrochemical immunoassay of prostate-specific antigen (PSA, used as a model analyte). We discovered that the as-synthesized NiCoBP–MWCNT had the ability to catalyze the glucose oxidization with a stable and well-defined redox peak. The catalytic current increased with the increment of the immobilized NiCoBP–MWCNT on the electrode. Transmission electron microscope (TEM) and energy dispersive X-ray spectrometry (EDX) were employed to characterize the as-prepared NiCoBP–MWCNT. Using the NiCoBP–MWCNT-conjugated anti-PSA antibody as the signal-transduction tag, a new enzyme-free electrochemical immunoassay protocol could be designed for the detection of target PSA on the capture antibody-functionalized immunosensing interface. Experimental results revealed that the designed immunoassay system could exhibit good electrochemical responses toward target PSA, and allowed the detection of PSA at a concentration as low as 0.035 ng mL{sup −1}. More importantly, the NiCoBP-MWCNT-based oxidase mimetic system could be further extended for the monitoring of other low-abundance proteins or disease-related biomarkers by tuning the target antibody.

Exciton-polaritons in cuprous oxide: Theory and comparison with experiment

Science.gov (United States)

Schweiner, Frank; Ertl, Jan; Main, Jörg; Wunner, Günter; Uihlein, Christoph

2017-12-01

The observation of giant Rydberg excitons in cuprous oxide (Cu2O ) up to a principal quantum number of n =25 by T. Kazimierczuk et al. [Nature (London) 514, 343 (2014), 10.1038/nature13832] inevitably raises the question whether these quasiparticles must be described within a multipolariton framework since excitons and photons are always coupled in the solid. In this paper we present the theory of exciton-polaritons in Cu2O . To this end we extend the Hamiltonian which includes the complete valence-band structure, the exchange interaction, and the central-cell corrections effects, and which has been recently deduced by F. Schweiner et al. [Phys. Rev. B 95, 195201 (2017), 10.1103/PhysRevB.95.195201], for finite values of the exciton momentum ℏ K . We derive formulas to calculate not only dipole but also quadrupole oscillator strengths when using the complete basis of F. Schweiner et al., which has recently been proven as a powerful tool to calculate exciton spectra. Very complex polariton spectra for the three orientations of K along the axes [001 ] , [110 ] , and [111 ] of high symmetry are obtained and a strong mixing of exciton states is reported. The main focus is on the 1 S ortho-exciton-polariton, for which pronounced polariton effects have been measured in experiments. We set up a 5 ×5 matrix model, which accounts for both the polariton effect and the K -dependent splitting, and which allows treating the anisotropic polariton dispersion for any direction of K . We especially discuss the dispersions for K being oriented in the planes perpendicular to [1 1 ¯0 ] and [111 ] , for which experimental transmission spectra have been measured. Furthermore, we compare our results with experimental values of the K -dependent splitting, the group velocity, and the oscillator strengths of this exciton-polariton. The results are in good agreement. This proves the validity of the 5 ×5 matrix model as a useful theoretical model for further investigations on the 1 S

Isolated sulfite oxidase deficiency.

Science.gov (United States)

Rupar, C A; Gillett, J; Gordon, B A; Ramsay, D A; Johnson, J L; Garrett, R M; Rajagopalan, K V; Jung, J H; Bacheyie, G S; Sellers, A R

1996-12-01

Isolated sulfite oxidase (SO) deficiency is an autosomal recessively inherited inborn error of sulfur metabolism. In this report of a ninth patient the clinical history, laboratory results, neuropathological findings and a mutation in the sulfite oxidase gene are described. The data from this patient and previously published patients with isolated sulfite oxidase deficiency and molybdenum cofactor deficiency are summarized to characterize this rare disorder. The patient presented neonatally with intractable seizures and did not progress developmentally beyond the neonatal stage. Dislocated lenses were apparent at 2 months. There was increased urine excretion of sulfite and S-sulfocysteine and a decreased concentration of plasma cystine. A lactic acidemia was present for 6 months. Liver sulfite oxidase activity was not detectable but xanthine dehydrogenase activity was normal. The boy died of respiratory failure at 32 months. Neuropathological findings of cortical necrosis and extensive cavitating leukoencephalopathy were reminiscent of those seen in severe perinatal asphyxia suggesting an etiology of energy deficiency. A point mutation that resulted in a truncated protein missing the molybdenum-binding site has been identified.

Characterization of wheat germin (oxalate oxidase) expressed by Pichia pastoris

International Nuclear Information System (INIS)

Pan, Heng-Yen; Whittaker, Mei M.; Bouveret, Romaric; Berna, Anne; Bernier, Francois; Whittaker, James W.

2007-01-01

High-level secretory expression of wheat (Triticum aestivum) germin/oxalate oxidase was achieved in Pichia pastoris fermentation cultures as an α-mating factor signal peptide fusion, based on the native wheat cDNA coding sequence. The oxalate oxidase activity of the recombinant enzyme is substantially increased (7-fold) by treatment with sodium periodate, followed by ascorbate reduction. Using these methods, approximately 1 g (4 x 10 4 U) of purified, activated enzyme was obtained following eight days of induction of a high density Pichia fermentation culture, demonstrating suitability for large-scale production of oxalate oxidase for biotechnological applications. Characterization of the recombinant protein shows that it is glycosylated, with N-linked glycan attached at Asn47. For potential biomedical applications, a nonglycosylated (S49A) variant was also prepared which retains essentially full enzyme activity, but exhibits altered protein-protein interactions

Facile synthesis of cuprous oxide nanowires decorated graphene oxide nanosheets nanocomposites and its application in label-free electrochemical immunosensor.

Science.gov (United States)

Wang, Huan; Zhang, Yong; Wang, Yulan; Ma, Hongmin; Du, Bin; Wei, Qin

2017-01-15

In this work, the assembly between one-dimensional (1D) nanomaterials and two-dimensional (2D) nanomaterials was achieved by a simple method. Cuprous oxide nanowires decorated graphene oxide nanosheets (Cu 2 O@GO) nanocomposites were synthesized for the first time by a simple electrostatic self-assembly process. The nanostructure was well confirmed by scanning electron microscope (SEM) and transmission electron microscope (TEM) images. Taking advantages of good electrocatalytic activity and high specific surface area of Cu 2 O@GO nanocomposites, a label-free electrochemical immunosensor was developed by employing Cu 2 O@GO as signal amplification platform for the quantitative detection of alpha fetoprotein (AFP). In addition, toluidine blue (TB) was used as the electron transfer mediator to provide the electrochemical signal, which was adsorbed on graphene oxide nanosheets (GO NSs) by electrostatic attraction. The detection mechanism was based on the monitoring of the electrochemical current response change of TB by the square wave voltammetry (SWV) when immunoreaction occurred on the surface of electrode. Under optimal conditions, the proposed immunosensor displayed a high sensitivity and a low detection limit. This designed method may provide an effective method in the clinical diagnosis of AFP and other tumor markers. Copyright © 2016 Elsevier B.V. All rights reserved.

A role for NADPH oxidase in antigen presentation

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Gail J Gardiner

2013-09-01

Full Text Available The nicotinamide adenine dinucleotide phosphate (NADPH oxidase expressed in phagocytes is a multi-subunit enzyme complex that generates superoxide (O2.-. This radical is an important precursor of hydrogen peroxide (H2O2 and other reactive oxygen species (ROS needed for microbicidal activity during innate immune responses. Inherited defects in NADPH oxidase give rise to chronic granulomatous disease (CGD, a primary immunodeficiency characterized by recurrent infections and granulomatous inflammation. Interestingly, CGD, CGD carrier status, and oxidase gene polymorphisms have all been associated with autoinflammatory and autoimmune disorders, suggesting a potential role for NADPH oxidase in regulating adaptive immune responses. Here, NADPH oxidase function in antigen processing and presentation is reviewed. NADPH oxidase influences dendritic cell (DC crosspresentation by major histocompatibility complex class I molecules (MHC-I through regulation of the phagosomal microenvironment, while in B lymphocytes, NADPH oxidase alters epitope selection by major histocompatibility complex class II molecules (MHC-II.

Role of pH in oxidase variability of Aeromonas hydrophila.

OpenAIRE

Hunt, L K; Overman, T L; Otero, R B

1981-01-01

Some strains of Aeromonas hydrophila may be oxidase negative or only weakly oxidase positive by the Kovacs method taken from the surface of a differential medium, such as MacConkey agar. Six strains of A. hydrophila, two oxidase variable, one oxidase constant, and three weakly oxidase positive on MacConkey agar, were studied to determine the cause of oxidase variability. The bacteriostatic dyes in MacConkey agar were considered possible inhibitors of the oxidase reaction. The concentration of...

Discovery, characterization, and kinetic analysis of an alditol oxidase from streptomyces coelicolor

NARCIS (Netherlands)

Heuts, Dominic P. H. M.; van Hellemond, Erik W.; Janssen, Dick B.; Fraaije, Marco W.

2007-01-01

A gene encoding an alditol oxidase was found in the genome of Streptomyces coelicolor A3(2). This newly identified oxidase, AldO, was expressed at extremely high levels in Escherichia coli when fused to maltose-binding protein. AldO is a soluble monomeric flavoprotein with subunits of 45.1 kDa, each

Phospholipid alterations in cardiac sarcoplasmic reticulum induced by xanthine oxidase: contamination of commercial preparations of xanthine oxidase by phospholipase A2

International Nuclear Information System (INIS)

Gamache, D.A.; Kornberg, L.J.; Bartolf, M.; Franson, R.C.

1986-01-01

Incubation of cardiac sarcoplasmic reticulum with xanthine oxidase alone at pH 7.0 resulted in a loss of lipid phosphorus that was potentiated by the addition of xanthine. Using autoclaved E.coli with 1- 14 C-oleate in the 2-acyl position of membrane phospholipids, the authors demonstrate that many, but not all, commercial preparations of xanthine oxidase contain significant phospholipase A 2 (PLA 2 ) activity (64.3-545.6 nmols/min/mg). The PLA 2 was maximally active in the neutral-alkaline pH range, was Ca 2+ -dependent, and was unaffected by the addition of xanthine. PLA 2 activity was totally inhibited by 1mM EDTA whereas radical production by optimal concentrations of xanthine/xanthine oxidase (X/XO) was unaffected by EDTA. Chromatographically purified xanthine oxidase (Sigma Grade III) contained high levels of PLA 2 activity (64.3 nmols/min/mg) compared to endogenous levels of neutral-active, Ca 2+ -dependent PLA 2 measured in various tissue homogenates (≤ 0.5 nmols/ min/mg). Because X/XO mixtures are used extensively to study oxygen free radical-induced cell injury and membrane phospholipid alterations, the presence of a potent extracellular PLA 2 may have influenced previously published reports, and such studies should be interpreted cautiously

Multilayer core-shell structured composite paper electrode consisting of copper, cuprous oxide and graphite assembled on cellulose fibers for asymmetric supercapacitors

Science.gov (United States)

Wan, Caichao; Jiao, Yue; Li, Jian

2017-09-01

An easily-operated and inexpensive strategy (pencil-drawing-electrodeposition-electro-oxidation) is proposed to synthesize a novel class of multilayer core-shell structured composite paper electrode, which consists of copper, cuprous oxide and graphite assembled on cellulose fibers. This interesting electrode structure plays a pivotal role in providing more active sites for electrochemical reactions, facilitating ion and electron transport and shorting their diffusion pathways. This electrode demonstrates excellent electrochemical properties with a high specific capacitance of 601 F g-1 at 2 A g-1 and retains 83% of this capacitance when operated at an ultrahigh current density of 100 A g-1. In addition, a high energy density of 13.4 W h kg-1 at the power density of 0.40 kW kg-1 and a favorable cycling stability (95.3%, 8000 cycles) were achieved for this electrode. When this electrode was assembled into an asymmetric supercapacitor with carbon paper as negative electrode, the device displays remarkable electrochemical performances with a large areal capacitances (122 mF cm-2 at 1 mA cm-2), high areal energy density (10.8 μW h cm-2 at 402.5 μW cm-2) and outstanding cycling stability (91.5%, 5000 cycles). These results unveil the potential of this composite electrode as a high-performance electrode material for supercapacitors.

Preparation, characterization and nonlinear absorption studies of cuprous oxide nanoclusters, micro-cubes and micro-particles

Science.gov (United States)

Sekhar, H.; Narayana Rao, D.

2012-07-01

Cuprous oxide nanoclusters, micro-cubes and micro-particles were successfully synthesized by reducing copper(II) salt with ascorbic acid in the presence of sodium hydroxide via a co-precipitation method. The X-ray diffraction and FTIR studies revealed that the formation of pure single-phase cubic. Raman and EPR spectral studies show the presence of CuO in as-synthesized powders of Cu2O. Transmission electron microscopy and field emission scanning electron microscopy data revealed that the morphology evolves from nanoclusters to micro-cubes and micro-particles by increasing the concentration of NaOH. Linear optical measurements show absorption peak maximum shifts towards red with changing morphology from nanoclusters to micro-cubes and micro-particles. The nonlinear optical properties were studied using open aperture Z-scan technique with 532 nm 6 ns laser pulses. Samples-exhibited both saturable as well as reverse saturable absorption. Due to confinement effects (enhanced band gap), we observed enhanced nonlinear absorption coefficient (β) in the case of nanoclusters compared to their micro-cubes and micro-particles.

A novel reducing graphene/polyaniline/cuprous oxide composite hydrogel with unexpected photocatalytic activity for the degradation of Congo red

International Nuclear Information System (INIS)

Miao, Jie; Xie, Anjian; Li, Shikuo; Huang, Fangzhi; Cao, Juan; Shen, Yuhua

2016-01-01

Graphical abstract: Excellent photocatalytic activity of the RGO/PANI/Cu_2O composite hydrogel for CR degradation under UV–vis light irradiation. - Highlights: • The RGO/PANI/Cu_2O composite hydrogel was first synthesized via a facile method. • Photocatalytic performance was studied under UV–vis light. • The ternary composite hydrogel shows unexpected photocatalytic activity. • A possible photocatalysis mechanism was illustrated. - Abstract: In this work, a novel reducing graphene/polyaniline/cuprous oxide (RGO/PANI/Cu_2O) composite hydrogel with a 3D porous network has been successfully prepared via a one-pot method in the presence of cubic Cu_2O nanoparticles. The as-synthesized ternary composites hydrogel shows unexpected photocatalytic activity such that Congo red (CR) degradation efficiency can reaches 97.91% in 20 min under UV–vis light irradiation, which is much higher than that of either the single component (Cu_2O nanoparticles), or two component systems (RGO/Cu_2O composite hydrogel and PANI/Cu_2O nanocomposites). Furthermore, the ternary composite hydrogel exhibits high stability and do not show any significant loss after five recycles. Such outstanding photocatalytic activity of the RGO/PANI/Cu_2O composite hydrogel was ascribed to the high absorption ability of the product for CR and the synergic effect among RGO, PANI and Cu_2O in photocatalytic process. The product of this work would provide a new sight for the construction of UV–vis light responsive photocatalyst with high performance.

Indanones as high-potency reversible inhibitors of monoamine oxidase.

Science.gov (United States)

Mostert, Samantha; Petzer, Anél; Petzer, Jacobus P

2015-05-01

Recent reports document that α-tetralone (3,4-dihydro-2H-naphthalen-1-one) is an appropriate scaffold for the design of high-potency monoamine oxidase (MAO) inhibitors. Based on the structural similarity between α-tetralone and 1-indanone, the present study involved synthesis of 34 1-indanone and related indane derivatives as potential inhibitors of recombinant human MAO-A and MAO-B. The results show that C6-substituted indanones are particularly potent and selective MAO-B inhibitors, with IC50 values ranging from 0.001 to 0.030 μM. C5-Substituted indanone and indane derivatives are comparatively weaker MAO-B inhibitors. Although the 1-indanone and indane derivatives are selective inhibitors of the MAO-B isoform, a number of homologues are also potent MAO-A inhibitors, with three homologues possessing IC50 values 1-indanone as a reversible MAO inhibitor with a competitive mode of inhibition. It may be concluded that 1-indanones are promising leads for the design of therapies for neurodegenerative and neuropsychiatric disorders such as Parkinson's disease and depression. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

High resolution structure of the ba3 cytochrome c oxidase from Thermus thermophilus in a lipidic environment.

Directory of Open Access Journals (Sweden)

Theresa Tiefenbrunn

Full Text Available The fundamental chemistry underpinning aerobic life on Earth involves reduction of dioxygen to water with concomitant proton translocation. This process is catalyzed by members of the heme-copper oxidase (HCO superfamily. Despite the availability of crystal structures for all types of HCO, the mode of action for this enzyme is not understood at the atomic level, namely how vectorial H(+ and e(- transport are coupled. Toward addressing this problem, we report wild type and A120F mutant structures of the ba(3-type cytochrome c oxidase from Thermus thermophilus at 1.8 Å resolution. The enzyme has been crystallized from the lipidic cubic phase, which mimics the biological membrane environment. The structures reveal 20 ordered lipid molecules that occupy binding sites on the protein surface or mediate crystal packing interfaces. The interior of the protein encloses 53 water molecules, including 3 trapped in the designated K-path of proton transfer and 8 in a cluster seen also in A-type enzymes that likely functions in egress of product water and proton translocation. The hydrophobic O(2-uptake channel, connecting the active site to the lipid bilayer, contains a single water molecule nearest the Cu(B atom but otherwise exhibits no residual electron density. The active site contains strong electron density for a pair of bonded atoms bridging the heme Fe(a3 and Cu(B atoms that is best modeled as peroxide. The structure of ba(3-oxidase reveals new information about the positioning of the enzyme within the membrane and the nature of its interactions with lipid molecules. The atomic resolution details provide insight into the mechanisms of electron transfer, oxygen diffusion into the active site, reduction of oxygen to water, and pumping of protons across the membrane. The development of a robust system for production of ba(3-oxidase crystals diffracting to high resolution, together with an established expression system for generating mutants, opens the

Observation of confinement effects through liner and nonlinear absorption spectroscopy in cuprous oxide

Science.gov (United States)

Sekhar, H.; Rakesh Kumar, Y.; Narayana Rao, D.

2015-02-01

Cuprous oxide nano clusters, micro cubes and micro particles were successfully synthesized by reducing copper (II) salt with ascorbic acid in the presence of sodium hydroxide via a co-precipitation method. The X-ray diffraction studies revealed the formation of pure single phase cubic. Raman spectrum shows the inevitable presence of CuO on the surface of the Cu2O powders which may have an impact on the stability of the phase. Transmission electron microscopy (TEM) data revealed that the morphology evolves from nanoclusters to micro cubes and micro particles by increasing the concentration of NaOH. Linear optical measurements show that the absorption peak maximum shifts towards red with changing morphology from nano clusters to micro cubes and micro particles. The nonlinear optical properties were studied using open aperture Z-scan technique with 532 nm, 6 ns laser pulses. Samples exhibited saturable as well as reverse saturable absorption. The results show that the transition from SA to RSA is ascribed to excited-state absorption (ESA) induced by two-photon absorption (TPA) process. Due to confinement effects (enhanced band gap) we observed enhanced nonlinear absorption coefficient (βeff) in the case of nano-clusters compared to their micro-cubes and micro-particles.

Intracellular lysyl oxidase: Effect of a specific inhibitor on nuclear mass in proliferating cells

Energy Technology Data Exchange (ETDEWEB)

Saad, Fawzy A. [Laboratory for the Study of Skeletal Disorders and Rehabilitation, Department of Orthopedics, Children' s Hospital Boston, 300 Longwood Avenue EN926, Boston, MA 02115 (United States); Harvard Medical School, Boston, MA 02115 (United States); Torres, Marie [Laboratory for the Study of Skeletal Disorders and Rehabilitation, Department of Orthopedics, Children' s Hospital Boston, 300 Longwood Avenue EN926, Boston, MA 02115 (United States); Wang, Hao [Laboratory for the Study of Skeletal Disorders and Rehabilitation, Department of Orthopedics, Children' s Hospital Boston, 300 Longwood Avenue EN926, Boston, MA 02115 (United States); Harvard Medical School, Boston, MA 02115 (United States); Graham, Lila, E-mail: lilagraham@cs.com [Laboratory for the Study of Skeletal Disorders and Rehabilitation, Department of Orthopedics, Children' s Hospital Boston, 300 Longwood Avenue EN926, Boston, MA 02115 (United States); Harvard Medical School, Boston, MA 02115 (United States)

2010-06-11

LOX, the principal enzyme involved in crosslinking of collagen, was the first of several lysyl oxidase isotypes to be characterized. Its active form was believed to be exclusively extracellular. Active LOX was later reported to be present in cell nuclei; its function there is unknown. LOX expression opposes the effect of mutationally activated Ras, which is present in about 30% of human cancers. The mechanism of LOX in countering the action of Ras is also unknown. In the present work, assessment of nuclear protein for possible effects of lysyl oxidase activity led to the discovery that proliferating cells dramatically increase their nuclear protein content when exposed to BAPN ({beta}-aminopropionitrile), a highly specific lysyl oxidase inhibitor that reportedly blocks LOX inhibition of Ras-induced oocyte maturation. In three cell types (PC12 cells, A7r5 smooth muscle cells, and NIH 3T3 fibroblasts), BAPN caused a 1.8-, 1.7-, and 2.1-fold increase in total nuclear protein per cell, respectively, affecting all major components in both nuclear matrix and chromatin fractions. Since nuclear size is correlated with proliferative status, enzyme activity restricting nuclear growth may be involved in the lysyl oxidase tumor suppressive effect. Evidence is also presented for the presence of apparent lysyl oxidase isotype(s) containing a highly conserved LOX active site sequence in the nuclei of PC12 cells, which do not manufacture extracellular lysyl oxidase substrates. Results reported here support the hypothesis that nuclear lysyl oxidase regulates nuclear growth, and thereby modulates cell proliferation.

A new procedure for the purification of monodisperse highly active cytochrome c oxidase from bovine heart.

OpenAIRE

Li, Y; Naqui, A; Frey, T G; Chance, B

1987-01-01

A simple and rapid method for the isolation of a large quantity of cytochrome c oxidase from bovine heart mitochondria was developed, based on selective solubilization of mitochondrial protein with first Triton and then lauryl maltoside. Gel filtration shows that the lauryl maltoside-solubilized oxidase preparation is in a hydrodynamically homogeneous state with a Stokes radius of 7.5 +/- 0.2 nm. It contains 8.0 mumol of haem (with an a/a3 ratio of 1)/g of protein. The catalytic constant (max...

Biocatalytic potential of laccase-like multicopper oxidases from Aspergillus niger

Directory of Open Access Journals (Sweden)

Tamayo-Ramos Juan Antonio

2012-12-01

Full Text Available Abstract Background Laccase-like multicopper oxidases have been reported in several Aspergillus species but they remain uncharacterized. The biocatalytic potential of the Aspergillus niger fungal pigment multicopper oxidases McoA and McoB and ascomycete laccase McoG was investigated. Results The laccase-like multicopper oxidases McoA, McoB and McoG from the commonly used cell factory Aspergillus niger were homologously expressed, purified and analyzed for their biocatalytic potential. All three recombinant enzymes were monomers with apparent molecular masses ranging from 80 to 110 kDa. McoA and McoG resulted to be blue, whereas McoB was yellow. The newly obtained oxidases displayed strongly different activities towards aromatic compounds and synthetic dyes. McoB exhibited high catalytic efficiency with N,N-dimethyl-p-phenylenediamine (DMPPDA and 2,2-azino-di(3-ethylbenzthiazoline sulfonic acid (ABTS, and appeared to be a promising biocatalyst. Besides oxidizing a variety of phenolic compounds, McoB catalyzed successfully the decolorization and detoxification of the widely used textile dye malachite green. Conclusions The A. niger McoA, McoB, and McoG enzymes showed clearly different catalytic properties. Yellow McoB showed broad substrate specificity, catalyzing the oxidation of several phenolic compounds commonly present in different industrial effluents. It also harbored high decolorization and detoxification activity with the synthetic dye malachite green, showing to have an interesting potential as a new industrial biocatalyst.

The terminal oxidases of Paracoccus denitrificans

NARCIS (Netherlands)

de Gier, J.-W.; Lübben, M; Reijnders, W N; Tipker, C A; Slotboom, D.J.; van Spanning, R J; Stouthamer, A.H.; van der Oost, J.

Three distinct types of terminal oxidases participate in the aerobic respiratory pathways of Paracoccus denitrificans. Two alternative genes encoding subunit I of the aa3-type cytochrome c oxidase have been isolated before, namely ctaDI and ctaDII. Each of these genes can be expressed separately to

Quantitation of immunoadsorbed flavoprotein oxidases by luminol-mediated chemiluminescence.

Science.gov (United States)

Hinkkanen, A; Maly, F E; Decker, K

1983-04-01

The detection of the flavoenzymes 6-hydroxy-L-nicotine oxidase and 6-hydroxy-D-nicotine oxidase at the sub-femtomol level was achieved by coupling the reaction of the immunoadsorbed proteins to the peroxidase-catalysed oxidation of luminol. The H2O2-producing oxidases retained their full activity when bound to the respective immobilized antibodies. This fact allowed the concentration of the enzymes from very dilute solutions and the quantitative assay of their activities in the microU range. Due to strict stereoselectivity and the absence of immunological cross-reactivity, the two flavoproteins could be determined in the same solution. This method was used to measure the 6-hydroxy-D-nicotine oxidase and 6-hydroxy-L-nicotine oxidase activities in Escherichia coli RR1 and different Arthrobacter strains cultured under non-inducing conditions. The same activity ratio of 6-hydroxy-L-nicotine oxidase/6-hydroxy-D-nicotine oxidase as in D L-nicotine-induced cells of A. oxidans was observed in non-induced wild type and in riboflavin-requiring (rf-) mutant cells of this aerob.

Effects of glucose oxidase on the growth performance, serum ...

African Journals Online (AJOL)

Martina

2016-02-06

Feb 6, 2016 ... The experiment was conducted to investigate the effects of diets supplemented with ... Keywords: Glucose oxidase, intestinal health, performance, swine ..... This work was supported by the National High-Tech Research and ...

Fabrication of cuprous chloride films on copper substrate by chemical bath deposition

Energy Technology Data Exchange (ETDEWEB)

Lin, Yu-Ting; Ci, Ji-Wei; Tu, Wei-Chen [Department of Electronic Engineering, College of Electrical Engineering and Computer Science, Chung Yuan Christian University, Chung-Li 32023, Taiwan (China); Uen, Wu-Yih, E-mail: uenwuyih@ms37.hinet.net [Department of Electronic Engineering, College of Electrical Engineering and Computer Science, Chung Yuan Christian University, Chung-Li 32023, Taiwan (China); Lan, Shan-Ming [Department of Electronic Engineering, College of Electrical Engineering and Computer Science, Chung Yuan Christian University, Chung-Li 32023, Taiwan (China); Yang, Tsun-Neng; Shen, Chin-Chang; Wu, Chih-Hung [Institute of Nuclear Energy Research, P.O. Box 3-11, Lungtan 32500, Taiwan (China)

2015-09-30

Polycrystalline CuCl films were fabricated by chemical bath deposition (CBD) on a Cu substrate at a low solution temperature of 90 °C. Continuous CuCl films were prepared using the copper (II) chloride (CuCl{sub 2}) compound as the precursor for both the Cu{sup 2+} and Cl{sup −} sources, together with repeated HCl dip treatments. An HCl dip pretreatment of the substrate favored the nucleation of CuCl crystallites. Further, interrupting the film deposition and including an HCl dip treatment of the film growth surface facilitated the deposition of a full-coverage CuCl film. A dual beam (FIB/SEM) system with energy dispersive spectrometry facilities attached revealed a homogeneous CuCl layer with a flat-top surface and an average thickness of about 1 μm. Both the excitonic and biexcitonic emission lines were well-resolved in the 6.4 K photoluminescence spectra. In particular, the free exciton emission line was observable at room temperature, indicating the good quality of the CuCl films prepared by CBD. - Highlights: • Cuprous chloride (CuCl) was prepared on Cu substrate by chemical bath deposition. • HCl dip treatments facilitated the deposition of a full-coverage CuCl film. • A homogeneous elemental distribution was recognized for the deposited CuCl layer. • Excitonic and biexcitonic photoluminescence lines of CuCl films were well-resolved. • The free exciton emission line of CuCl films was observable at room temperature.

A novel reducing graphene/polyaniline/cuprous oxide composite hydrogel with unexpected photocatalytic activity for the degradation of Congo red

Energy Technology Data Exchange (ETDEWEB)

Miao, Jie; Xie, Anjian; Li, Shikuo; Huang, Fangzhi; Cao, Juan; Shen, Yuhua, E-mail: yhshen@ahu.edu.cn

2016-01-01

Graphical abstract: Excellent photocatalytic activity of the RGO/PANI/Cu{sub 2}O composite hydrogel for CR degradation under UV–vis light irradiation. - Highlights: • The RGO/PANI/Cu{sub 2}O composite hydrogel was first synthesized via a facile method. • Photocatalytic performance was studied under UV–vis light. • The ternary composite hydrogel shows unexpected photocatalytic activity. • A possible photocatalysis mechanism was illustrated. - Abstract: In this work, a novel reducing graphene/polyaniline/cuprous oxide (RGO/PANI/Cu{sub 2}O) composite hydrogel with a 3D porous network has been successfully prepared via a one-pot method in the presence of cubic Cu{sub 2}O nanoparticles. The as-synthesized ternary composites hydrogel shows unexpected photocatalytic activity such that Congo red (CR) degradation efficiency can reaches 97.91% in 20 min under UV–vis light irradiation, which is much higher than that of either the single component (Cu{sub 2}O nanoparticles), or two component systems (RGO/Cu{sub 2}O composite hydrogel and PANI/Cu{sub 2}O nanocomposites). Furthermore, the ternary composite hydrogel exhibits high stability and do not show any significant loss after five recycles. Such outstanding photocatalytic activity of the RGO/PANI/Cu{sub 2}O composite hydrogel was ascribed to the high absorption ability of the product for CR and the synergic effect among RGO, PANI and Cu{sub 2}O in photocatalytic process. The product of this work would provide a new sight for the construction of UV–vis light responsive photocatalyst with high performance.

NADPH oxidases as novel pharmacologic targets against influenza A virus infection.

Science.gov (United States)

Vlahos, Ross; Selemidis, Stavros

2014-12-01

Influenza A viruses represent a major global health care challenge, with imminent pandemics, emerging antiviral resistance, and long lag times for vaccine development, raising a pressing need for novel pharmacologic strategies that ideally target the pathology irrespective of the infecting strain. Reactive oxygen species (ROS) pervade all facets of cell biology with both detrimental and protective properties. Indeed, there is compelling evidence that activation of the NADPH oxidase 2 (NOX2) isoform of the NADPH oxidase family of ROS-producing enzymes promotes lung oxidative stress, inflammation, injury, and dysfunction resulting from influenza A viruses of low to high pathogenicity, as well as impeding virus clearance. By contrast, the dual oxidase isoforms produce ROS that provide vital protective antiviral effects for the host. In this review, we propose that inhibitors of NOX2 are better alternatives than broad-spectrum antioxidant approaches for treatment of influenza pathologies, for which clinical efficacy may have been limited owing to poor bioavailability and inadvertent removal of beneficial ROS. Finally, we briefly describe the current suite of NADPH oxidase inhibitors and the molecular features of the NADPH oxidase enzymes that could be exploited by drug discovery for development of more specific and novel inhibitors to prevent or treat disease caused by influenza. Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.

Confirmation of a blocked amino terminus of sulfhydryl oxidase

International Nuclear Information System (INIS)

Janolino, V.G.; Morrison-Rowe, S.J.; Swaisgood, H.E.

1990-01-01

The isolation of sulfhydryl oxidase from bovine milk in a suitably pure form for sequencing was carried out by transient covalent affinity chromatography of diafiltered whey using cysteinylsuccinamidopropyl-glass as matrix. The glutathione-eluted proteins were separated by SDS-PAGE. By radiolabeling the affinity chromatography-purified enzyme with [ 14 C]iodoacetate before subjecting to SDS-PAGE, the sulfhydryl oxidase band was identified, because sulfhydryl oxidase is known to be inactivated by alkylation of one sulfhydryl group per mole. The results confirmed that sulfhydryl oxidase corresponds to the 85 (± 5)-kDa band observed on SDS-PAGE. The protein band corresponding to radiolabeled sulfhydryl oxidase was recovered from SDS-PAGE gels by electrophoretic elution and by electroblotting on polyvinylidene difluoride membrane and subjected to gas phase sequencing. Precautions were taken during electrophoretic elution to prevent reactions that result in N-terminal blocking. Both methods of protein recovery yielded negative results when subjected to sequence analysis indicating that the N-terminus of sulfhydryl oxidase is blocked

Immobilization of oxidases and their analytical applications

International Nuclear Information System (INIS)

Yasinzai, M.

2007-01-01

Immobilized enzymes are replacing their soluble counter-parts in nearly every field of application. These enzyme modifications have evolved from a research curiosity into an entire branch of Biotechnology. An immobilization method for flavin containing oxidases and their use in flow injection system is described. An electrochemical detector for H/sub 2/O/sub 2/ is assembled which is used effectively for the determination of glucose using more common glucose oxidase and the simultaneous determination of sugars. The combination of oxidases with hydrolases have been used for the determination of maltose and starch. (author)

NADPH Oxidases: Progress and Opportunities

OpenAIRE

San Martin, Alejandra; Griendling, Kathy K.

2014-01-01

From the initial discovery in 1999 that NADPH oxidases comprise a family of enzymes to our current focus on drug development to treat multiple pathologies related to this enzyme family, progress has been swift and impressive. We have expanded our understanding of the extent of the family, the basic enzymatic biochemistry, the multiple cellular functions controlled by NADPH oxidases, and their varied roles in physiology and diseases. We have developed numerous cell culture tools, animal models...

Band offsets of n-type electron-selective contacts on cuprous oxide (Cu{sub 2}O) for photovoltaics

Energy Technology Data Exchange (ETDEWEB)

Brandt, Riley E., E-mail: rbrandt@alum.mit.edu, E-mail: buonassisi@mit.edu; Lee, Yun Seog; Buonassisi, Tonio, E-mail: rbrandt@alum.mit.edu, E-mail: buonassisi@mit.edu [Massachusetts Institute of Technology, Cambridge, Massachusetts 02139 (United States); Young, Matthew; Dameron, Arrelaine; Teeter, Glenn [National Renewable Energy Laboratory, Golden, Colorado 80401 (United States); Park, Helen Hejin; Chua, Danny; Gordon, Roy G. [Harvard University, Cambridge, Massachusetts 02139 (United States)

2014-12-29

The development of cuprous oxide (Cu{sub 2}O) photovoltaics (PVs) is limited by low device open-circuit voltages. A strong contributing factor to this underperformance is the conduction-band offset between Cu{sub 2}O and its n-type heterojunction partner or electron-selective contact. In the present work, a broad range of possible n-type materials is surveyed, including ZnO, ZnS, Zn(O,S), (Mg,Zn)O, TiO{sub 2}, CdS, and Ga{sub 2}O{sub 3}. Band offsets are determined through X-ray photoelectron spectroscopy and optical bandgap measurements. A majority of these materials is identified as having a negative conduction-band offset with respect to Cu{sub 2}O; the detrimental impact of this on open-circuit voltage (V{sub OC}) is evaluated through 1-D device simulation. These results suggest that doping density of the n-type material is important as well, and that a poorly optimized heterojunction can easily mask changes in bulk minority carrier lifetime. Promising heterojunction candidates identified here include Zn(O,S) with [S]/[Zn] ratios >70%, and Ga{sub 2}O{sub 3}, which both demonstrate slightly positive conduction-band offsets and high V{sub OC} potential. This experimental protocol and modeling may be generalized to evaluate the efficiency potential of candidate heterojunction partners for other PV absorbers, and the materials identified herein may be promising for other absorbers with low electron affinities.

Xanthine oxidase activity and free radical generation in patients with sepsis syndrome

DEFF Research Database (Denmark)

Galley, H F; Davies, Michael Jonathan; Webster, N R

1996-01-01

OBJECTIVE: To determine xanthine oxidase activity, free radical concentrations, and lipid peroxidation in patients with sepsis syndrome compared with noninfected critically ill patients. DESIGN: A prospective observational study. SETTING: A nine-bed intensive care unit in a university teaching......). CONCLUSIONS: Patients with sepsis have xanthine oxidase activation, high free-radical concentrations, and evidence of free radical damage. The finding that xanthine oxidase activity was lower in those patients who died, coupled with increased lactate concentrations implies more severe ischemia with incomplete...... to the Acute Physiology and Chronic Health Evaluation (APACHE) II score or to the presence of organ dysfunction. The mean ascorbyl radical concentration (arbitrary units) determined by electron paramagnetic resonance following spin trapping was increased in patients compared with healthy subjects (p

Carbon-layer-protected cuprous oxide nanowire arrays for efficient water reduction

KAUST Repository

Zhang, Zhonghai

2013-02-26

In this work, we propose a solution-based carbon precursor coating and subsequent carbonization strategy to form a thin protective carbon layer on unstable semiconductor nanostructures as a solution to the commonly occurring photocorrosion problem of many semiconductors. A proof-of-concept is provided by using glucose as the carbon precursor to form a protective carbon coating onto cuprous oxide (Cu2O) nanowire arrays which were synthesized from copper mesh. The carbon-layer-protected Cu2O nanowire arrays exhibited remarkably improved photostability as well as considerably enhanced photocurrent density. The Cu2O nanowire arrays coated with a carbon layer of 20 nm thickness were found to give an optimal water splitting performance, producing a photocurrent density of -3.95 mA cm-2 and an optimal photocathode efficiency of 0.56% under illumination of AM 1.5G (100 mW cm-2). This is the highest value ever reported for a Cu 2O-based electrode coated with a metal/co-catalyst-free protective layer. The photostability, measured as the percentage of the photocurrent density at the end of 20 min measurement period relative to that at the beginning of the measurement, improved from 12.6% on the bare, nonprotected Cu2O nanowire arrays to 80.7% on the continuous carbon coating protected ones, more than a 6-fold increase. We believe that the facile strategy presented in this work is a general approach that can address the stability issue of many nonstable photoelectrodes and thus has the potential to make a meaningful contribution in the general field of energy conversion. © 2013 American Chemical Society.

Modulation of NADPH oxidase activity by known uraemic retention solutes

DEFF Research Database (Denmark)

Schulz, Anna Marta; Terne, Cindy; Jankowski, Vera

2014-01-01

chloride (DPI), an inhibitor of NADPH oxidase. The effect on enzymatic activity of NADPH oxidase was quantified within an incubation time of 120 min. RESULTS: Thirty-nine of the 48 uraemic retention solutes tested had a significant decreasing effect on NADPH oxidase activity. Oxalate has been characterized......BACKGROUND: Uraemia and cardiovascular disease appear to be associated with an increased oxidative burden. One of the key players in the genesis of reactive oxygen species (ROS) is nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Based on initial experiments demonstrating a decreased...... inhibitory effect on NADPH oxidase activity in the presence of plasma from patients with CKD-5D after dialysis compared with before dialysis, we investigated the effect of 48 known and commercially available uraemic retention solutes on the enzymatic activity of NADPH oxidase. METHODS: Mononuclear leucocytes...

Gravity Responsive NADH Oxidase of the Plasma Membrane

Science.gov (United States)

Morre, D. James (Inventor)

2002-01-01

A method and apparatus for sensing gravity using an NADH oxidase of the plasma membrane which has been found to respond to unit gravity and low centrifugal g forces. The oxidation rate of NADH supplied to the NADH oxidase is measured and translated to represent the relative gravitational force exerted on the protein. The NADH oxidase of the plasma membrane may be obtained from plant or animal sources or may be produced recombinantly.

Salicylic Acid Regulation of Respiration in Higher Plants: Alternative Oxidase Expression.

Science.gov (United States)

Rhoads, DM; McIntosh, L

1992-01-01

Alternative respiratory pathway capacity increases during the development of the thermogenic appendix of a voodoo lily inflorescence. The levels of the alternative oxidase proteins increased dramatically between D-4 (4 days prior to the day of anthesis) and D-3 and continued to increase until the day of anthesis (D-day). The level of salicylic acid (SA) in the appendix is very low early on D-1, but increases to a high level in the evening of D-1. Thermogenesis occurs after a few hours of light on D-day. Therefore, the initial accumulation of the alternative oxidase proteins precedes the increase in SA by 3 days, indicating that other regulators may be involved. A 1.6-kb transcript encoding the alternative oxidase precursor protein accumulated to a high level in the appendix tissue by D-1. Application of SA to immature appendix tissue caused an increase in alternative pathway capacity and a dramatic accumulation of the alternative oxidase proteins and the 1.6-kb transcript. Time course experiments showed that the increase in capacity, protein levels, and transcript level corresponded precisely. The response to SA was blocked by cycloheximide or actinomycin D, indicating that de novo transcription and translation are required. However, nuclear, in vitro transcription assays indicated that the accumulation of the 1.6-kb transcript did not result from a simple increase in the rate of transcription of aox1. PMID:12297672

Hot or not? Discovery and characterization of a thermostable alditol oxidase from Acidothermus cellulolyticus 11B

NARCIS (Netherlands)

Winter, Remko T.; Heuts, Dominic P. H. M.; Rijpkema, Egon M. A.; van Bloois, Edwin; Wijma, Hein J.; Fraaije, Marco W.

We describe the discovery, isolation and characterization of a highly thermostable alditol oxidase from Acidothermus cellulolyticus 11B. This protein was identified by searching the genomes of known thermophiles for enzymes homologous to Streptomyces coelicolor A3(2) alditol oxidase (AldO). A gene

Multi-Copper Oxidases and Human Iron Metabolism

Science.gov (United States)

Vashchenko, Ganna; MacGillivray, Ross T. A.

2013-01-01

Multi-copper oxidases (MCOs) are a small group of enzymes that oxidize their substrate with the concomitant reduction of dioxygen to two water molecules. Generally, multi-copper oxidases are promiscuous with regards to their reducing substrates and are capable of performing various functions in different species. To date, three multi-copper oxidases have been detected in humans—ceruloplasmin, hephaestin and zyklopen. Each of these enzymes has a high specificity towards iron with the resulting ferroxidase activity being associated with ferroportin, the only known iron exporter protein in humans. Ferroportin exports iron as Fe2+, but transferrin, the major iron transporter protein of blood, can bind only Fe3+ effectively. Iron oxidation in enterocytes is mediated mainly by hephaestin thus allowing dietary iron to enter the bloodstream. Zyklopen is involved in iron efflux from placental trophoblasts during iron transfer from mother to fetus. Release of iron from the liver relies on ferroportin and the ferroxidase activity of ceruloplasmin which is found in blood in a soluble form. Ceruloplasmin, hephaestin and zyklopen show distinctive expression patterns and have unique mechanisms for regulating their expression. These features of human multi-copper ferroxidases can serve as a basis for the precise control of iron efflux in different tissues. In this manuscript, we review the biochemical and biological properties of the three human MCOs and discuss their potential roles in human iron homeostasis. PMID:23807651

Calcium transport in vesicles energized by cytochrome oxidase

Energy Technology Data Exchange (ETDEWEB)

Rosier, Randy N. [Univ. of Rochester, NY (United States)

1979-01-01

Experiments on the reconstitution of cytochrome oxidase into phospholipid vesicles were carried out using techniques of selectivity energizing the suspensions with ascorbate and cytochrome c or ascorbate, PMS, and internally trapped cytochrome c. It was found that the K⁺ selective ionophore valinomycin stimulated the rate of respiration of cytochrome oxidase vesicles regardless of the direction of the K⁺ flux across the vesicle membranes. The stimulation occurred in the presence of protonophoric uncouplers and in the complete absence of potassium or in detergent-lysed suspensions. Gramicidin had similar effects and it was determined that the ionophores acted by specific interaction with cytochrome oxidase rather than by the previously assumed collapse of membrane potentials. When hydrophobic proteins and appropriate coupling factors were incorporated into the cytochrome oxidase, vesicles phosphorylation of ADP could be coupled to the oxidation reaction of cytochrome oxidase. Relatively low P:O, representing poor coupling of the system, were problematical and precluded measurements of protonmotive force. However the system was used to study ion translocation.

Cytotoxicity of cuprous oxide nanoparticles to fish blood cells: hemolysis and internalization

Energy Technology Data Exchange (ETDEWEB)

Chen Liqiang, E-mail: chenlq@ynu.edu.cn; Kang Bin [Yunnan University, Asian International Rivers Center, Yunnan Key Laboratory of International Rivers and Trans-boundary Eco-security (China); Ling Jian [Yunnan University, College of Chemistry and Chemical Engineering (China)

2013-03-15

Cuprous oxide nanoparticles (Cu{sub 2}O NPs) possess unique physical and chemical properties which are employed in a broad variety of applications. However, little is known about the adverse effects of Cu{sub 2}O NPs on organisms. In the current study, in vitro cytotoxicity of Cu{sub 2}O NPs (ca. 60 nm in diameter) to the blood cells of freshwater fish Carassius auratus was evaluated. A concentration-dependent hemolytic activity of Cu{sub 2}O NPs to red blood cells (RBCs) and the phagocytosis of Cu{sub 2}O NPs by leukocytes were revealed. The results showed that dosages of Cu{sub 2}O NPs greater than 40 {mu}g/mL were toxic to blood cells, and could cause serious membrane damage to RBCs. The EC{sub 50} value of Cu{sub 2}O NPs as obtained from RBCs and whole blood exposure was 26 and 63 {mu}g/mL, respectively. The generation of reactive oxygen species and the direct interaction between Cu{sub 2}O NPs and the cell membrane were suggested as the possible mechanism for cytotoxicity, and the intrinsic hemolytic active of Cu{sub 2}O NPs was the main contributor to the toxicity rather than solubilized copper ions. The adsorption of plasma proteins on the surfaces of Cu{sub 2}O NPs led to their aggregation in whole blood, and aggregate formation can significantly alleviate the hemolytic effect and subsequently mediate the phagocytosis of Cu{sub 2}O NPs by leukocytes.

Ultra-high-throughput screening method for the directed evolution of glucose oxidase.

Science.gov (United States)

Ostafe, Raluca; Prodanovic, Radivoje; Nazor, Jovana; Fischer, Rainer

2014-03-20

Glucose oxidase (GOx) is used in many industrial processes that could benefit from improved versions of the enzyme. Some improvements like higher activity under physiological conditions and thermal stability could be useful for GOx applications in biosensors and biofuel cells. Directed evolution is one of the currently available methods to engineer improved GOx variants. Here, we describe an ultra-high-throughput screening system for sorting the best enzyme variants generated by directed evolution that incorporates several methodological refinements: flow cytometry, in vitro compartmentalization, yeast surface display, fluorescent labeling of the expressed enzyme, delivery of glucose substrate to the reaction mixture through the oil phase, and covalent labeling of the cells with fluorescein-tyramide. The method enables quantitative screening of gene libraries to identify clones with improved activity and it also allows cells to be selected based not only on the overall activity but also on the specific activity of the enzyme. Copyright © 2014 Elsevier Ltd. All rights reserved.

Significance of membrane bioreactor design on the biocatalytic performance of glucose oxidase and catalase: Free vs. immobilized enzyme systems

DEFF Research Database (Denmark)

Morthensen, Sofie Thage; Meyer, Anne S.; Jørgensen, Henning

2017-01-01

Membrane separation of xylose and glucose can be accomplished via oxidation of glucose to gluconic acid by enzymatic glucose oxidase catalysis. Oxygen for this reaction can be supplied via decomposition of hydrogen peroxide by enzymatic catalase catalysis. In order to maximize the biocatalytic...... productivity of glucose oxidase and catalase (gluconic acid yield per total amount of enzyme) the following system set-ups were compared: immobilization of glucose oxidase alone; co-immobilization of glucose oxidase and catalase; glucose oxidase and catalase free in the membrane bioreactor. Fouling......-induced enzyme immobilization in the porous support of an ultrafiltration membrane was used as strategy for entrapment of glucose oxidase and catalase. The biocatalytic productivity of the membrane reactor was found to be highly related to the oxygen availability, which in turn depended on the reactor...

High electro-catalytic activities of glucose oxidase embedded one-dimensional ZnO nanostructures

International Nuclear Information System (INIS)

Sarkar, Nirmal K; Bhattacharyya, Swapan K

2013-01-01

One-dimensional ZnO nanorods and nanowires are separately synthesized on Zn substrate by simple hydrothermal processes at low temperatures. Electro-catalytic responses of glucose oxidase/ZnO/Zn electrodes using these two synthesized nanostructures of ZnO are reported and compared with others available in literature. It is apparent the Michaelis–Menten constant, K M app , for the present ZnO nanowire, having a greater aspect ratio, is found to be the lowest when compared with others. This sensor shows lower oxidation peak potential with a long detection range of 6.6 μM–380 mM and the highest sensitivity of ∼35.1 μA cm −2 mM −1 , among the reported values in the literature. Enzyme catalytic efficiency and turnover numbers are also found to be remarkably high. (paper)

Modulation of NADPH oxidase activity by known uraemic retention solutes.

Science.gov (United States)

Schulz, Anna Marta; Terne, Cindy; Jankowski, Vera; Cohen, Gerald; Schaefer, Mandy; Boehringer, Falko; Tepel, Martin; Kunkel, Desiree; Zidek, Walter; Jankowski, Joachim

2014-08-01

Uraemia and cardiovascular disease appear to be associated with an increased oxidative burden. One of the key players in the genesis of reactive oxygen species (ROS) is nicotinamide adenine dinucleotide phosphate (NADPH) oxidase. Based on initial experiments demonstrating a decreased inhibitory effect on NADPH oxidase activity in the presence of plasma from patients with CKD-5D after dialysis compared with before dialysis, we investigated the effect of 48 known and commercially available uraemic retention solutes on the enzymatic activity of NADPH oxidase. Mononuclear leucocytes isolated from buffy coats of healthy volunteers were isolated, lysed and incubated with NADH in the presence of plasma from healthy controls and patients with CKD-5D. Furthermore, the leucocytes were lysed and incubated in the presence of uraemic retention solute of interest and diphenyleneiodonium chloride (DPI), an inhibitor of NADPH oxidase. The effect on enzymatic activity of NADPH oxidase was quantified within an incubation time of 120 min. Thirty-nine of the 48 uraemic retention solutes tested had a significant decreasing effect on NADPH oxidase activity. Oxalate has been characterized as the strongest inhibitor of NADPH oxidase (90% of DPI inhibition). Surprisingly, none of the uraemic retention solutes we investigated was found to increase NADPH oxidase activity. Furthermore, plasma from patients with CKD-5D before dialysis caused significantly higher inhibitory effect on NADPH oxidase activity compared with plasma from healthy subjects. However, this effect was significantly decreased in plasma from patients with CKD-5D after dialysis. The results of this study show that uraemic retention solutes modulated the activity of the NADPH oxidase. The results of this study might be the basis for the development of inhibitors applicable as drug in the situation of increased oxidative stress. © 2014 Stichting European Society for Clinical Investigation Journal Foundation.

Effect of cuprous oxide with different sizes on thermal and combustion behaviors of unsaturated polyester resin.

Science.gov (United States)

Hou, Yanbei; Hu, Weizhao; Gui, Zhou; Hu, Yuan

2017-07-15

Cuprous oxide (Cu 2 O) as an effective catalyst has been applied to enhance the fire safety of unsaturated polyester resin (UPR), but the particle size influence on combustion behaviors has not been previously reported. Herein, the UPR/Cu 2 O composites (metal oxide particles with average particle-size of 10, 100, and 200nm) were successfully synthesized by thermosetting process. The effects of Cu 2 O with different sizes on thermostability and combustion behaviors of UPR were characterized by TGA, MCC, TG-IR, FTIR, and SSTF. The results revel that the addition of Cu 2 O contributes to sufficient decomposition of oxygen-containing compounds, which is beneficial to the release of nontoxic compounds. The smallest-sized Cu 2 O performs the excellent catalytic decomposition effect and promotes the complete combustion of UPR, which benefits the enhancement of fire safety. While the other additives retard pyrolysis process and yield more char residue, and thus the flame retardancy of UPR composites was improved. Therefore, catalysis plays a major role for smaller-sized particles during thermal decomposition of matrix, while flame retarded effect became gradual distinctly for the larger-sized additives. Copyright © 2017 Elsevier B.V. All rights reserved.

Lysyl oxidase in colorectal cancer

DEFF Research Database (Denmark)

Cox, Thomas R; Erler, Janine T

2013-01-01

Colorectal cancer is the third most prevalent form of cancer worldwide and fourth-leading cause of cancer-related mortality, leading to ~600,000 deaths annually, predominantly affecting the developed world. Lysyl oxidase is a secreted, extracellular matrix-modifying enzyme previously suggested...... to act as a tumor suppressor in colorectal cancer. However, emerging evidence has rapidly implicated lysyl oxidase in promoting metastasis of solid tumors and in particular colorectal cancer at multiple stages, affecting tumor cell proliferation, invasion, and angiogenesis. This emerging research has...... advancements in the field of colorectal cancer....

Plasma diamine oxidase activity in asthmatic children

Directory of Open Access Journals (Sweden)

Kyoichiro Toyoshima

1996-01-01

Full Text Available Histamine plays an important role in the development of asthmatic symptoms. Diamine oxidase (DAO histaminase, which inactivates histamine, is located in the intestine and kidney and is released into plasma. Plasma DAO activity in asthmatic children was measured by a recently developed high performance liquid chromatographic method using histamine as the DAO substrate. Diamine oxidase activity was higher in severely asthmatic children than in those with mild asthma. A time course study during the acute exacerbation phase revealed that DAO activity rose during acute asthmatic attacks and then decreased gradually over several days. Although the mechanisms of plasma DAO activity increase during acute asthmatic attacks could not be explained, data showed that plasma DAO activity is an important index of histamine metabolism in asthmatics and may relate to some mechanisms of acute exacerbation of airway inflammation. Consequently, fluctuations in plasma DAO can be used as one of various indices of instability in management of asthma.

Chemoenzymatic combination of glucose oxidase with titanium silicalite -1

DEFF Research Database (Denmark)

Vennestrøm, Peter Nicolai Ravnborg; Taarning, Esben; Christensen, Claus H.

2010-01-01

Zeozymes: A proof-of-concept is presented for the chemoenzymatic combination of titanium silicalite-1 zeolite with glucose oxidase. In this combination, glucose is oxidized to gluconic acid and the H2O2 byproduct formed in situ is used for the simultaneous oxidation of chemical substrates. Both...... a soluble glucose oxidase and a truly integrated heterogeneous combination whereby the oxidase enzyme is anchored onto the zeolite surface are reported....

Passivation Of High-Temperature Superconductors

Science.gov (United States)

Vasquez, Richard P.

1991-01-01

Surfaces of high-temperature superconductors passivated with native iodides, sulfides, or sulfates formed by chemical treatments after superconductors grown. Passivating compounds nearly insoluble in and unreactive with water and protect underlying superconductors from effects of moisture. Layers of cuprous iodide and of barium sulfate grown. Other candidate passivating surface films: iodides and sulfides of bismuth, strontium, and thallium. Other proposed techniques for formation of passivating layers include deposition and gas-phase reaction.

Amine oxidases as important agents of pathological processes of rhabdomyolysis in rats.

Science.gov (United States)

Gudkova, O O; Latyshko, N V; Shandrenko, S G

2016-01-01

In this study we have tested an idea on the important role of amine oxidases (semicarbazide-sensitive amine oxidase, diamine oxidase, polyamine oxidase) as an additional source of oxidative/carbonyl stress under glycerol-induced rhabdomyolysis, since the enhanced formation of reactive oxygen species and reactive carbonyl species in a variety of tissues is linked to various diseases. In our experiments we used the sensitive fluorescent method devised for estimation of amine oxidases activity in the rat kidney and thymus as targeted organs under rhabdomyolysis. We have found in vivo the multiple rises in activity of semicarbazide-sensitive amine oxidase, diamine oxidase, polyamine oxidase (2-4.5 times) in the corresponding cell fractions, whole cells or their lysates at the 3-6th day after glycerol injection. Aberrant antioxidant activities depended on rhabdomyolysis stage and had organ specificity. Additional treatment of animals with metal chelator ‘Unithiol’ adjusted only the activity of antioxidant enzymes but not amine oxidases in both organs. Furthermore the in vitro experiment showed that Fenton reaction (hydrogen peroxide in the presence of iron) products alone had no effect on semicarbazide-sensitive amine oxidase activity in rat liver cell fraction whereas supplementation with methylglyoxal resulted in its significant 2.5-fold enhancement. Combined action of the both agents had additive effect on semicarbazide-sensitive amine oxidase activity. We can assume that biogenic amine and polyamine catabolism by amine oxidases is upregulated by oxidative and carbonyl stress factors directly under rhabdomyolysis progression, and the increase in catabolic products concentration contributes to tissue damage in glycerol-induced acute renal failure and apoptosis stimulation in thymus.

Laboratory-evolved vanillyl-alcohol oxidase produces natural vanillin

NARCIS (Netherlands)

Heuvel, van den R.H.H.; Berg, van den W.A.M.; Rovida, S.; Berkel, van W.J.H.

2004-01-01

The flavoenzyme vanillyl-alcohol oxidase was subjected to random mutagenesis to generate mutants with enhanced reactivity to creosol (2-methoxy-4-methylphenol). The vanillyl-alcohol oxidase-mediated conversion of creosol proceeds via a two-step process in which the initially formed vanillyl alcohol

In Situ Enzymatically Generated Photoswitchable Oxidase Mimetics and Their Application for Colorimetric Detection of Glucose Oxidase.

Science.gov (United States)

Cao, Gen-Xia; Wu, Xiu-Ming; Dong, Yu-Ming; Li, Zai-Jun; Wang, Guang-Li

2016-07-09

In this study, a simple and amplified colorimetric assay is developed for the detection of the enzymatic activity of glucose oxidase (GOx) based on in situ formation of a photoswitchable oxidase mimetic of PO₄(3-)-capped CdS quantum dots (QDs). GOx catalyzes the oxidation of 1-thio-β-d-glucose to give 1-thio-β-d-gluconic acid which spontaneously hydrolyzes to β-d-gluconic acid and H₂S; the generated H₂S instantly reacts with Cd(2+) in the presence of Na₃PO₄ to give PO₄(3-)-stabilized CdS QDs in situ. Under visible-light (λ ≥ 400 nm) stimulation, the PO₄(3-)-capped CdS QDs are a new style of oxidase mimic derived by producing some active species, such as h⁺, (•)OH, O₂(•-) and a little H₂O₂, which can oxidize the typical substrate (3,3,5,5-tetramethylbenzydine (TMB)) with a color change. Based on the GOx-triggered growth of the oxidase mimetics of PO₄(3-)-capped CdS QDs in situ, we developed a simple and amplified colorimetric assay to probe the enzymatic activity of GOx. The proposed method allowed the detection of the enzymatic activity of GOx over the range from 25 μg/L to 50 mg/L with a low detection limit of 6.6 μg/L. We believe the PO₄(3-)-capped CdS QDs generated in situ with photo-stimulated enzyme-mimicking activity may find wide potential applications in biosensors.

Astragaloside IV prevents damage to human mesangial cells through the inhibition of the NADPH oxidase/ROS/Akt/NF‑κB pathway under high glucose conditions.

Science.gov (United States)

Sun, Li; Li, Weiping; Li, Weizu; Xiong, Li; Li, Guiping; Ma, Rong

2014-07-01

Glomerular hypertrophy and hyperfiltration are the two major pathological characteristics of the early stages of diabetic nephropathy (DN), which are respectively related to mesangial cell (MC) proliferation and a decrease in calcium influx conducted by canonical transient receptor potential cation channel 6 (TRPC6). The marked increase in the production of reactive oxygen species (ROS) induced by hyperglycemia is the main sponsor of multiple pathological pathways in DN. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase is an important source of ROS production in MCs. Astragaloside IV (AS‑IV) is an active ingredient of Radix Astragali which has a potent antioxidative effect. In this study, we aimed to investigate whether high glucose (HG)‑induced NADPH oxidase activation and ROS production contribute to MC proliferation and the downregulation of TRPC6 expression; we also wished to determine the effects of AS‑IV on MCs under HG conditions. Using a human glomerular mesangial cell line, we found that treatment with AS‑IV for 48 h markedly attenuated HG‑induced proliferation and the hypertrophy of MCs in a dose‑dependent manner. The intracellular ROS level was also markedly reduced following treatment with AS‑IV. In addition, the enhanced activity of NADPH oxidase and the expression level of NADPH oxidase 4 (Nox4) protein were decreased. Treatment with AS‑IV also inhibited the phosphorylation level of Akt and IκBα in the MCs. In addition, TRPC6 protein expression and the intracellular free calcium concentration were also markedly reduced following treatment with AS‑IV under HG conditions. These results suggest that AS‑IV inhibits HG‑induced mesangial cell proliferation and glomerular contractile dysfunction through the NADPH oxidase/ROS/Akt/nuclear factor‑κB (NF‑κB) pathway, providing a new perspective for the clinical treatment of DN.

Thermal Characterization of Purified Glucose Oxidase from A Newly Isolated Aspergillus Niger UAF-1

Science.gov (United States)

Anjum Zia, Muhammad; Khalil-ur-Rahman; K. Saeed, Muhammad; Andaleeb, Fozia; I. Rajoka, Muhammad; A. Sheikh, Munir; A. Khan, Iftikhar; I. Khan, Azeem

2007-01-01

An intracellular glucose oxidase was isolated from the mycelium extract of a locally isolated strain of Aspergillus niger UAF-1. The enzyme was purified to a yield of 28.43% and specific activity of 135 U mg−1 through ammonium sulfate precipitation, anion exchange and gel filtration chromatography. The enzyme showed high affinity for D-glucose with a Km value of 2.56 mM. The enzyme exhibited optimum catalytic activity at pH 5.5. Temperature optimum for glucose oxidase, catalyzed D-glucose oxidation was 40°C. The enzyme showed a high thermostability having a half-life 30 min, enthalpy of denaturation 99.66 kJ mol−1 and free energy of denaturation 103.63 kJ mol−1. These characteristics suggest the use of glucose oxidase from Aspergillus niger UAF-1 as an analytical reagent and in the design of biosensors for clinical, biochemical and diagnostic assays. PMID:18193107

Layer-by-Layer Assembly of Glucose Oxidase on Carbon Nanotube Modified Electrodes.

Science.gov (United States)

Suroviec, Alice H

2017-01-01

The use of enzymatically modified electrodes for the detection of glucose or other non-electrochemically active analytes is becoming increasingly common. Direct heterogeneous electron transfer to glucose oxidase has been shown to be kinetically difficult, which is why electron transfer mediators or indirect detection is usually used for monitoring glucose with electrochemical sensors. It has been found, however, that electrodes modified with single or multi-walled carbon nanotubes (CNTs) demonstrate fast heterogeneous electron transfer kinetics as compared to that found for traditional electrodes. Incorporating CNTs into the assembly of electrochemical glucose sensors, therefore, affords the possibility of facile electron transfer to glucose oxidase, and a more direct determination of glucose. This chapter describes the methods used to use CNTs in a layer-by-layer structure along with glucose oxidase to produce an enzymatically modified electrode with high turnover rates, increased stability and shelf-life.

RXR agonists inhibit high glucose-induced upregulation of inflammation by suppressing activation of the NADPH oxidase-nuclear factor-κB pathway in human endothelial cells.

Science.gov (United States)

Ning, R B; Zhu, J; Chai, D J; Xu, C S; Xie, H; Lin, X Y; Zeng, J Z; Lin, J X

2013-12-13

An inflammatory response induced by high glucose is a cause of endothelial dysfunction in diabetes and is an important contributing link to atherosclerosis. Diabetes is an independent risk factor of atherosclerosis and activation of retinoid X receptor (RXR) has been shown to exert anti-atherogenic effects. In the present study, we examined the effects of the RXR ligands 9-cis-retinoic acid (9-cis-RA) and SR11237 on high glucose-induced inflammation in human umbilical endothelial vein endothelial cells (HUVECs) and explored the potential mechanism. Our results showed that the inflammation induced by high-glucose in HUVECs was mainly mediated by the activation of nuclear factor-B (NF- κB). High glucose-induced expression of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) were in comparison, significantly decreased by treatment with RXR. The effect of RXR agonists was mainly due to the inhibition of NF-κB activation. Using pharmacological inhibitors and siRNA, we confirmed that nicotinamide adenine dinucleotide phosphate (NADPH) oxidase was an upstream activator of NF-κB. Furthermore, RXR agonists significantly inhibited high glucose-induced activation of NADPH oxidase and significantly decreased the production of reactive oxygen species (ROS). To explore whether the rapid inhibitory effects of RXR agonists were in fact mediated by RXR, we examined the effect of RXR downregulation by RXR siRNA. Our results showed that RXR siRNA largely abrogated the effects of RXR agonists, suggesting the requirement of RXR expression. Therefore, we have shown that RXR is involved in the regulation of NADPH oxidase- NF-κB signal pathway, as the RXR ligands antagonized the inflammatory response in HUVECs induced by high glucose.

Monoamine Oxidase Inhibitors (MAOIs)

Science.gov (United States)

... health-medications/index.shtml. Accessed May 16, 2016. Hirsch M, et al. Monoamine oxidase inhibitors (MAOIs) for ... www.uptodate.com/home. Accessed May 16, 2016. Hirsch M, et al. Discontinuing antidepressant medications in adults. ...

Functional Assembly of Soluble and Membrane Recombinant Proteins of Mammalian NADPH Oxidase Complex.

Science.gov (United States)

Souabni, Hajer; Ezzine, Aymen; Bizouarn, Tania; Baciou, Laura

2017-01-01

Activation of phagocyte cells from an innate immune system is associated with a massive consumption of molecular oxygen to generate highly reactive oxygen species (ROS) as microbial weapons. This is achieved by a multiprotein complex, the so-called NADPH oxidase. The activity of phagocyte NADPH oxidase relies on an assembly of more than five proteins, among them the membrane heterodimer named flavocytochrome b 558 (Cytb 558 ), constituted by the tight association of the gp91 phox (also named Nox2) and p22 phox proteins. The Cytb 558 is the membrane catalytic core of the NADPH oxidase complex, through which the reducing equivalent provided by NADPH is transferred via the associated prosthetic groups (one flavin and two hemes) to reduce dioxygen into superoxide anion. The other major proteins (p47 phox , p67 phox , p40 phox , Rac) requisite for the complex activity are cytosolic proteins. Thus, the NADPH oxidase functioning relies on a synergic multi-partner assembly that in vivo can be hardly studied at the molecular level due to the cell complexity. Thus, a cell-free assay method has been developed to study the NADPH oxidase activity that allows measuring and eventually quantifying the ROS generation based on optical techniques following reduction of cytochrome c. This setup is a valuable tool for the identification of protein interactions, of crucial components and additives for a functional enzyme. Recently, this method was improved by the engineering and the production of a complete recombinant NADPH oxidase complex using the combination of purified proteins expressed in bacterial and yeast host cells. The reconstitution into artificial membrane leads to a fully controllable system that permits fine functional studies.

NADPH Oxidase-Mediated ROS Production Determines Insulin's Action on the Retinal Microvasculature.

Science.gov (United States)

Kida, Teruyo; Oku, Hidehiro; Horie, Taeko; Matsuo, Junko; Kobayashi, Takatoshi; Fukumoto, Masanori; Ikeda, Tsunehiko

2015-10-01

To determine whether insulin induces nitric oxide (NO) formation in retinal microvessels and to examine the effects of high glucose on the formation of NO. Freshly isolated rat retinal microvessels were incubated in normal (5.5 mM) or high (20 mM) glucose with or without insulin (100 nM). The levels of insulin-induced NO and reactive oxygen species (ROS) in the retinal microvessels were determined semiquantitatively using fluorescent probes, 4,5-diaminofluorescein diacetate, and hydroethidine, respectively, and a laser scanning confocal microscope. The insulin-induced changes of NO in rat retinal endothelial cells and pericytes cultured at different glucose concentrations (5.5 and 25 mM) were determined using flow cytometry. Nitric oxide synthase (NOS) protein levels were determined by Western blot analysis; intracellular levels of ROS were determined using fluorescence-activated cell sorting (FACS) analysis of ethidium fluorescence; and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase RNA expression was quantified using real-time PCR. Exposure of microvessels to insulin under normal glucose conditions led to a significant increase in NO levels; however, this increase was significantly suppressed when the microvessels were incubated under high glucose conditions. Intracellular levels of ROS were significantly increased in both retinal microvessels and cultured microvascular cells under high glucose conditions. The expression of NOS and NADPH oxidase were significantly increased in endothelial cells and pericytes under high glucose conditions. The increased formation of NO by insulin and its suppression by high glucose conditions suggests that ROS production mediated by NADPH oxidase is important by insulin's effect on the retinal microvasculature.

Optimization of glucose oxidase production by Aspergillus niger

African Journals Online (AJOL)

user

2011-02-28

Feb 28, 2011 ... manganese, cobalt, thioglycolic acid, and gluconic acid according to (Liu et al., .... In this experiment duplicate media of glucose 10% were adjusted at different ... Glucose oxidase as a pharmaceutical anti oxidant Drug. Devt. ... Plush KS, Hellmuth K, Rinas U (1996). kinetics of glucose oxidase excretion by ...

Cytochrome cbb3 of Thioalkalivibrio is a Na+-pumping cytochrome oxidase

NARCIS (Netherlands)

Muntyan, M.S.; Cherepanov, D.A.; Malinen, A.M.; Bloch, D.A.; Sorokin, D.Y.; Severina, I.I.; Ivashina, T.V.; Lahti, R.; Muyzer, G.; Skulachev, V.P.

2015-01-01

Cytochrome c oxidases (Coxs) are the basic energy transducers in the respiratory chain of the majority of aerobic organisms. Coxs studied to date are redox-driven proton-pumping enzymes belonging to one of three subfamilies: A-, B-, and C-type oxidases. The C-type oxidases (cbb3 cytochromes), which

Amide group anchored glucose oxidase based anodic catalysts for high performance enzymatic biofuel cell

Science.gov (United States)

Chung, Yongjin; Ahn, Yeonjoo; Kim, Do-Heyoung; Kwon, Yongchai

2017-01-01

A new enzyme catalyst is formed by fabricating gold nano particle (GNP)-glucose oxidase (GOx) clusters that are then attached to polyethyleneimine (PEI) and carbon nanotube (CNT) with cross-linkable terephthalaldehyde (TPA) (TPA/[CNT/PEI/GOx-GNP]). Especially, amide bonds belonging to TPA play an anchor role for incorporating rigid bonding among GNP, GOx and CNT/PEI, while middle size GNP is well bonded with thiol group of GOx to form strong GNP-GOx cluster. Those bonds are identified by chemical and electrochemical characterizations like XPS and cyclic voltammogram. The anchording effect of amide bonds induces fast electron transfer and strong chemical bonding, resulting in enhancements in (i) catalytic activity, (ii) amount of immobilized GOx and (ii) performance of enzymatic biofuel cell (EBC) including the catalyst. Regarding the catalytic activity, the TPA/[CNT/PEI/GOx-GNP] produces high electron transfer rate constant (6 s-1), high glucose sensitivity (68 μA mM-1 cm-2), high maximum current density (113 μA cm-2), low charge transfer resistance (17.0 Ω cm2) and long-lasting durability while its chemical structure is characterized by XPS confirming large portion of amide bond. In EBC measurement, it has high maximum power density (0.94 mW cm-2) compatible with catalytic acitivity measurements.

Serum diamine oxidase activity in patients with histamine intolerance.

Science.gov (United States)

Manzotti, G; Breda, D; Di Gioacchino, M; Burastero, S E

2016-03-01

Intolerance to various foods, excluding bona fide coeliac disease and lactose intolerance, represents a growing cause of patient visits to allergy clinics.Histamine intolerance is a long-known, multifaceted clinical condition triggered by histamine-rich foods and alcohol and/or by drugs that liberate histamine or block diamine oxidase (DAO), the main enzyme involved in the metabolism of ingested histamine. Histamine limitation diets impose complex, non-standardized restrictions that may severely impact the quality of life of patients. We retrospectively evaluated 14 patients who visited allergy outpatient facilities in northern Italy with a negative diagnosis for IgE-mediated food hypersensitivity, coeliac disease, conditions related to gastric hypersecretion, and systemic nickel hypersensitivity, and who previously underwent a histamine limitation diet with benefits for their main symptoms. Serum diamine oxidase levels and the clinical response to diamine oxidase supplementation were investigated. We found that 10 out of 14 patients had serum DAO activityintolerance. Moreover, 13 out of 14 patients subjectively reported a benefit in at least one of the disturbances related to food intolerances following diamine oxidase supplementation. The mean value (±SD) of diamine oxidase activity in the cohort of patients with histamine intolerance symptoms was 7.04±6.90 U/mL compared to 39.50±18.16 U/mL in 34 healthy controls (P=0.0031). In patients with symptoms triggered by histamine-rich food, measuring the serum diamine oxidase activity can help identify subjects who can benefit from a histamine limitation diet and/or diamine oxidase supplementation.Properly designed, controlled studies investigating histamine intolerance that include histamine provocation are indispensable for providing insights into the area of food intolerances, which are currently primarily managed with non-scientific approaches in Italy. © The Author(s) 2015.

Structure and activity of Aspergillus nidulans copper amine oxidase

DEFF Research Database (Denmark)

McGrath, Aaron P; Mithieux, Suzanne M; Collyer, Charles A

2011-01-01

Aspergillus nidulans amine oxidase (ANAO) has the unusual ability among the family of copper and trihydroxyphenylalanine quinone-containing amine oxidases of being able to oxidize the amine side chains of lysine residues in large peptides and proteins. We show here that in common with the related...... enzyme from the yeast Pichia pastoris, ANAO can promote the cross-linking of tropoelastin and oxidize the lysine residues in α-casein proteins and tropoelastin. The crystal structure of ANAO, the first for a fungal enzyme in this family, has been determined to a resolution of 2.4 Å. The enzyme is a dimer...... with the archetypal fold of a copper-containing amine oxidase. The active site is the most open of any of those of the structurally characterized enzymes in the family and provides a ready explanation for its lysine oxidase-like activity....

One-step synthesis and properties of monolithic photoluminescent ruby colored cuprous oxide antimony oxide glass nanocomposites

Energy Technology Data Exchange (ETDEWEB)

Som, Tirtha [Glass Science and Technology Section, Glass Division, Central Glass and Ceramic Research Institute, Council of Scientific and Industrial Research (CSIR, India), 196, Raja S.C. Mullick Road, Kolkata 700032 (India); Karmakar, Basudeb, E-mail: basudebk@cgcri.res.in [Glass Science and Technology Section, Glass Division, Central Glass and Ceramic Research Institute, Council of Scientific and Industrial Research (CSIR, India), 196, Raja S.C. Mullick Road, Kolkata 700032 (India)

2011-04-14

Research highlights: > Single-step synthesis of Cu{sub 2}O, Cu{sub y}Sb{sub 2-x}(O,OH){sub 6-7} (y {<=} 2, x {<=} 1) and Cu nanocrystals co-doped novel antimony oxide glass hybrid nanocomposites. > Yellow and orange colored nanocomposites shows size-controlled band gap shift of Cu{sub 2}O. > Red nanocomposite exhibits surface plasmon resonance band due to metallic Cu. > They exhibit broad deep-red photoluminescence emission under various UV excitation wavelengths. - Abstract: Cuprous oxide (Cu{sub 2}O) antimony glass (K{sub 2}O-B{sub 2}O{sub 3}-Sb{sub 2}O{sub 3}) monolithic nanocomposites having brilliant yellow to ruby red color have been synthesized by a single-step melt-quench technique involving in situ thermochemical reduction of Cu{sup 2+} (CuO) by the reducing glass matrix without using any external reducing agent. The X-ray diffraction (XRD), infrared transmission and reflection spectra, and selected area electron diffraction analysis support the reduction of Cu{sup 2+} to Cu{sup +} with the formation of Cu{sub 2}O nanoclusters along with Cu{sub y}Sb{sub 2-x}(O,OH){sub 6-7} (y {<=} 2, x {<=} 1) nanocrystalline phases while Cu{sup 0} nanoclusters are formed at very high Cu concentration. The UV-vis spectra of the yellow and orange colored nanocomposites show size-controlled band gap shift of the semiconductor (Cu{sub 2}O) nanocrystallites embedded in the glasses while the red nanocomposite exhibits surface plasmon resonance band at 529 nm due to metallic Cu. Transmission electron microscopic image advocates the formation of nanocystallites (5-42 nm). Photoluminescence emission studies show broad red emission band around 626 nm under various excitation wavelengths from 210 to 270 nm.

Expression and Chloroplast Targeting of Cholesterol Oxidase in Transgenic Tobacco Plants

Science.gov (United States)

Corbin, David R.; Grebenok, Robert J.; Ohnmeiss, Thomas E.; Greenplate, John T.; Purcell, John P.

2001-01-01

Cholesterol oxidase represents a novel type of insecticidal protein with potent activity against the cotton boll weevil (Anthonomus grandis grandis Boheman). We transformed tobacco (Nicotiana tabacum) plants with the cholesterol oxidase choM gene and expressed cytosolic and chloroplast-targeted versions of the ChoM protein. Transgenic leaf tissues expressing cholesterol oxidase exerted insecticidal activity against boll weevil larvae. Our results indicate that cholesterol oxidase can metabolize phytosterols in vivo when produced cytosolically or when targeted to chloroplasts. The transgenic plants exhibiting cytosolic expression accumulated low levels of saturated sterols known as stanols, and displayed severe developmental aberrations. In contrast, the transgenic plants expressing chloroplast-targeted cholesterol oxidase maintained a greater accumulation of stanols, and appeared phenotypically and developmentally normal. These results are discussed within the context of plant sterol distribution and metabolism. PMID:11457962

High throughput Screening to Identify Natural Human Monoamine Oxidase B Inhibitors

Science.gov (United States)

Mazzio, E; Deiab, S; Park, K; Soliman, KFA

2012-01-01

Age-related increase in monoamine oxidase B (MAO-B) may contribute to CNS neurodegenerative diseases. Moreover, MAO-B inhibitors are used in the treatment of idiopathic Parkinson disease as preliminary monotherapy or adjunct therapy with L-dopa. To date, meager natural sources of MAO-B inhibitors have been identified, and the relative strength, potency and rank of many plants relative to standard drugs such as Selegiline (L-deprenyl, Eldepryl) are not known. In this work, we developed and utilized a high throughput enzyme microarray format to screen and evaluate 905 natural product extracts (0.025–.7 mg/ml) to inhibit human MAO-B derived from BTI-TN-5B1-4 cells infected with recombinant baculovirus. The protein sequence of purified enzyme was confirmed using 1D gel electrophoresis-matrix assisted laser desorption ionization-time-of-flight-tandem mass spectroscopy, and enzyme activity was confirmed by [1] substrate conversion (3-mM benzylamine) to H202 and [2] benzaldehyde. Of the 905 natural extracts tested, the lowest IC50s [Comfrey, Bringraj, Skullcap, Kava-kava, Wild Indigo, Gentian and Green Tea. In conclusion, the data reflect relative potency information by rank of commonly used herbs and plants that contain human MAO-B inhibitory properties in their natural form. PMID:22887993

Plasma diamine oxidase levels in pregnancy complicated by threatened abortion.

OpenAIRE

Legge, M; Duff, G B

1981-01-01

Plasma diamine oxidase levels were assayed in 66 patients who presented with pregnancy complicated by threatened abortion. Levels within the normal range were associated with continuing pregnancies, whereas levels below the normal range were associated with subsequent abortion. Among those patients in whom gestation was greater than eight weeks, 66.6% of diamine oxidase levels correctly predicted the pregnancy outcome. Assay of the diamine oxidase levels at eight weeks of gestation or less ga...

Metavanadate causes cellular accumulation of copper and decreased lysyl oxidase activity

International Nuclear Information System (INIS)

Cui, Changtai T.; Uriu-Adams, Janet Y.; Tchaparian, Eskouhie H.; Keen, Carl L.; Rucker, Robert B.

2004-01-01

Selected indices of copper metabolism in weanling rats and fibroblast cultures were progressively altered in response to increased levels of sodium metavanadate. In diets, vanadium was added in amounts ranging from 0 to 80 μg V/g of diet, that is, 0-1.6 μmol V/g of diet. In fibroblast cultures, vanadium ranged from 0 to 400 nmol V/ml. The inhibition of P-ATPase-7A activity by metavanadate, important to copper egress from cells, was a primary focus. In skin, and tendon, the copper concentration was increased in response to increased dietary levels of metavanadate, whereas lysyl oxidase activity, a secreted cuproprotein, was reduced. The reduction in lysyl oxidase activity was also accompanied by reduced redox cycling potential of isolated fractions of lysyl oxidase, presumably due to reduced lysyltyrosyl quinone (LTQ) formation at the active site of lysyl oxidase. In contrast, liver copper concentrations and plasma ceruloplasmin activity were not affected by metavanadate exposure. However, semicarbazide-sensitive benzylamine oxidase (SCBO) activity, which was taken as an indirect measure of vascular adhesive protein-1 (VAP-1), was increased. In cultured fibroblasts, cellular copper was also increased and lysyl oxidase decreased in response to metavanadate. Moreover, the steady-state levels of atp7a and lysyl oxidase mRNAs were not affected by addition of metavanadate to culture medium up to 200 nmol/ml. Taken together, these data suggest that pathways involving copper egress and lysyl oxidase activation are particularly sensitive to metavanadate exposure through processes that are predominately posttranslational

Molecular Modeling of Peroxidase and Polyphenol Oxidase: Substrate Specificity and Active Site Comparison

Directory of Open Access Journals (Sweden)

Lalida Shank

2010-09-01

Full Text Available Peroxidases (POD and polyphenol oxidase (PPO are enzymes that are well known to be involved in the enzymatic browning reaction of fruits and vegetables with different catalytic mechanisms. Both enzymes have some common substrates, but each also has its specific substrates. In our computational study, the amino acid sequence of grape peroxidase (ABX was used for the construction of models employing homology modeling method based on the X-ray structure of cytosolic ascorbate peroxidase from pea (PDB ID:1APX, whereas the model of grape polyphenol oxidase was obtained directly from the available X-ray structure (PDB ID:2P3X. Molecular docking of common substrates of these two enzymes was subsequently studied. It was found that epicatechin and catechin exhibited high affinity with both enzymes, even though POD and PPO have different binding pockets regarding the size and the key amino acids involved in binding. Predicted binding modes of substrates with both enzymes were also compared. The calculated docking interaction energy of trihydroxybenzoic acid related compounds shows high affinity, suggesting specificity and potential use as common inhibitor to grape ascorbate peroxidase and polyphenol oxidase.

Oxidases as Breast Cancer Oncogens

National Research Council Canada - National Science Library

Yeldandi, Anjana

2000-01-01

...) in a non-tumorigenic human mammary epithelial cell line to ascertain whether oxidase overexpressing cells undergo transformation when exposed to substrate xanthine for XOX and uric acid for UOX...

Plasma diamine oxidase levels in pregnancy complicated by threatened abortion.

Science.gov (United States)

Legge, M; Duff, G B

1981-02-01

Plasma diamine oxidase levels were assayed in 66 patients who presented with pregnancy complicated by threatened abortion. Levels within the normal range were associated with continuing pregnancies, whereas levels below the normal range were associated with subsequent abortion. Among those patients in whom gestation was greater than eight weeks, 66.6% of diamine oxidase levels correctly predicted the pregnancy outcome. Assay of the diamine oxidase levels at eight weeks of gestation or less gave little useful information.

Biocompatibility selenium nanoparticles with an intrinsic oxidase-like activity

International Nuclear Information System (INIS)

Guo, Leilei; Huang, Kaixun; Liu, Hongmei

2016-01-01

Selenium nanoparticles (SeNPs) are considered to be the new selenium supplement forms with high biological activity and low toxicity; however, the molecular mechanism by which SeNPs exert the biological function is unclear. Here, we reported that biocompatibility SeNPs possessed intrinsic oxidase-like activity. Using Na 2 SeO 3 as a precursor and glutathione as a reductant, biocompatibility SeNPs were synthesized by the wet chemical reduction method in the presence of bovine serum albumin (BSA). The results of structure characterization revealed that synthesized SeNPs were amorphous red elementary selenium with spherical morphology, and ranged in size from 25 to 70 nm size with a narrow distribution (41.4 ± 6.7 nm). The oxidase-like activity of the as-synthesized SeNPs was tested with 3,3′,5,5′-tetramethylbenzidine (TMB) as a substrate. The results indicated that SeNPs could catalyze the oxidization of TMB by dissolved oxygen. These SeNPs showed an optimum catalytic activity at pH 4 and 30 °C, and the oxidase-like activity was higher as the concentration of SeNPs increased and the size of SeNPs decreased. The Michaelis constant (K m ) values and maximal reaction velocity (V max ) of the SeNPs for TMB oxidation were 0.0083 mol/L and 3.042 μmol/L min, respectively.

Biocompatibility selenium nanoparticles with an intrinsic oxidase-like activity

Science.gov (United States)

Guo, Leilei; Huang, Kaixun; Liu, Hongmei

2016-03-01

Selenium nanoparticles (SeNPs) are considered to be the new selenium supplement forms with high biological activity and low toxicity; however, the molecular mechanism by which SeNPs exert the biological function is unclear. Here, we reported that biocompatibility SeNPs possessed intrinsic oxidase-like activity. Using Na2SeO3 as a precursor and glutathione as a reductant, biocompatibility SeNPs were synthesized by the wet chemical reduction method in the presence of bovine serum albumin (BSA). The results of structure characterization revealed that synthesized SeNPs were amorphous red elementary selenium with spherical morphology, and ranged in size from 25 to 70 nm size with a narrow distribution (41.4 ± 6.7 nm). The oxidase-like activity of the as-synthesized SeNPs was tested with 3,3',5,5'-tetramethylbenzidine (TMB) as a substrate. The results indicated that SeNPs could catalyze the oxidization of TMB by dissolved oxygen. These SeNPs showed an optimum catalytic activity at pH 4 and 30 °C, and the oxidase-like activity was higher as the concentration of SeNPs increased and the size of SeNPs decreased. The Michaelis constant ( K m) values and maximal reaction velocity ( V max) of the SeNPs for TMB oxidation were 0.0083 mol/L and 3.042 μmol/L min, respectively.

Effect of gamma irradiation on aspergillus niger for enhanced production of glucose oxidase

International Nuclear Information System (INIS)

Zia, M.A.; Rasul, S.

2012-01-01

Developing countries have a high prevalence of diabetes and their populations are suffering from associated adverse factors. Such a frequency requires more effective diagnosis, mostly achieved by glucose diagnostic kits. Although high priced kits are available in market but local production of such kits can be highly cost effective and may confer the decline in incidence of the disease. Glucose oxidase is the key enzyme for the determination of glucose in such analytical tools. Enhanced production of glucose oxidase was performed by mutagenesis of Aspergillus niger by gamma irradiation. A dose of 80 krad was found as optimum for derivation of positive mutant strains. Following the screening by triton X-100 and 2-deoxy-D-glucose, the selected strains A. niger G-80-A, A. niger G-80-B and A. niger G-80-C showed 27.5, 23.20 and 20.55 UmL/sub -1/ glucose oxidase activity in enzyme diffusion zone test; which is much higher to parental strain (7.5 UmL/sup -1/). A. niger G-80-A was subjected to submerged fermentation and obtained highest yields after 36 h, at CSL 2%, pH 6.5, 30 degree C, KH/sub 2/PO/sub 4/ 0.8% and urea 0.3%. Partial purification by ammonium sulfate resulted in 175 UmL/sup -1/ of glucose oxidase activity after dialysis. Kinetic parameters like optimum pH, temperature, K/sub m/ and V/sub max/ were found to be 6.0 (180 +- 2 UmL/sup -1/), 30 degree C (185 +- 0.5 UmL/sup -1/), 5.26 mM and 400 U mL/sup -1/, respectively. Active inhibition of the enzyme by increasing concentration of PLP in reaction mixture confirmed the presence of functional lysyl residue on the active site of enzyme. (author)

21 CFR 866.2420 - Oxidase screening test for gonorrhea.

Science.gov (United States)

2010-04-01

... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Oxidase screening test for gonorrhea. 866.2420... screening test for gonorrhea. (a) Identification. An oxidase screening test for gonorrhea is an in vitro... of gonorrhea. (b) Classification. Class III (premarket approval) (transitional device). (c) Date PMA...

Association of molecular markers with polyphenol oxidase activity in selected wheat genotypes

International Nuclear Information System (INIS)

Abbas, Z.; Javad, B.; Majeed, N.; Naqvi, S.

2016-01-01

Wheat (Triticum aestivum L.), a major staple food for the people of Pakistan and other Asian countries, is used as bread, chapatti, porridge, noodles and many other. It is established that color quality of wheat products depend on chemical and enzymatic factors especially the polyphenol oxidases (PPOs). These are copper containing enzymes which induce browning in wheat-based products. Various procedures for determining PPO activity available and differences in PPO activity among wheat genotypes have been documented. In present study, an attempt was made to establish the association of molecular markers with polyphenol oxidase activity in wheat genotypes having very high or very low PPO activities. Twelve pairs of markers were used out of which only three primer pairs viz. PPO43, PPO30 and WP2-2 yielded specific pattern discriminating high and low PPO genotypes. Cluster analysis for all 12 markers revealed that all the low PPO lline share the same sub cluster, but high PPO lines were dispersed in different clusters. (author)

Effect of cuprous halide interlayers on the device performance of ZnPc/C60 organic solar cells

International Nuclear Information System (INIS)

Lee, Jinho; Park, Dasom; Heo, Ilsu; Yim, Sanggyu

2014-01-01

Highlights: • Effect of CuX interlayers on subsequently deposited films and devices was studied. • CuI is the most effective for the performance of ZnPc/C 60 -based solar cells. • Results were related to the molecular geometry of ZnPc and HOMO level of interlayers. - Abstract: The effect of various cuprous halide (CuX) interlayers introduced between a poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) layer and zinc phthalocyanine (ZnPc) layer on the physical properties of the ZnPc thin films and device performances of ZnPc/C 60 -based small-molecule organic solar cells was studied. Strong substrate–molecule interaction between the CuX and ZnPc partly converted surface-perpendicular stacking geometry of ZnPc molecules into surface-parallel one. This flat-lying geometry led to an enhancement in electronic absorption and charge transport within the ZnPc films. As a result, the overall power conversion efficiency of the cell with CuI interlayer increased by ∼37%. In the case of the cells with CuBr and CuCl interlayer, however, the enhancement in device performances was limited because of the reduced conversion of the molecular geometry and increased energy barrier for hole extraction due to the low highest occupied molecular orbital level of the interlayer

Dynamic 1-aminocyclopropane-1-carboxylate-synthase and -oxidase transcript accumulation patterns during pollen tube growth in tobacco styles.

Science.gov (United States)

Weterings, Koen; Pezzotti, Mario; Cornelissen, Marc; Mariani, Celestina

2002-11-01

In flowering plants, pollination of the stigma sets off a cascade of responses in the distal flower organs. Ethylene and its biosynthetic precursor 1-aminocyclopropane-1-carboxylate (ACC) play an important role in regulating these responses. Because exogenous application of ethylene or ACC does not invoke the full postpollination syndrome, the pollination signal probably consists of a more complex set of stimuli. We set out to study how and when the pollination signal moves through the style of tobacco (Nicotiana tabacum) by analyzing the expression patterns of pistil-expressed ACC-synthase and -oxidase genes. Results from this analysis showed that pollination induces high ACC-oxidase transcript levels in all cells of the transmitting tissue. ACC-synthase mRNA accumulated only in a subset of transmitting tract cells and to lower levels as compared with ACC-oxidase. More significantly, we found that although ACC-oxidase transcripts accumulate to uniform high levels, the ACC-synthase transcripts accumulate in a wave-like pattern in which the peak coincides with the front of the ingrowing pollen tube tips. This wave of ACC-synthase expression can also be induced by incongruous pollination and (partially) by wounding. This indicates that wounding-like features of pollen tube invasion might be part of the stimuli evoking the postpollination response and that these stimuli are interpreted differently by the regulatory mechanisms of the ACC-synthase and -oxidase genes.

Catalytic aspects of a copper(II) complex: biological oxidase to ...

Indian Academy of Sciences (India)

BISWAJIT CHOWDHURY

2017-10-03

Oct 3, 2017 ... made with a Jasco model V-730 UV-Vis spectrophotometer. ..... Ligand-induced coordination changes ... Fet3 protein from yeast, a multinuclear copper oxidase ... of mutants of the multicopper oxidase Fet3p Biochem-.

Controlling the Optical and Magnetic Properties of Nanostructured Cuprous Oxide Synthesized from Waste Electric Cables

Science.gov (United States)

Abdelbasir, S. M.; El-Sheikh, S. M.; Rashad, M. M.; Rayan, D. A.

2018-03-01

Cuprous oxide Cu2O nanopowders were purposefully synthesised from waste electric cables (WECs) via a simple precipitation route at room temperature using lactose as a reducing agent. In this regard, dimethyl sulfoxide (DMSO) was first applied as an organic solvent for the dissolution of the cable insulating materials. Several parameters were investigated during dissolution of WECs such as dissolution temperature, time and solid/liquid ratio to determine the dissolution percentage of the insulating materials in DMSO. The morphology and the optical properties of the formed Cu2O particles were investigated using X-ray diffraction (XRD), field emission-scanning electron microscopy (FE-SEM), Fourier-transform infrared spectroscopy and UV-visible-near IR spectrophotometer. XRD data confirmed the presence of single crystalline phase of Cu2O nanoparticles. FE-SEM and TEM images revealed spherical, cubic and octahedral shapes with the various particle sizes ranged from 16 to 57 nm depending on the synthesis conditions. A possible mechanism explaining the Cu2O nanostructures formation was proposed. The band gap energies of the Cu2O nanostructures were estimated and the values were located between 1.5 and 2.08 eV. Photoluminescence spectroscopy analysis clearly showed a noticeably blue-shifted emission for the synthesized samples compared to spectrum of the bulk. Eventually, magnetic properties of the synthesized nanoparticles have been measured by vibrating sample magnetometer and the attained results implied that the synthesized particles are weakly ferromagnetic in nature at normal temperature.

The effect of heat on DNA degradation by the 1, 10-phenanthroline-cuprous ion complex

International Nuclear Information System (INIS)

Nagle, W.A.; Henle, K.J.; Willingham, W.M.; Sorenson, J.R.J.; McClellan, J.L.; Moss, A.J.

1987-01-01

The 1, 10-phenanthroline-cuprous ion complex (OP)/sub 2/Cu/sup +/ exhibits artificial DNase activity which closely parallels micrococcal nuclease. Using cell-free systems and in situ generated (OP)/sub 2/Cu/sup +/, other studies have shown a requirement for a reducing agent as well as O/sub 2/ or H/sub 2/O/sub 2/ to degrade DNA to acid-soluble fragments. The authors investigated the influence of hyperthermia on the degradation of V79 cell DNA using the (OP)/sub 2/Cu/sup +/-ascorbate system. The (OP)/sub 2/Cu/sup +/ complex was synthesized and characterized prior to cell treatment. Cells were prelabeled with /sup 3/H-TdR (control) or /sup 14/C-TdR (treated) and exposed 10 minutes at 45 0 C, followed by a 30 minute incubation with lμM (OP)/sub 2/Cu/sup +/ and 10μM as corbate in balanced salts solution. Cellular DNA was assayed using the alkaline elution technique. Heated cells incubated with lμM (OP)/sub 2/Cu/sup +/ or 10μM ascorbate exhibited a 300 rad equivalent increase in strand breaks over the unheated control. Incubation of cells with either lμM (OP)/sub 2/Cu/sup +/ or 10μM ascorbate alone did not induce strand breaks. These results suggests that heating initially increases the susceptibility of DNA to attack by the (OP)/sub 2/Cu/sup +/-ascorbate system

Immobilization of xanthine oxidase on a polyaniline silicone support.

Science.gov (United States)

Nadruz, W; Marques, E T; Azevedo, W M; Lima-Filho, J L; Carvalho, L B

1996-03-01

A polyaniline silicone support to immobilize xanthine oxidase is proposed as a reactor coil to monitor the action of xanthine oxidase on hypoxanthine, xanthine and 6-mercaptopurine. A purified xanthine oxidase immobilized on this support lost 80% of the initial activity after 12 min of use. Co-immobilization of superoxide dismutase and catalase increased the stability of immobilized xanthine oxidase so that the derivative maintained 79% of its initial activity after 4.6 h of continuous use in which 1.5 mumol purine bases were converted by the immobilized enzyme system. There is no evidence of either polyaniline or protein leaching from the coil during 3 h of continuous use. When solutions (10 ml) of hypoxanthine, xanthine and 6-mercaptopurine were circulated individually through the xanthine oxidase-superoxide dismutase-catalase-polyaniline coil (1 mm internal diameter and 3 m in length, 3 ml internal volume) activities of 8.12, 11.17 and 1.09 nmol min-1 coil-1, respectively, were obtained. The advantages of the reactor configuration and the redox properties of the polymer, particularly with respect to immobilized oxidoreductases, make this methodology attractive for similar enzyme systems. This immobilized enzyme system using polyaniline-silicone as support converted 6-mercaptopurine to 6-thiouric acid with equal efficiency as resins based on polyacrylamide and polyamide 11.

Identification and characterization of aldehyde oxidases (AOXs) in the cotton bollworm

Science.gov (United States)

Xu, Wei; Liao, Yalin

2017-12-01

Aldehyde oxidases (AOXs) are a family of metabolic enzymes that oxidize aldehydes into carboxylic acids; therefore, they play critical roles in detoxification and degradation of chemicals. By using transcriptomic and genomic approaches, we successfully identified six putative AOX genes (HarmAOX1-6) from cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae). In silico expression profile, reverse transcription (RT)-PCR, and quantitative PCR (qPCR) analyses showed that HarmAOX1 is highly expressed in adult antennae, tarsi, and larval mouthparts, so they may play an important role in degrading plant-derived compounds. HarmAOX2 is highly and specifically expressed in adult antennae, suggesting a candidate pheromone-degrading enzyme (PDE) to inactivate the sex pheromone components (Z)-11-hexadecenal and (Z)-9-hexadecenal. RNA sequencing data further demonstrated that a number of host plants they feed on could significantly upregulate the expression levels of HarmAOX1 in larvae. This study improves our understanding of insect aldehyde oxidases and insect-plant interactions.

Interference of aldehyde metabolizing enzyme with diamine oxidase/histaminase/activity as determined by 14C putrescine method

International Nuclear Information System (INIS)

Fogel, W.A.; Bieganski, T.; Wozniak, J.; Maslinski, C.

1978-01-01

The Δ 1 pyrroline formation, as an indicator of diamine oxidase activity according to Okuyama and Kobayashi 14 C putrescine test (1961, Archs Biochem. Biophys., vol.95, 242), has been investigated in several tissue homogenates. When guinea pig liver homogenate was used as a source of enzyme in the presence of aldehyde dehydrogenase inhibitors chlorate hydrate and acetaldehyde the level of formation Δ 1 pyrroline was strongly increased in a dose-dependent manner. Also inhibition of aldehyde reductase by phenobarbital enhanced Δ 1 pyrroline formation, but to a lesser degree. In other tissues, with very high initial diamine oxidase activity (rat intestine, dog kidney) or with very low diamine oxidase activity (guinea pig skin, dog liver) the influence of these inhibitors was only slight. Pyrazole, an inhibitor of alcohol dehydrogenase exerted only a small effect on Δ 1 pyrroline formation. All aldehyde-metabolizing enzymes inhibitors, except pyrazole, were without effect on purified pea seddling and hog kidney diamine oxidases. The use of aldehyde-metabolizing enzymes inhibitors may help to reveal the real values of diamine oxidase activity, when tissues homogenates are used as a source of enzyme. (author)

Xanthine oxidase in human skeletal muscle following eccentric exercise

DEFF Research Database (Denmark)

Hellsten, Ylva; Frandsen, Ulrik; Orthenblad, N.

1997-01-01

the increase in xanthine oxidase in the muscle there were no detectable changes in the levels of muscle malondialdehyde or in plasma antioxidant capacity up to 4 days post-exercise. 5. It is concluded that eccentric exercise leads to an increased level of xanthine oxidase in human muscle and that the increase...

Construction of mutant glucose oxidases with increased dye-mediated dehydrogenase activity.

Science.gov (United States)

Horaguchi, Yohei; Saito, Shoko; Kojima, Katsuhiro; Tsugawa, Wakako; Ferri, Stefano; Sode, Koji

2012-11-02

Mutagenesis studies on glucose oxidases (GOxs) were conducted to construct GOxs with reduced oxidase activity and increased dehydrogenase activity. We focused on two representative GOxs, of which crystal structures have already been reported—Penicillium amagasakiense GOx (PDB ID; 1gpe) and Aspergillus niger GOx (PDB ID; 1cf3). We constructed oxygen-interacting structural models for GOxs, and predicted the residues responsible for oxidative half reaction with oxygen on the basis of the crystal structure of cholesterol oxidase as well as on the fact that both enzymes are members of the glucose/methanol/choline (GMC) oxidoreductase family. Rational amino acid substitution resulted in the construction of an engineered GOx with drastically decreased oxidase activity and increased dehydrogenase activity, which was higher than that of the wild-type enzyme. As a result, the dehydrogenase/oxidase ratio of the engineered enzyme was more than 11-fold greater than that of the wild-type enzyme. These results indicate that alteration of the dehydrogenase/oxidase activity ratio of GOxs is possible by introducing a mutation into the putative functional residues responsible for oxidative half reaction with oxygen of these enzymes, resulting in a further increased dehydrogenase activity. This is the first study reporting the alteration of GOx electron acceptor preference from oxygen to an artificial electron acceptor.

Construction of Mutant Glucose Oxidases with Increased Dye-Mediated Dehydrogenase Activity

Science.gov (United States)

Horaguchi, Yohei; Saito, Shoko; Kojima, Katsuhiro; Tsugawa, Wakako; Ferri, Stefano; Sode, Koji

2012-01-01

Mutagenesis studies on glucose oxidases (GOxs) were conducted to construct GOxs with reduced oxidase activity and increased dehydrogenase activity. We focused on two representative GOxs, of which crystal structures have already been reported—Penicillium amagasakiense GOx (PDB ID; 1gpe) and Aspergillus niger GOx (PDB ID; 1cf3). We constructed oxygen-interacting structural models for GOxs, and predicted the residues responsible for oxidative half reaction with oxygen on the basis of the crystal structure of cholesterol oxidase as well as on the fact that both enzymes are members of the glucose/methanol/choline (GMC) oxidoreductase family. Rational amino acid substitution resulted in the construction of an engineered GOx with drastically decreased oxidase activity and increased dehydrogenase activity, which was higher than that of the wild-type enzyme. As a result, the dehydrogenase/oxidase ratio of the engineered enzyme was more than 11-fold greater than that of the wild-type enzyme. These results indicate that alteration of the dehydrogenase/oxidase activity ratio of GOxs is possible by introducing a mutation into the putative functional residues responsible for oxidative half reaction with oxygen of these enzymes, resulting in a further increased dehydrogenase activity. This is the first study reporting the alteration of GOx electron acceptor preference from oxygen to an artificial electron acceptor. PMID:23203056

Biocatalytic potential of laccase-like multicopper oxidases from Aspergillus niger

NARCIS (Netherlands)

Tamayo Ramos, J.A.; Berkel, van W.J.H.; Graaff, de L.H.

2012-01-01

BACKGROUND: Laccase-like multicopper oxidases have been reported in several Aspergillus species but they remain uncharacterized. The biocatalytic potential of the Aspergillus niger fungal pigment multicopper oxidases McoA and McoB and ascomycete laccase McoG was investigated. RESULTS: The

Biocompatibility selenium nanoparticles with an intrinsic oxidase-like activity

Energy Technology Data Exchange (ETDEWEB)

Guo, Leilei; Huang, Kaixun; Liu, Hongmei, E-mail: hmliu2004@126.com [Huazhong University of Science and Technology, School of Chemistry and Chemical Engineering (China)

2016-03-15

Selenium nanoparticles (SeNPs) are considered to be the new selenium supplement forms with high biological activity and low toxicity; however, the molecular mechanism by which SeNPs exert the biological function is unclear. Here, we reported that biocompatibility SeNPs possessed intrinsic oxidase-like activity. Using Na{sub 2}SeO{sub 3} as a precursor and glutathione as a reductant, biocompatibility SeNPs were synthesized by the wet chemical reduction method in the presence of bovine serum albumin (BSA). The results of structure characterization revealed that synthesized SeNPs were amorphous red elementary selenium with spherical morphology, and ranged in size from 25 to 70 nm size with a narrow distribution (41.4 ± 6.7 nm). The oxidase-like activity of the as-synthesized SeNPs was tested with 3,3′,5,5′-tetramethylbenzidine (TMB) as a substrate. The results indicated that SeNPs could catalyze the oxidization of TMB by dissolved oxygen. These SeNPs showed an optimum catalytic activity at pH 4 and 30 °C, and the oxidase-like activity was higher as the concentration of SeNPs increased and the size of SeNPs decreased. The Michaelis constant (K{sub m}) values and maximal reaction velocity (V{sub max}) of the SeNPs for TMB oxidation were 0.0083 mol/L and 3.042 μmol/L min, respectively.

Cytochemical Localization of Glucose Oxidase in Peroxisomes of Aspergillus niger

NARCIS (Netherlands)

Veenhuis, Marten; Dijken, Johannes Pieter van

1980-01-01

The subcellular localization of glucose oxidase (E.C. 1.1.3.4) in mycelia of Aspergillus niger has been investigated using cytochemical staining techniques. Mycelia from fermenter cultures, which produced gluconic acid from glucose, contained elevated levels of glucose oxidase and catalase. Both

Reduced graphene oxide–cuprous oxide composite via facial deposition for photocatalytic dye-degradation

Energy Technology Data Exchange (ETDEWEB)

Wang, MingYan, E-mail: mingyanlyg@hotmail.com [Department of Chemical Engineering, Huaihai Institute of Technology, Lianyungang 222005 (China); Intelligent Polymer Research Institute, ARC Centre of Excellence for Electromaterials Science, Australian Institute of Innovative Materials, University of Wollongong, Northfields Avenue, Wollongong, NSW 2522 (Australia); Huang, JunRao; Tong, ZhiWei [Department of Chemical Engineering, Huaihai Institute of Technology, Lianyungang 222005 (China); Li, WeiHua [School of Mechanical, Materials and Mechatronic Engineering, University of Wollongong, Northfields Avenue, Wollongong, NSW 2522 (Australia); Chen, Jun, E-mail: junc@uow.edu.au [Intelligent Polymer Research Institute, ARC Centre of Excellence for Electromaterials Science, Australian Institute of Innovative Materials, University of Wollongong, Northfields Avenue, Wollongong, NSW 2522 (Australia)

2013-08-15

Highlights: •Cubic Cu{sub 2}O were effectively loaded on n-propylamine (PA) intercalated graphene oxide. •The addition of PA on the carbon sheets supports the stable structure of the composites. •Cu{sub 2}O/PA/rGO showed superior adsorption capacity and photocatalytic activity. -- Abstract: Cubic Cu{sub 2}O nanoparticles have been successfully synthesized on n-propylamine (PA) intercalated graphene oxide (GO) with uniform distribution followed with a subsequent hydrazine hydrate reduction process to generate Cu{sub 2}O/PA/rGO composite. For comparison, Cu{sub 2}O conjugated reduced graphene oxide (Cu{sub 2}O/rGO) composite was also synthesized using the same method. The as-prepared Cu{sub 2}O/PA/rGO and Cu{sub 2}O/rGO nanocomposites are characterized by X-ray photoelectron spectroscopy (XPS), X-ray diffraction (XRD) spectroscopy, infrared spectroscopy (IR), scanning electron microscopy (SEM), transmission electron microscopy (TEM), Brunauer–Emmett–Teller (BET) surface area analysis, and Electrochemical impedance spectra (EIS) measurements. UV/vis diffuse reflectance spectroscopy was employed to estimate band gap energies of cuprous oxide composites. The results show that the intercalation of PA into the layered GO increases the surface area of the composites and provides an efficient strategy to load Cu{sub 2}O due to the large and uniform distribution of active sites for anchoring copper ions. The surface area of the Cu{sub 2}O/PA/rGO (123 m{sup 2}/g) nanocomposite was found to be almost 2.5 times higher than that of Cu{sub 2}O/rGO (55.7 m{sup 2}/g). The as-prepared Cu{sub 2}O/PA/rGO show significant improvement on both adsorption capacity and photocatalytic activity towards organic pigment pollution compared with Cu{sub 2}O/rGO under identical performance conditions.

Construction of Mutant Glucose Oxidases with Increased Dye-Mediated Dehydrogenase Activity

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Koji Sode

2012-11-01

Full Text Available Mutagenesis studies on glucose oxidases (GOxs were conducted to construct GOxs with reduced oxidase activity and increased dehydrogenase activity. We focused on two representative GOxs, of which crystal structures have already been reported—Penicillium amagasakiense GOx (PDB ID; 1gpe and Aspergillus niger GOx (PDB ID; 1cf3. We constructed oxygen-interacting structural models for GOxs, and predicted the residues responsible for oxidative half reaction with oxygen on the basis of the crystal structure of cholesterol oxidase as well as on the fact that both enzymes are members of the glucose/methanol/choline (GMC oxidoreductase family. Rational amino acid substitution resulted in the construction of an engineered GOx with drastically decreased oxidase activity and increased dehydrogenase activity, which was higher than that of the wild-type enzyme. As a result, the dehydrogenase/oxidase ratio of the engineered enzyme was more than 11-fold greater than that of the wild-type enzyme. These results indicate that alteration of the dehydrogenase/oxidase activity ratio of GOxs is possible by introducing a mutation into the putative functional residues responsible for oxidative half reaction with oxygen of these enzymes, resulting in a further increased dehydrogenase activity. This is the first study reporting the alteration of GOx electron acceptor preference from oxygen to an artificial electron acceptor.

Protein structural development of threadfin bream ( Nemipterus spp.) surimi gels induced by glucose oxidase.

Science.gov (United States)

Wang, Lei; Fan, Daming; Fu, Lulu; Jiao, Xidong; Huang, Jianlian; Zhao, Jianxin; Yan, Bowen; Zhou, Wenguo; Zhang, Wenhai; Ye, Weijian; Zhang, Hao

2018-01-01

This study investigated the effect of glucose oxidase on the gel properties of threadfin bream surimi. The gel strength of surimi increased with the addition of 0.5‰ glucose oxidase after two-step heating. Based on the results of the chemical interactions, the hydrophobic interaction and disulfide bond of glucose oxidase-treated surimi samples increased compared with the control samples at the gelation temperature and gel modori temperature. The surface hydrophobicity of samples with glucose oxidase and glucose increased significantly ( p glucose oxidase induced more α-helixes to turn into a more elongated random and flocculent structure. Glucose oxidase changes the secondary structure of the surimi protein, making more proteins depolarize and stretch and causing actomyosin to accumulate to each other, resulting in the formation of surimi gel.

Functional expression of amine oxidase from Aspergillus niger (AO-I) in Saccharomyces cerevisiae.

Science.gov (United States)

Kolaríková, Katerina; Galuszka, Petr; Sedlárová, Iva; Sebela, Marek; Frébort, Ivo

2009-01-01

The aim of this work was to prepare recombinant amine oxidase from Aspergillus niger after overexpressing in yeast. The yeast expression vector pDR197 that includes a constitutive PMA1 promoter was used for the expression in Saccharomyces cerevisiae. Recombinant amine oxidase was extracted from the growth medium of the yeast, purified to homogeneity and identified by activity assay and MALDI-TOF peptide mass fingerprinting. Similarity search in the newly published A. niger genome identified six genes coding for copper amine oxidase, two of them corresponding to the previously described enzymes AO-I a methylamine oxidase and three other genes coding for FAD amine oxidases. Thus, A. niger possesses an enormous metabolic gear to grow on amine compounds and thus support its saprophytic lifestyle.

The immobilisation of trypsin and glucose oxidase onto natural rubber-g.co-HEMA - high energy radiation derived copolymeric support systems

International Nuclear Information System (INIS)

Devi, S.; Guthrie, J.T.; Beddows, C.G.

1990-01-01

Natural rubber has been grafted with 2-HEMA by three different methods each involving Co(60)γ-radiation as the initiation source. The grafted samples were used in the immobilisation of glucose oxidase and trypsin. Optimisation of immobilisation involving trypsin was studied with regard to the pH and the type of crosslinking agent. It was observed that the immobilised enzyme had superior stability over a wider pH range when compared to the free trypsin. The retention of activity demonstrated by the immobilised trypsin was significant. That of immobilised glucose oxidase was far from being satisfactory. (author)

Biofilm Inhibition by Novel Natural Product- and Biocide-Containing Coatings Using High-Throughput Screening

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Maria Salta

2018-05-01

Full Text Available The use of natural products (NPs as possible alternative biocidal compounds for use in antifouling coatings has been the focus of research over the past decades. Despite the importance of this field, the efficacy of a given NP against biofilm (mainly bacteria and diatoms formation is tested with the NP being in solution, while almost no studies test the effect of an NP once incorporated into a coating system. The development of a novel bioassay to assess the activity of NP-containing and biocide-containing coatings against marine biofilm formation has been achieved using a high-throughput microplate reader and highly sensitive confocal laser scanning microscopy (CLSM, as well as nucleic acid staining. Juglone, an isolated NP that has previously shown efficacy against bacterial attachment, was incorporated into a simple coating matrix. Biofilm formation over 48 h was assessed and compared against coatings containing the NP and the commonly used booster biocide, cuprous oxide. Leaching of the NP from the coating was quantified at two time points, 24 h and 48 h, showing evidence of both juglone and cuprous oxide being released. Results from the microplate reader showed that the NP coatings exhibited antifouling efficacy, significantly inhibiting biofilm formation when compared to the control coatings, while NP coatings and the cuprous oxide coatings performed equally well. CLSM results and COMSTAT analysis on biofilm 3D morphology showed comparable results when the NP coatings were tested against the controls, with higher biofilm biovolume and maximum thickness being found on the controls. This new method proved to be repeatable and insightful and we believe it is applicable in antifouling and other numerous applications where interactions between biofilm formation and surfaces is of interest.

Vanillyl alcohol oxidases produced in Komagataella phaffii contain a highly stable non-covalently bound anionic FAD semiquinone

NARCIS (Netherlands)

Gygli, G.A.; Berkel, van W.J.H.

2017-01-01

Vanillyl alcohol oxidase (VAO) from Penicillium simplicissimum is a covalent flavoprotein that has emerged as a promising biocatalyst for the production of aromatic fine chemicals such as vanillin, coniferyl alcohol and enantiopure 1-(4’-hydroxyphenyl) alcohols. The largescale production of this

Interference of aldehyde metabolizing enzyme with diamine oxidase/histaminase/activity as determined by /sup 14/C putrescine method

Energy Technology Data Exchange (ETDEWEB)

Fogel, W A [Polish Academy of Sciences, Cracow (Poland). Inst. of Pharmacology; Bieganski, T; Wozniak, J; Maslinski, C

1978-01-01

The ..delta../sup 1/ pyrroline formation, as an indicator of diamine oxidase activity according to Okuyama and Kobayashi /sup 14/C putrescine test (1961, Archs Biochem. Biophys., vol.95, 242), has been investigated in several tissue homogenates. When guinea pig liver homogenate was used as a source of enzyme in the presence of aldehyde dehydrogenase inhibitors chlorate hydrate and acetaldehyde the level of formation ..delta../sup 1/ pyrroline was strongly increased in a dose-dependent manner. Also inhibition of aldehyde reductase by phenobarbital enhanced ..delta../sup 1/ pyrroline formation, but to a lesser degree. In other tissues, with very high initial diamine oxidase activity (rat intestine, dog kidney) or with very low diamine oxidase activity (guinea pig skin, dog liver) the influence of these inhibitors was only slight. Pyrazole, an inhibitor of alcohol dehydrogenase exerted only a small effect on ..delta../sup 1/ pyrroline formation. All aldehyde-metabolizing enzymes inhibitors, except pyrazole, were without effect on purified pea seddling and hog kidney diamine oxidases. The use of aldehyde-metabolizing enzymes inhibitors may help to reveal the real values of diamine oxidase activity, when tissues homogenates are used as a source of enzyme.

The use of galactose oxidase in lipid labeling

International Nuclear Information System (INIS)

Radin, N.S.; Evangelatos, G.P.

1981-01-01

Galactose oxidase can be used to oxidize the terminal carbon atom of lipids containing galactose or N-acetylgalactosamine, and the resultant aldehyde group can be reduced back to the original carbinol with radioactive borohydride. The efficiency of the first reaction has been investigated systematically by using [6- 3 H]galactosyl ceramide as substrate and measuring the amount of radioactive water formed. This enabled us to establish that the addition of catalase and peroxidase greatly speeded the oxidation, that phosphate and PIPES buffers were the best among those tested, that the reaction continued for 24 hr without a second addition of galactose oxidase, and that the optimum concentration of organic solvent (tetrahydrofuran) was 50%. The suggestion if made that a similar set of variables be studied for each lipid or nonlipid by the same basic technique: labeling by the oxidase/borohydride method and use of the resultant compound as substrate

Effect of cuprous halide interlayers on the device performance of ZnPc/C{sub 60} organic solar cells

Energy Technology Data Exchange (ETDEWEB)

Lee, Jinho; Park, Dasom; Heo, Ilsu; Yim, Sanggyu, E-mail: sgyim@kookmin.ac.kr

2014-10-15

Highlights: • Effect of CuX interlayers on subsequently deposited films and devices was studied. • CuI is the most effective for the performance of ZnPc/C{sub 60}-based solar cells. • Results were related to the molecular geometry of ZnPc and HOMO level of interlayers. - Abstract: The effect of various cuprous halide (CuX) interlayers introduced between a poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate) layer and zinc phthalocyanine (ZnPc) layer on the physical properties of the ZnPc thin films and device performances of ZnPc/C{sub 60}-based small-molecule organic solar cells was studied. Strong substrate–molecule interaction between the CuX and ZnPc partly converted surface-perpendicular stacking geometry of ZnPc molecules into surface-parallel one. This flat-lying geometry led to an enhancement in electronic absorption and charge transport within the ZnPc films. As a result, the overall power conversion efficiency of the cell with CuI interlayer increased by ∼37%. In the case of the cells with CuBr and CuCl interlayer, however, the enhancement in device performances was limited because of the reduced conversion of the molecular geometry and increased energy barrier for hole extraction due to the low highest occupied molecular orbital level of the interlayer.

Molecular Basis for Converting (2S-Methylsuccinyl-CoA Dehydrogenase into an Oxidase

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Simon Burgener

2017-12-01

Full Text Available Although flavoenzymes have been studied in detail, the molecular basis of their dioxygen reactivity is only partially understood. The members of the flavin adenosine dinucleotide (FAD-dependent acyl-CoA dehydrogenase and acyl-CoA oxidase families catalyze similar reactions and share common structural features. However, both enzyme families feature opposing reaction specificities in respect to dioxygen. Dehydrogenases react with electron transfer flavoproteins as terminal electron acceptors and do not show a considerable reactivity with dioxygen, whereas dioxygen serves as a bona fide substrate for oxidases. We recently engineered (2S-methylsuccinyl-CoA dehydrogenase towards oxidase activity by rational mutagenesis. Here we characterized the (2S-methylsuccinyl-CoA dehydrogenase wild-type, as well as the engineered (2S-methylsuccinyl-CoA oxidase, in detail. Using stopped-flow UV-spectroscopy and liquid chromatography-mass spectrometry (LC-MS based assays, we explain the molecular base for dioxygen reactivity in the engineered oxidase and show that the increased oxidase function of the engineered enzyme comes at a decreased dehydrogenase activity. Our findings add to the common notion that an increased activity for a specific substrate is achieved at the expense of reaction promiscuity and provide guidelines for rational engineering efforts of acyl-CoA dehydrogenases and oxidases.

The increasing role of monoamine oxidase type B inhibitors in Parkinson's disease therapy.

Science.gov (United States)

Elmer, Lawrence W; Bertoni, John M

2008-11-01

The role of monoamine oxidase type B inhibitors in the treatment of Parkinson's disease has expanded with the new monoamine oxidase B inhibitor rasagiline and a new formulation, selegiline oral disintegrating tablets. As primary therapy in early disease monoamine oxidase B inhibitors reduce motor disability and delay the need for levodopa. In more advanced disease requiring levodopa, adjunctive monoamine oxidase B inhibitors reduce 'off' time and may improve gait and freezing. Rasagiline and selegiline oral disintegrating tablets may reduce the safety risks associated with the amfetamine and methamfetamine metabolites of conventional oral selegiline while retaining or improving therapeutic efficacy. Articles were identified by searches of PubMed and searches on the Internet and reviewed. All articles and other referenced materials were retrieved using the keywords 'Parkinson's disease', 'treatment' and 'monoamine oxidase B inhibitor' and were published between 1960 and 2007, with older references selected for historical significance. Only papers published in English were reviewed. Accumulating data support the use of monoamine oxidase B inhibitors as monotherapy for early and mild Parkinson's disease and as adjunctive therapy for more advanced Parkinson's disease with levodopa-associated motor fluctuations. The recently released monoamine oxidase B inhibitor rasagiline and a new formulation, selegiline oral disintegrating tablets, have potential advantages over conventional oral selegiline.

Screening of Bothrops snake venoms for L-amino acid oxidase activity

Energy Technology Data Exchange (ETDEWEB)

Pessati, M.L.; Fontana, J.D.; Guimaraes, M.F. [Federal Univ. of Parana, Curitiba (Brazil)

1995-12-31

Toxins, enzymes, and biologically active peptides are the main components of snake venoms from the genus Bothrops. Following the venom inoculation, the local effects are hemorrhage, edema, and myonecrosis. Nineteen different species of Brazilian Bothrops were screened for protein content and L-amino acid oxidase activity. B. cotiara, formerly found in the South of Brazil, is now threatened with extinction. Its venom contains a highly hemorrhagic fraction and, as expected from the deep yellow color of the corresponding lyophilized powder, a high L-amino acid oxidase (LAO) activity was also characterized. Flavin adenine dinucleotide (FAD) is its associate coenzyme. B. cotiara venom LAO catalyzed the oxidative deamination of several L-amino acids, and the best substrates were methionine, leucine, tryptophan, and phenylalanine, hence, its potential application for the use in biosensors for aspartame determination and for the removal of amino acids from plasma. High levels for LAO were also found in other species than B. cotiara. In addition, the technique of isoelectric focusing (IEF) was employed as a powerful tool to study the iso- or multi-enzyme distribution for LAO activity in the B. cotiara snake venom.

Rac1-NADPH oxidase signaling promotes CD36 activation under glucotoxic conditions in pancreatic beta cells.

Science.gov (United States)

Elumalai, Suma; Karunakaran, Udayakumar; Lee, In Kyu; Moon, Jun Sung; Won, Kyu Chang

2017-04-01

We recently reported that cluster determinant 36 (CD36), a fatty acid transporter, plays a pivotal role in glucotoxicity-induced β-cell dysfunction. However, little is known about how glucotoxicity influences CD36 expression. Emerging evidence suggests that the small GTPase Rac1 is involved in the pathogenesis of beta cell dysfunction in type 2 diabetes (T2D). The primary objective of the current study was to determine the role of Rac1 in CD36 activation and its impact on β-cell dysfunction in diabetes mellitus. To address this question, we subjected INS-1 cells and human beta cells (1.1B4) to high glucose conditions (30mM) in the presence or absence of Rac1 inhibition either by NSC23766 (Rac1 GTPase inhibitor) or small interfering RNA. High glucose exposure in INS-1 and human beta cells (1.1b4) resulted in the activation of Rac1 and induced cell apoptosis. Rac1 activation mediates NADPH oxidase (NOX) activation leading to elevated ROS production in both cells. Activation of the Rac1-NOX complex by high glucose levels enhanced CD36 expression in INS-1 and human 1.1b4 beta cell membrane fractions. The inhibition of Rac1 by NSC23766 inhibited NADPH oxidase activity and ROS generation induced by high glucose concentrations in INS-1 & human 1.1b4 beta cells. Inhibition of Rac1-NOX complex activation by NSC23766 significantly reduced CD36 expression in INS-1 and human 1.1b4 beta cell membrane fractions. In addition, Rac1 inhibition by NSC23766 significantly reduced high glucose-induced mitochondrial dysfunction. Furthermore, NADPH oxidase inhibition by VAS2870 also attenuated high glucose-induced ROS generation and cell apoptosis. These results suggest that Rac1-NADPH oxidase dependent CD36 expression contributes to high glucose-induced beta cell dysfunction and cell death. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Amine oxidase from lentil seedlings: energetic domains and effect of temperature on activity.

Science.gov (United States)

Moosavi-Nejad, S Z; Rezaei-Tavirani, M; Padiglia, A; Floris, G; Moosavi-Movahedi, A A

2001-07-01

Copper/TPQ amine oxidases from mammalian and plant sources have shown many differences in substrate specificity and molecular properties. In this work the activity of lentil seedling amine oxidase was followed at various temperatures in 100 mM potassium phosphate buffer, pH 7, using benzylamine as substrate. The discontinuous Arrhenius plot of lentil amine oxidase showed two distinct phases with a jump between them. Thermal denaturation of the enzyme, using differential scanning calorimetry under the same experimental conditions, showed a transition at the same temperature ranges in the absence of substrate, indicating the occurrence of conformational changes, with an enthalpy change of about 175.9 kJ/mole. The temperature-induced changes of the activity of lentil amine oxidase are compared with those of bovine serum amine oxidase (taken from the literature).

Integration of a highly ordered gold nanowires array with glucose oxidase for ultra-sensitive glucose detection.

Science.gov (United States)

Cui, Jiewu; Adeloju, Samuel B; Wu, Yucheng

2014-01-27

A highly sensitive amperometric nanobiosensor has been developed by integration of glucose oxidase (GO(x)) with a gold nanowires array (AuNWA) by cross-linking with a mixture of glutaraldehyde (GLA) and bovine serum albumin (BSA). An initial investigation of the morphology of the synthesized AuNWA by field emission scanning electron microscopy (FESEM) and field emission transmission electron microscopy (FETEM) revealed that the nanowires array was highly ordered with rough surface, and the electrochemical features of the AuNWA with/without modification were also investigated. The integrated AuNWA-BSA-GLA-GO(x) nanobiosensor with Nafion membrane gave a very high sensitivity of 298.2 μA cm(-2) mM(-1) for amperometric detection of glucose, while also achieving a low detection limit of 0.1 μM, and a wide linear range of 5-6000 μM. Furthermore, the nanobiosensor exhibited excellent anti-interference ability towards uric acid (UA) and ascorbic acid (AA) with the aid of Nafion membrane, and the results obtained for the analysis of human blood serum indicated that the device is capable of glucose detection in real samples. Copyright © 2013 Elsevier B.V. All rights reserved.

Lysyl oxidase in cancer research

DEFF Research Database (Denmark)

Perryman, Lara; Erler, Janine Terra

2014-01-01

Metastasis is the main reason for cancer-associated deaths and therapies are desperately needed to target the progression of cancer. Lysyl oxidase (LOX) plays a pivotal role in cancer progression, including metastasis, and is therefore is an attractive therapeutic target. In this review we...

An Oxidase-Based Electrochemical Fluidic Sensor with High-Sensitivity and Low-Interference by On-Chip Oxygen Manipulation

Directory of Open Access Journals (Sweden)

Chang-Soo Kim

2012-06-01

Full Text Available Utilizing a simple fluidic structure, we demonstrate the improved performance of oxidase-based enzymatic biosensors. Electrolysis of water is utilized to generate bubbles to manipulate the oxygen microenvironment close to the biosensor in a fluidic channel. For the proper enzyme reactions to occur, a simple mechanical procedure of manipulating bubbles was developed to maximize the oxygen level while minimizing the pH change after electrolysis. The sensors show improved sensitivities based on the oxygen dependency of enzyme reaction. In addition, this oxygen-rich operation minimizes the ratio of electrochemical interference signal by ascorbic acid during sensor operation (i.e., amperometric detection of hydrogen peroxide. Although creatinine sensors have been used as the model system in this study, this method is applicable to many other biosensors that can use oxidase enzymes (e.g., glucose, alcohol, phenol, etc. to implement a viable component for in-line fluidic sensor systems.

Angiotensin II inhibits the Na+-K+ pump via PKC-dependent activation of NADPH oxidase.

Science.gov (United States)

White, Caroline N; Figtree, Gemma A; Liu, Chia-Chi; Garcia, Alvaro; Hamilton, Elisha J; Chia, Karin K M; Rasmussen, Helge H

2009-04-01

The sarcolemmal Na(+)-K(+) pump, pivotal in cardiac myocyte function, is inhibited by angiotensin II (ANG II). Since ANG II activates NADPH oxidase, we tested the hypothesis that NADPH oxidase mediates the pump inhibition. Exposure to 100 nmol/l ANG II increased superoxide-sensitive fluorescence of isolated rabbit ventricular myocytes. The increase was abolished by pegylated superoxide dismutase (SOD), by the NADPH oxidase inhibitor apocynin, and by myristolated inhibitory peptide to epsilon-protein kinase C (epsilonPKC), previously implicated in ANG II-induced Na(+)-K(+) pump inhibition. A role for epsilonPKC was also supported by an ANG II-induced increase in coimmunoprecipitation of epsilonPKC with the receptor for the activated kinase and with the cytosolic p47(phox) subunit of NADPH oxidase. ANG II decreased electrogenic Na(+)-K(+) pump current in voltage-clamped myocytes. The decrease was abolished by SOD, by the gp91ds inhibitory peptide that blocks assembly and activation of NADPH oxidase, and by epsilonPKC inhibitory peptide. Since colocalization should facilitate NADPH oxidase-dependent regulation of the Na(+)-K(+) pump, we examined whether there is physical association between the pump subunits and NADPH oxidase. The alpha(1)-subunit coimmunoprecipitated with caveolin 3 and with membrane-associated p22(phox) and cytosolic p47(phox) NADPH oxidase subunits at baseline. ANG II had no effect on alpha(1)/caveolin 3 or alpha(1)/p22(phox) interaction, but it increased alpha(1)/p47(phox) coimmunoprecipitation. We conclude that ANG II inhibits the Na(+)-K(+) pump via PKC-dependent NADPH oxidase activation.

X-ray analysis of bilirubin oxidase from Myrothecium verrucaria at 2.3 Å resolution using a twinned crystal

International Nuclear Information System (INIS)

Mizutani, Kimihiko; Toyoda, Mayuko; Sagara, Kenta; Takahashi, Nobuyuki; Sato, Atsuko; Kamitaka, Yuji; Tsujimura, Seiya; Nakanishi, Yuji; Sugiura, Toshiyuki; Yamaguchi, Shotaro; Kano, Kenji; Mikami, Bunzo

2010-01-01

The crystal structure of bilirubin oxidase (BOD) from M. verrucaria has been determined at 2.3 Å resolution using a merohedrally twinned crystal. BOD has four copper-coordination sites that are almost identical to those of other multicopper oxidases and is also very similar to them in overall structure. Bilirubin oxidase (BOD), a multicopper oxidase found in Myrothecium verrucaria, catalyzes the oxidation of bilirubin to biliverdin. Oxygen is the electron acceptor and is reduced to water. BOD is used for diagnostic analysis of bilirubin in serum and has attracted considerable attention as an enzymatic catalyst for the cathode of biofuel cells that work under neutral conditions. Here, the crystal structure of BOD is reported for the first time. Blue bipyramid-shaped crystals of BOD obtained in 2-methyl-2,4-pentanediol (MPD) and ammonium sulfate solution were merohedrally twinned in space group P6 3 . Structure determination was achieved by the single anomalous diffraction (SAD) method using the anomalous diffraction of Cu atoms and synchrotron radiation and twin refinement was performed in the resolution range 33–2.3 Å. The overall organization of BOD is almost the same as that of other multicopper oxidases: the protein is folded into three domains and a total of four copper-binding sites are found in domains 1 and 3. Although the four copper-binding sites were almost identical to those of other multicopper oxidases, the hydrophilic Asn residue (at the same position as a hydrophobic residue such as Leu in other multicopper oxidases) very close to the type I copper might contribute to the characteristically high redox potential of BOD

Systemic Manifestations in Pyridox(am)ine 5'-Phosphate Oxidase Deficiency.

Science.gov (United States)

Guerriero, Réjean M; Patel, Archana A; Walsh, Brian; Baumer, Fiona M; Shah, Ankoor S; Peters, Jurriaan M; Rodan, Lance H; Agrawal, Pankaj B; Pearl, Phillip L; Takeoka, Masanori

2017-11-01

Pyridoxine is converted to its biologically active form pyridoxal-5-phosphate (P5P) by the enzyme pyridox(am)ine 5'-phosphate oxidase and serves as a cofactor in nearly 200 reactions in the central nervous system. Pyridox(am)ine 5'-phosphate oxidase deficiency leads to P5P dependent epilepsy, typically a neonatal- or infantile-onset epileptic encephalopathy treatable with P5P or in some cases, pyridoxine. Following identification of retinopathy in a patient with pyridox(am)ine 5'-phosphate oxidase deficiency that was reversible with P5P therapy, we describe the systemic manifestations of pyridox(am)ine 5'-phosphate oxidase deficiency. A series of six patients with homozygous mutations of PNPO, the gene coding pyridox(am)ine 5'-phosphate oxidase, were evaluated in our center over the course of two years for phenotyping of neurological and systemic manifestations. Five of six were born prematurely, three had anemia and failure to thrive, and two had elevated alkaline phosphatase. A movement disorder was observed in two children, and a reversible retinopathy was observed in the most severely affected infant. All patients had neonatal-onset epilepsy and were on a continuum of developmental delay to profound encephalopathy. Electroencephalographic features included background slowing and disorganization, absent sleep features, and multifocal and generalized epileptiform discharges. All the affected probands carried a homozygous PNPO mutation (c.674 G>T, c.686 G>A and c.352G>A). In addition to the well-described epileptic encephalopathy, pyridox(am)ine 5'-phosphate oxidase deficiency causes a range of neurological and systemic manifestations. A movement disorder, developmental delay, and encephalopathy, as well as retinopathy, anemia, and failure to thrive add to the broadening clinical spectrum of P5P dependent epilepsy. Copyright © 2017 Elsevier Inc. All rights reserved.

Fluorescent Probes for Analysis and Imaging of Monoamine Oxidase Activity

Energy Technology Data Exchange (ETDEWEB)

Kim, Dokyoung; Jun, Yong Woong; Ahn, Kyo Han [POSTECH, Pohang (Korea, Republic of)

2014-05-15

Monoamine oxidases catalyze the oxidative deamination of dietary amines and amine neurotransmitters, and assist in maintaining the homeostasis of the amine neurotransmitters in the brain. Dysfunctions of these enzymes can cause neurological and behavioral disorders including Parkinson's and Alzheimer's diseases. To understand their physiological roles, efficient assay methods for monoamine oxidases are essential. Reviewed in this Perspective are the recent progress in the development of fluorescent probes for monoamine oxidases and their applications to enzyme assays in cells and tissues. It is evident that still there is strong need for a fluorescent probe with desirable substrate selectivity and photophysical properties to challenge the much unsolved issues associated with the enzymes and the diseases.

[Experimental rationale for the parameters of a rapid method for oxidase activity determination].

Science.gov (United States)

Butorina, N N

2010-01-01

Experimental rationale is provided for the parameters of a rapid (1-2-min) test to concurrently determine the oxidase activity of all bacteria grown on the membrane filter after water filtration. Oxidase reagents that are the aqueous solutions of tetramethyl-p-phenylenediamine dihydrochloride and demethyl-p-phenylenediamine dihydrochloride have been first ascertained to exert no effect on the viability and enzymatic activity of bacteria after one-hour contact. An algorithm has been improved for the rapid oxidase activity test: the allowable time for bacteria to contact oxidase reagents and procedures for minimizing the effect on bacterial biochemical activity following the contact. An accelerated method based on lactose medium with tergitol 7 and Endo agar has been devised to determine coliform bacteria, by applying the rapid oxidase test: the time of a final response is 18-24 hours. The method has been included into GOST 52426-2005.

Action of DCCD on the H+/O stoichiometry of mitoplast cytochrome c oxidase.

Science.gov (United States)

Lehninger, A L; Reynafarje, B; Costa, L

1985-01-01

The mechanistic H+/O ejection stoichiometry of the cytochrome c oxidase reaction in rat liver mitoplasts is close to 4 at level flow when the reduced oxidase is pulsed with O2. Dicyclohexylcarbodiimide (DCCD) up to 30 nmol/mg protein fails to influence the rate of electron flow through the mitoplast oxidase, but inhibits H+ ejection. The inhibition of H+ ejection appears to be biphasic; ejection of 2-3 H+ per O is completely inhibited by very low DCCD, whereas inhibition of the remaining H+ ejection requires very much higher concentrations of DCCD. This effect suggests the occurrence of two types of H+ pumps in the native cytochrome oxidase of mitoplasts.

Characterization of Two VAO-Type Flavoprotein Oxidases from Myceliophthora thermophila

Directory of Open Access Journals (Sweden)

Alessandro R. Ferrari

2018-01-01

Full Text Available The VAO flavoprotein family consists mostly of oxidoreductases harboring a covalently linked flavin cofactor. The linkage can be either monocovalent at position 8 with a histidine or tyrosine or bicovalent at position 8 with a histidine and at position 6 with a cysteine. Bicovalently bound flavoproteins show a preference for bulkier substrates such as oligosaccharides or secondary metabolites. The genome of the thermophilic fungus Myceliophthora thermophila C1 was found to be rich in genes encoding putative covalent VAO-type flavoproteins. Enzymes from this fungus have the advantage of being rather thermostable and homologous overexpression in M. thermophila C1 is feasible. Recently we discovered a new and VAO-type carbohydrate oxidase from this fungus: xylooligosaccharide oxidase. In this study, two other putative VAO-type oxidases, protein sequence XP_003663615 (MtVAO615 and XP_003665713 (MtVAO713, were expressed in M. thermophila C1, purified and characterized. Enzyme MtVAO615 was found to contain a bicovalently bound FAD, while enzyme MtVAO713 contained a monocovalent histidyl-bound FAD. The crystal structures of both proteins were obtained which revealed atypical active site architectures. It could be experimentally verified that both proteins, when reduced, rapidly react with molecular oxygen, a hallmark of flavoprotein oxidases. A large panel of alcohols, including carbohydrates, steroids and secondary alcohols were tested as potential substrates. For enzyme MtVAO713 low oxidase activity was discovered towards ricinoleic acid.

Vanillyl-alcohol oxidase, a tasteful biocatalyst

NARCIS (Netherlands)

Heuvel, van den R.H.H.; Fraaije, M.W.; Mattevi, A.; Laane, C.; Berkel, van W.J.H.

2001-01-01

The covalent flavoenzyme vanillyl-alcohol oxidase (VAO) is a versatile biocatalyst. It converts a wide range of phenolic compounds by catalysing oxidation, deamination, demethylation, dehydrogenation and hydroxylation reactions. The production of natural vanillin, 4-hydroxybenzaldehyde, coniferyl

An intron capture strategy used to identify and map a lysyl oxidase-like gene on chromosome 9 in the mouse

Energy Technology Data Exchange (ETDEWEB)

Wydner, K.S.; Passmore, H.C. [Rutgers Univ., Piscataway, NJ (United States); Kim, Houngho; Csiszar, K.; Boyd, C.D. [UMDNJ, New Brunswick, NJ (United States)

1997-03-01

An intron capture strategy involving use of polymerase chain reaction was used to identify and map the mouse homologue of a human lysyl oxidase-like gene (LOXL). Oligonucleotides complementary to conserved domains within exons 4 and 5 of the human lysyl oxidase-like gene were used to amplify the corresponding segment from mouse genomic DNA. Sequencing of the resulting mouse DNA fragment of approximately 1 kb revealed that the exon sequences at the ends of the amplified fragment are highly homologous (90% nucleotide identity) to exons 4 and 5 of the human lysyl oxidase-like gene. An AluI restriction site polymorphism within intron 4 was used to map the mouse lysyl oxidase-like gene (Loxl) to mouse Chromosome 9 in a region that shares linkage conservation with human chromosome 15q24, to which the LOXL was recently mapped. 22 refs., 3 figs.

TtMCO: A highly thermostable laccase-like multicopper oxidase from the thermophilic Thermobaculum terrenum

DEFF Research Database (Denmark)

Brander, Søren; Mikkelsen, Jørn Dalgaard; Kepp, Kasper Planeta

2015-01-01

This paper reports the identification, heterologous expression in Escherichia coli and characterization of TtMCO from the thermophilic bacterium Thermobaculum terrenum, the first laccase-like multi-copper oxidase (LMCO) from the distinct Phylum Chloroflexi. TtMCO has only 39% identity to its...... closest characterized homologue, CotA from Bacillus subtilis, but sequence and spectrophotometry confirmed copper coordination similar to that of LMCOs. TtMCO is extremely thermophilic with a half-time of inactivation of 2.24 days at 70 degrees C and 350 min at 80°C and pH 7, consistent...

Immobilization of Glucose Oxidase on Modified-Carbon-Paste-Electrodes for Microfuel Cell

Directory of Open Access Journals (Sweden)

Laksmi Ambarsari

2016-03-01

Full Text Available Glucose oxidase (GOx is being developed for many applications such as an implantable fuel cell, due to its attractive property of operating under physiological conditions. This study reports the functional immobilization of glucose oxidase onto polyaniline-nanofiber-modified-carbon-paste-electrodes (GOx/MCPE as bioanodes in fuel cell applications. In particular, GOx is immobilized onto the electrode surface via a linker molecule (glutaraldehyde. Polyaniline, synthesized by the interfacial polymerization method, produces a morphological form of nanofibers (100-120 nm which have good conductivity. The performance of the polyaniline-modified-carbon-paste-electrode (MCPE was better than the carbon- paste-electrode (CPE alone. The optimal pH and temperature of the GOx/MCPE were 4.5 (in 100 mM acetate buffer and 65 °C, respectively. The GOx/MCPE exhibit high catalytic performances (activation energy 16.4 kJ mol-1, have a high affinity for glucose (Km value 37.79 µM and can have a maximum current (Imax of 3.95 mA. The sensitivity of the bioelectrode also was high at 57.79 mA mM-1 cm-2.

Absorption of enzymatically active 125I-labeled bovine milk xanthine oxidase fed to rabbits

International Nuclear Information System (INIS)

Rzucidlo, S.J.; Zikakis, J.P.

1990-01-01

Rabbits fed a regular laboratory diet supplemented with a high-fat milk containing xanthine oxidase (XO) were studied to determine the presence of active XO in the blood. A pilot feeding study, where rabbits consumed a high-fat diet containing xanthine oxidase, showed a correlation between dairy food consumption and XO activity in the blood. Antibody to dietary XO was also found. In a second study, rabbits were fed ad libitum the high-fat milk and blood serum samples were tested weekly for XO activity. No elevation in serum XO activity was found. A third study showed that serum XO activity was increased when rabbits were force fed the high-fat milk. The final study consisted of force feeding 125 I-labeled XO to one rabbit to ascertain whether the observed increase in serum XO was due to dietary or endogenous XO. Isoelectric focusing of sera collected from the test rabbit strongly suggested that at least a portion of the serum XO contained the radioactive label. This is the first direct evidence showing the uptake of dietary active XO from the gut

Substrate specificity of flavin-dependent vanillyl-alcohol oxidase from Penicillium simplicissimum. Evidence for the production of 4-hydroxycinnamyl alcohols from 4-allylphenols.

Science.gov (United States)

Fraaije, M W; Veeger, C; van Berkel, W J

1995-11-15

The substrate specificity of the flavoprotein vanillyl-alcohol oxidase from Penicillium simplicissimum was investigated. Vanillyl-alcohol oxidase catalyzes besides the oxidation of 4-hydroxybenzyl alcohols, the oxidative deamination of 4-hydroxybenzylamines and the oxidative demethylation of 4-(methoxymethyl)phenols. During the conversion of vanillylamine to vanillin, a transient intermediate, most probably vanillylimine, is observed. Vanillyl-alcohol oxidase weakly interacts with 4-hydroxyphenylglycols and a series of catecholamines. These compounds are converted to the corresponding ketones. Both enantiomers of (nor)epinephrine are substrates for vanillyl-alcohol oxidase, but the R isomer is preferred. Vanillyl-alcohol oxidase is most active with chavicol and eugenol. These 4-allylphenols are converted to coumaryl alcohol and coniferyl alcohol, respectively. Isotopic labeling experiments show that the oxygen atom inserted at the C gamma atom of the side chain is derived from water. The 4-hydroxycinnamyl alcohol products and the substrate analog isoeugenol are competitive inhibitors of vanillyl alcohol oxidation. The binding of isoeugenol to the oxidized enzyme perturbs the optical spectrum of protein-bound FAD. pH-dependent binding studies suggest that vanillyl-alcohol oxidase preferentially binds the phenolate form of isoeugenol (pKa < 6, 25 degrees C). From this and the high pH optimum for turnover, a hydride transfer mechanism involving a p-quinone methide intermediate is proposed for the vanillyl-alcohol-oxidase-catalyzed conversion of 4-allylphenols.

The Role of Aldehyde Oxidase and Xanthine Oxidase in the Biotransformation of a Novel Negative Allosteric Modulator of Metabotropic Glutamate Receptor Subtype 5

Science.gov (United States)

Morrison, Ryan D.; Blobaum, Anna L.; Byers, Frank W.; Santomango, Tammy S.; Bridges, Thomas M.; Stec, Donald; Brewer, Katrina A.; Sanchez-Ponce, Raymundo; Corlew, Melany M.; Rush, Roger; Felts, Andrew S.; Manka, Jason; Bates, Brittney S.; Venable, Daryl F.; Rodriguez, Alice L.; Jones, Carrie K.; Niswender, Colleen M.; Conn, P. Jeffrey; Lindsley, Craig W.; Emmitte, Kyle A.

2012-01-01

Negative allosteric modulation (NAM) of metabotropic glutamate receptor subtype 5 (mGlu5) represents a therapeutic strategy for the treatment of childhood developmental disorders, such as fragile X syndrome and autism. VU0409106 emerged as a lead compound within a biaryl ether series, displaying potent and selective inhibition of mGlu5. Despite its high clearance and short half-life, VU0409106 demonstrated efficacy in rodent models of anxiety after extravascular administration. However, lack of a consistent correlation in rat between in vitro hepatic clearance and in vivo plasma clearance for the biaryl ether series prompted an investigation into the biotransformation of VU0409106 using hepatic subcellular fractions. An in vitro appraisal in rat, monkey, and human liver S9 fractions indicated that the principal pathway was NADPH-independent oxidation to metabolite M1 (+16 Da). Both raloxifene (aldehyde oxidase inhibitor) and allopurinol (xanthine oxidase inhibitor) attenuated the formation of M1, thus implicating the contribution of both molybdenum hydroxylases in the biotransformation of VU0409106. The use of 18O-labeled water in the S9 experiments confirmed the hydroxylase mechanism proposed, because 18O was incorporated into M1 (+18 Da) as well as in a secondary metabolite (M2; +36 Da), the formation of which was exclusively xanthine oxidase-mediated. This unusual dual and sequential hydroxylase metabolism was confirmed in liver S9 and hepatocytes of multiple species and correlated with in vivo data because M1 and M2 were the principal metabolites detected in rats administered VU0409106. An in vitro-in vivo correlation of predicted hepatic and plasma clearance was subsequently established for VU0409106 in rats and nonhuman primates. PMID:22711749

Cytochemical localization of catalase and several hydrogen peroxide-producing oxidases in the nucleoids and matrix of rat liver peroxisomes

NARCIS (Netherlands)

Veenhuis, M.; Wendelaar Bonga, S.E.

1979-01-01

The distribution of catalase, amino acid oxidase, α-hydroxy acid oxidase, urate oxidase and alcohol oxidase was studied cytochemically in rat hepatocytes. The presence of catalase was demonstrated with the conventional diaminobenzidine technique. Oxidase activities were visualized with methods based

Lysyl Oxidase and the Tumor Microenvironment

Directory of Open Access Journals (Sweden)

Tong-Hong Wang

2016-12-01

Full Text Available The lysyl oxidase (LOX family of oxidases contains a group of extracellular copper-dependent enzymes that catalyze the cross-linking of collagen and elastin by oxidation, thus maintaining the rigidity and structural stability of the extracellular matrix (ECM. Aberrant expression or activation of LOX alters the cellular microenvironment, leading to many diseases, including atherosclerosis, tissue fibrosis, and cancer. Recently, a number of studies have shown that LOX is overexpressed in most cancers and that it is involved in the regulation of tumor progression and metastasis. In contrast, a few reports have also indicated the tumor-suppressing role of LOX. In this short review, we discuss recent research on the correlations between LOX and cancer. Further, the role of LOX in tumor microenvironment remodeling, tumorigenesis, and metastasis and the underlying mechanisms have also been elucidated.

Are colorimetric assays appropriate for measuring phenol oxidase activity in peat soils?

Science.gov (United States)

Magdalena M. Wiedermann; Evan S. Kane; Timothy J. Veverica; Erik A. Lilleskov

2017-01-01

The activity of extracellular phenol oxidases is believed to play a critical role in decomposition processes in peatlands. The water logged, acidic conditions, and recalcitrant litter from the peatland vegetation, lead to exceptionally high phenolics in the peat. In order to quantify the activity of oxidative enzymes involved in the modification and break down of...

Plasma amine oxidase activities in Norrie disease patients with an X-chromosomal deletion affecting monoamine oxidase.

Science.gov (United States)

Murphy, D L; Sims, K B; Karoum, F; Garrick, N A; de la Chapelle, A; Sankila, E M; Norio, R; Breakefield, X O

1991-01-01

Two individuals with an X-chromosomal deletion were recently found to lack the genes encoding monoamine oxidase type A (MAO-A) and MAO-B. This abnormality was associated with almost total (90%) reductions in the oxidatively deaminated urinary metabolites of the MAO-A substrate, norepinephrine, and with marked (100-fold) increases in an MAO-B substrate, phenylethylamine, confirming systemic functional consequences of the genetic enzyme deficiency. However, urinary concentrations of the deaminated metabolites of dopamine and serotonin (5-HT) were essentially normal. To investigate other deaminating systems besides MAO-A and MAO-B that might produce these metabolites of dopamine and 5-HT, we examined plasma amine oxidase (AO) activity in these two patients and two additional patients with the same X-chromosomal deletion. Normal plasma AO activity was found in all four Norrie disease-deletion patients, in four patients with classic Norrie disease without a chromosomal deletion, and in family members of patients from both groups. Marked plasma amine metabolite abnormalities and essentially absent platelet MAO-B activity were found in all four Norrie disease-deletion patients, but in none of the other subjects in the two comparison groups. These results indicate that plasma AO is encoded by gene(s) independent of those for MAO-A and MAO-B, and raise the possibility that plasma AO, and perhaps the closely related tissue AO, benzylamine oxidase, as well as other atypical AOs or MAOs encoded independently from MAO-A and MAO-B may contribute to the oxidative deamination of dopamine and 5-HT in humans.

The microglial NADPH oxidase complex as a source of oxidative stress in Alzheimer's disease

Directory of Open Access Journals (Sweden)

Landreth Gary E

2006-11-01

Full Text Available Abstract Alzheimer's disease is the most common cause of dementia in the elderly, and manifests as progressive cognitive decline and profound neuronal loss. The principal neuropathological hallmarks of Alzheimer's disease are the senile plaques and the neurofibrillary tangles. The senile plaques are surrounded by activated microglia, which are largely responsible for the proinflammatory environment within the diseased brain. Microglia are the resident innate immune cells in the brain. In response to contact with fibrillar beta-amyloid, microglia secrete a diverse array of proinflammatory molecules. Evidence suggests that oxidative stress emanating from activated microglia contribute to the neuronal loss characteristic of this disease. The source of fibrillar beta-amyloid induced reactive oxygen species is primarily the microglial nicotinamide adenine dinucleotide phosphate (NADPH oxidase. The NADPH oxidase is a multicomponent enzyme complex that, upon activation, produces the highly reactive free radical superoxide. The cascade of intracellular signaling events leading to NADPH oxidase assembly and the subsequent release of superoxide in fibrillar beta-amyloid stimulated microglia has recently been elucidated. The induction of reactive oxygen species, as well as nitric oxide, from activated microglia can enhance the production of more potent free radicals such as peroxynitrite. The formation of peroxynitrite causes protein oxidation, lipid peroxidation and DNA damage, which ultimately lead to neuronal cell death. The elimination of beta-amyloid-induced oxidative damage through the inhibition of the NADPH oxidase represents an attractive therapeutic target for the treatment of Alzheimer's disease.

Inactivation of 1-aminocyclopropane-1-carboxylate oxidase involves oxidative modifications.

Science.gov (United States)

Barlow, J N; Zhang, Z; John, P; Baldwin, J E; Schofield, C J

1997-03-25

1-Aminocyclopropane-1-carboxylate (ACC) oxidase catalyzes the final step in the biosynthesis of the plant signaling molecule ethylene. It is a member of the ferrous iron dependent family of oxidases and dioxygenases and is unusual in that it displays a very short half-life under catalytic conditions, typically less than 20 min, and a requirement for CO2 as an activator. The rates of inactivation of purified, recombinant ACC oxidase from tomato under various combinations of substrates and cofactors were measured. Inactivation was relatively slow in the presence of buffer alone (t1/2 > 1 h), but fast in the presence of ferrous iron and ascorbate (t1/2 approximately 10 min). The rate of iron/ascorbate-mediated inactivation was increased by the addition of ACC, unaffected by the addition of CO2 at saturation (supplied as bicarbonate) but decreased by the addition of catalase or ACC + CO2 at saturation (supplied as bicarbonate). Iron/ascorbate-mediated inactivation was accompanied by partial proteolysis as observed by SDS-PAGE analysis. The fragmentation pattern was altered when ACC was also included, suggesting that ACC can bind to ACC oxidase in the absence of bicarbonate. N-terminal sequencing of fragments resulted in identification of an internal cleavage site which we propose is proximate to active-site bound iron. Thus, ACC oxidase inactivates via relatively slow partial unfolding of the catalytically active conformation, oxidative damage mediated via hydrogen peroxide which is catalase protectable and oxidative damage to the active site which results in partial proteolysis and is not catalase protectable.

Genetics Home Reference: isolated sulfite oxidase deficiency

Science.gov (United States)

... and Management Resources (1 link) GeneReview: Isolated Sulfite Oxidase Deficiency General Information from MedlinePlus (5 links) Diagnostic Tests Drug Therapy Genetic Counseling Palliative Care Surgery and ...

Up-Streaming Process for Glucose Oxidase by Thermophilic Penicillium sp. in Shake Flask

OpenAIRE

Muhammad Mohsin JAVED; Aroosh SHABIR; Sana ZAHOOR; Ikram UL-HAQ

2012-01-01

The present study is concerned with the production of glucose oxidase (GOD) from thermophilic Penicillium sp. in 250 mL shake flask. Fourteen different strains of thermophilic Penicillium sp. were isolated from the soil and were screened for glucose oxidase production. IIBP-13 strain gave maximum extra-cellular glucose oxidase production as compared to other isolates. Effect of submerged fermentation in shaking and static conditions, different carbon sources and incubation period on the produ...

Preparation of Cu{sub 2}O nanoparticles in cupric chloride solutions with a simple mechanochemical approach

Energy Technology Data Exchange (ETDEWEB)

Chen, D., E-mail: ma97chen@hotamil.co [School of Materials Science and Engineering, Hunan University, Changsha, 410082 (China); Graduate School of Energy Science, Kyoto University, 606-8501, Kyoto (Japan); Ni, S. [School of Materials Science and Engineering, Hunan University, Changsha, 410082 (China); Fang, J.J. [College of Electromechanical Engineering, North China University of Technology, Beijing, 100041 (China); Xiao, T. [School of Materials Science and Engineering, Hunan University, Changsha, 410082 (China)

2010-08-15

The cuprous oxide (Cu{sub 2}O) nanoparticles with diameter of 50-150 nm are prepared by high-energy ball milling in the various CuCl{sub 2} solutions with different [Cl{sup -}] concentration. The as-synthesized products are characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM) and high-resolution transmission electron microscopy (HRTEM). Finally, the effects of [Cl{sup -}] concentrations on the formation of cuprous oxide and reaction mechanism are discussed.

In vitro analysis of the role of glucose oxidase from Talaromyces flavus in biocontrol of the plant pathogen Verticillium dahliae.

OpenAIRE

Stosz, S K; Fravel, D R; Roberts, D P

1996-01-01

Culture filtrates from Talaromyces flavus grown on glucose contained high levels of glucose oxidase activity, while culture filtrates from T. flavus grown on xylan contained negligible glucose oxidase activity. Culture filtrates from T-flavus grown on both media contained complex protein profiles. However, only culture filtrates from T. flavus grown on glucose inhibited germination of microsclerotia of Verticillium dahliae in in vitro inhibition assays. A polyclonal antiserum preparation, pAB...

Barley polyamine oxidase: Characterisation and analysis of the cofactor and the N-terminal amino acid sequence

DEFF Research Database (Denmark)

Radova, A.; Sebela, M.; Galuszka, P.

2001-01-01

This paper reports the first purification method developed for the isolation of an homogeneous polyamine oxidase (PAO) from etiolated barley seedlings. The crude enzyme preparation was obtained after initial precipitation of the extract with protamine sulphate and ammonium sulphate. The enzyme...... was further confirmed by measuring the fluorescence spectra, Barley PAO is an acidic protein (pI 5.4) containing 3% of neutral sugars: its molecular mass determined by SDS-PAGE was 56 kDa, whilst gel permeation chromatography revealed the higher value of 76 kDa. The N-terminal amino acid sequence of barley...... PAO shows a high degree of similarity to that of maize PAO and to several other flavoprotein oxidases. The polyamines spermine and spermidine were the only two substrates of the enzyme with K-m values 4 x 10(-5) and 3 x 10(-5) M and pH optima of 5.0 and 6.0, respectively. Barley polyamine oxidase...

Two X-linked chronic granulomatous disease patients with unusual NADPH oxidase properties

NARCIS (Netherlands)

Wolach, Baruch; Broides, Arnon; Zeeli, Tal; Gavrieli, Ronit; de Boer, Martin; van Leeuwen, Karin; Levy, Jacov; Roos, Dirk

2011-01-01

Chronic granulomatous disease (CGD) is an immune deficiency syndrome caused by defects in the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, the enzyme that generates reactive oxygen species (ROS) in phagocytizing leukocytes. This study evaluates the NADPH oxidase capacity in two

Effects of high hydrostatic pressure or hydrophobic modification on thermal stability of xanthine oxidase.

Science.gov (United States)

Halalipour, Ali; Duff, Michael R; Howell, Elizabeth E; Reyes-De-Corcuera, José I

2017-08-01

The effect of high hydrostatic pressure (HHP) on the kinetics of thermal inactivation of xanthine oxidase (XOx) from bovine milk was studied. Inactivation of XOx followed pseudo-first-order kinetics at 0.1-300MPa and 55.0-70.0°C. High pressure up to at least 300MPa stabilized XOx at all the studied temperatures. The highest stabilization effect of HHP on XOx was at 200-300MPa at 55.0 and 58.6°C, and at 250-300MPa at 62.3-70.0°C. The stability of XOx increased 9.5 times at 300MPa and 70.0°C compared to atmospheric pressure at the same temperature. The activation energy of inactivation of XOx decreased with pressure and was 1.9 times less at 300MPa (97.0±8.2kJmol -1 ) than at 0.1MPa (181.7±12.1kJmol -1 ). High pressure decreased the dependence of the rate constant of inactivation to temperature effects compared to atmospheric pressure. The stabilizing effect of HHP on XOx was highest at 70.0°C where the activation volume of inactivation of XOx was 28.9±2.9cm 3 mol -1 . A second approach to try to increase XOx stability involved hydrophobic modification using aniline or benzoate. However, the thermal stability of XOx remained unaffected after 8-14 modifications of carboxyl side groups per XOx monomer with aniline, or 12-17 modifications of amino side groups per XOx monomer with benzoate. Copyright © 2017 Elsevier Inc. All rights reserved.

Biogenic manganese oxide nanoparticle formation by a multimeric multicopper oxidase Mnx

Energy Technology Data Exchange (ETDEWEB)

Romano, Christine A.; Zhou, Mowei; Song, Yang; Wysocki, Vicki H.; Dohnalkova, Alice C.; Kovarik, Libor; Paša-Tolić, Ljiljana; Tebo, Bradley M.

2017-09-29

Bacteria that produce Mn oxides are extraordinarily skilled engineers of nanomaterials that contribute significantly to global biogeochemical cycles. Their enzyme-based reaction mechanisms may be genetically tailored for environmental remediation applications or bioenergy production. However, significant challenges exist for structural characterization of the enzymes responsible for biomineralization. The active Mn oxidase, Mnx, in Bacillus sp. PL-12 is a complex composed of a multicopper oxidase (MCO), MnxG, and two accessory proteins MnxE and MnxF. MnxG shares sequence similarity with other, structurally characterized MCOs. However, MnxE and MnxF have no similarity to any characterized proteins. The ~200 kDa complex has been recalcitrant to crystallization, so its structure is unknown. In this study, native mass spectrometry defines the subunit topology and copper binding of the Mnx complex, while high resolution electron microscopy visualizes the protein and nascent Mn oxide minerals. These data provide critical structural information for conceptualizing how Mnx produces nanoparticulate Mn oxides.

Characterization of the cydAB-Encoded Cytochrome bd Oxidase from Mycobacterium smegmatis

Science.gov (United States)

Kana, Bavesh D.; Weinstein, Edward A.; Avarbock, David; Dawes, Stephanie S.; Rubin, Harvey; Mizrahi, Valerie

2001-01-01

The cydAB genes from Mycobacterium smegmatis have been cloned and characterized. The cydA and cydB genes encode the two subunits of a cytochrome bd oxidase belonging to the widely distributed family of quinol oxidases found in prokaryotes. The cydD and cydC genes located immediately downstream of cydB encode a putative ATP-binding cassette-type transporter. At room temperature, reduced minus oxidized difference spectra of membranes purified from wild-type M. smegmatis displayed spectral features that are characteristic of the γ-proteobacterial type cytochrome bd oxidase. Inactivation of cydA or cydB by insertion of a kanamycin resistance marker resulted in loss of d-heme absorbance at 631 nm. The d-heme could be restored by transformation of the M. smegmatis cyd mutants with a replicating plasmid carrying the highly homologous cydABDC gene cluster from Mycobacterium tuberculosis. Inactivation of cydA had no effect on the ability of M. smegmatis to exit from stationary phase at 37 or 42°C. The growth rate of the cydA mutant was tested under oxystatic conditions. Although no discernible growth defect was observed under moderately aerobic conditions (9.2 to 37.5 × 102 Pa of pO2 or 5 to 21% air saturation), the mutant displayed a significant growth disadvantage when cocultured with the wild type under extreme microaerophilia (0.8 to 1.7 × 102 Pa of pO2 or 0.5 to 1% air saturation). These observations were in accordance with the two- to threefold increase in cydAB gene expression observed upon reduction of the pO2 of the growth medium from 21 to 0.5% air saturation and with the concomitant increase in d-heme absorbance in spectra of membranes isolated from wild-type M. smegmatis cultured at 1% air saturation. Finally, the cydA mutant displayed a competitive growth disadvantage in the presence of the terminal oxidase inhibitor, cyanide, when cocultured with wild type at 21% air saturation in an oxystat. In conjunction with these findings, our results suggest that

Purification, characterization and decolorization of bilirubin oxidase from Myrothecium verrucaria 3.2190

Science.gov (United States)

Myrothecium verrucaria 3.2190 is a nonligninolytic fungus that produces bilirubin oxidase. Both Myrothecium verrucaria and the extracellular bilirubin oxidase were tested for their ability to decolorize indigo carmine. The biosorption and biodegradation of the dye were detected during the process of...

Estrogen inhibits lysyl oxidase and decreases mechanical function in engineered ligaments.

Science.gov (United States)

Lee, Cassandra A; Lee-Barthel, Ann; Marquino, Louise; Sandoval, Natalie; Marcotte, George R; Baar, Keith

2015-05-15

Women are more likely to suffer an anterior cruciate ligament (ACL) rupture than men, and the incidence of ACL rupture in women rises with increasing estrogen levels. We used an engineered ligament model to determine how an acute rise in estrogen decreases the mechanical properties of ligaments. Using fibroblasts isolated from human ACLs from male or female donors, we engineered ligaments and determined that ligaments made from female ACL cells had more collagen and were equal in strength to those made from male ACL cells. We then treated engineered ligaments for 14 days with low (5 pg/ml), medium (50 pg/ml), or high (500 pg/ml) estrogen, corresponding to the range of in vivo serum estrogen concentrations and found that collagen within the grafts increased without a commensurate increase in mechanical strength. Mimicking the menstrual cycle, with 12 days of low estrogen followed by 2 days of physiologically high estrogen, resulted in a decrease in engineered ligament mechanical function with no change in the amount of collagen in the graft. The decrease in mechanical stiffness corresponded with a 61.7 and 76.9% decrease in the activity of collagen cross-linker lysyl oxidase with 24 and 48 h of high estrogen, respectively. Similarly, grafts treated with the lysyl oxidase inhibitor β-aminoproprionitrile (BAPN) for 24 h showed a significant decrease in ligament mechanical strength [control (CON) = 1.58 ± 0.06 N; BAPN = 1.06 ± 0.13 N] and stiffness (CON = 7.7 ± 0.46 MPa; BAPN = 6.1 ± 0.71 MPa) without changing overall collagen levels (CON = 396 ± 11.5 μg; BAPN = 382 ± 11.6 μg). Together, these data suggest that the rise in estrogen during the follicular phase decreases lysyl oxidase activity in our engineered ligament model and if this occurs in vivo may decrease the stiffness of ligaments and contribute to the elevated rate of ACL rupture in women. Copyright © 2015 the American Physiological Society.

Expanding the substrate scope of chitooligosaccharide oxidase from Fusarium graminearum by structure-inspired mutagenesis

NARCIS (Netherlands)

Ferrari, Alessandro; Lee, Misun; Fraaije, Marco

2015-01-01

Chitooligosaccharide oxidase from Fusarium graminearum (ChitO) oxidizes N-acetyl-D-glucosamine (GlcNAc) and its oligomers with high efficiency at the C1-hydroxyl moiety while it shows poor or no activity with other carbohydrates. By sequence and structural comparison with other known carbohydrate

Blockade of TGF-β 1 Signalling Inhibits Cardiac NADPH Oxidase Overactivity in Hypertensive Rats

Directory of Open Access Journals (Sweden)

José Luis Miguel-Carrasco

2012-01-01

Full Text Available NADPH oxidases constitute a major source of superoxide anion (⋅O2 - in hypertension. Several studies suggest an important role of NADPH oxidases in different effects mediated by TGF-β 1. In this study we show that chronic administration of P144, a peptide synthesized from type III TGF-β 1 receptor, significantly reduced the cardiac NADPH oxidase expression and activity as well as in the nitrotyrosine levels observed in control spontaneously hypertensive rats (V-SHR to levels similar to control normotensive Wistar Kyoto rats. In addition, P144 was also able to reduce the significant increases in the expression of collagen type I protein and mRNA observed in hearts from V-SHR. In addition, positive correlations between collagen expression, NADPH oxidase activity, and nitrotyrosine levels were found in all animals. Finally, TGF-β 1-stimulated Rat-2 exhibited significant increases in NADPH oxidase activity that was inhibited in the presence of P144. It could be concluded that the blockade of TGF-β 1 with P144 inhibited cardiac NADPH oxidase in SHR, thus adding new data to elucidate the involvement of this enzyme in the profibrotic actions of TGF-β 1.

Herbivore-plant interactions: mixed-function oxidases and secondary plant substances.

Science.gov (United States)

Brattsten, L B; Wilkinson, C F; Eisner, T

1977-06-17

The mixed-function oxidases of a polyphagous insect larva (the southern armyworm, Spodoptera eridania) were found to be induced by a diversity of secondary plant substances. The induction proceeds rapidly and in response to a small quantity of secondary substance. Following induction, the larva is less susceptible to dietary poisoning. It is argued that mixed-function oxidases play a major role in protecting herbivores against chemical stress from secondary plant substances.

Functional heterogeneity of NADPH oxidase-mediated contractions to endothelin with vascular aging.

Science.gov (United States)

Meyer, Matthias R; Barton, Matthias; Prossnitz, Eric R

2014-11-24

Aging, a physiological process and main risk factor for cardiovascular and renal diseases, is associated with endothelial cell dysfunction partly resulting from NADPH oxidase-dependent oxidative stress. Because increased formation of endothelium-derived endothelin-1 (ET-1) may contribute to vascular aging, we studied the role of NADPH oxidase function in age-dependent contractions to ET-1. Renal arteries and abdominal aortas from young and old C57BL6 mice (4 and 24 months of age) were prepared for isometric force measurements. Contractions to ET-1 (0.1-100 nmol/L) were determined in the presence and absence of the NADPH oxidase-selective inhibitor gp91ds-tat (3 μmol/L). To exclude age-dependent differential effects of NO bioactivity between vascular beds, all experiments were conducted in the presence of the NO synthase inhibitor L-NAME (300 μmol/L). In young animals, ET-1-induced contractions were 6-fold stronger in the renal artery than in the aorta (prenal artery and aorta, respectively (pAging had no effect on NADPH oxidase-dependent and -independent contractions to ET-1 in the renal artery. In contrast, contractions to ET-1 were markedly reduced in the aged aorta (5-fold, page-dependent heterogeneity of NADPH oxidase-mediated vascular contractions to ET-1, demonstrating an inherent resistance to functional changes in the renal artery but not in the aorta with aging. Thus, local activity of NADPH oxidase differentially modulates responses to ET-1 with aging in distinct vascular beds. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

Targeting NADPH oxidase decreases oxidative stress in the transgenic sickle cell mouse penis.

Science.gov (United States)

Musicki, Biljana; Liu, Tongyun; Sezen, Sena F; Burnett, Arthur L

2012-08-01

Sickle cell disease (SCD) is a state of chronic vasculopathy characterized by endothelial dysfunction and increased oxidative stress, but the sources and mechanisms responsible for reactive oxygen species (ROS) production in the penis are unknown. We evaluated whether SCD activates NADPH oxidase, induces endothelial nitric oxide synthase (eNOS) uncoupling, and decreases antioxidants in the SCD mouse penis. We further tested the hypothesis that targeting NADPH oxidase decreases oxidative stress in the SCD mouse penis. SCD transgenic (sickle) mice were used as an animal model of SCD. Hemizygous (hemi) mice served as controls. Mice received an NADPH oxidase inhibitor apocynin (10 mM in drinking water) or vehicle. Penes were excised at baseline for molecular studies. Markers of oxidative stress (4-hydroxy-2-nonenal [HNE]), sources of ROS (eNOS uncoupling and NADPH oxidase subunits p67(phox) , p47(phox) , and gp91(phox) ), and enzymatic antioxidants (superoxide dismutase [SOD]1, SOD2, catalase, and glutathione peroxidase-1 [GPx1]) were measured by Western blot in penes. Sources of ROS, oxidative stress, and enzymatic antioxidants in the SCD penis. Relative to hemi mice, SCD increased (Ppenis. Apocynin treatment of sickle mice reversed (P0.05) prevented eNOS uncoupling in the penis. Apocynin treatment of hemi mice did not affect any of these parameters. NADPH oxidase and eNOS uncoupling are sources of oxidative stress in the SCD penis; decreased GPx1 further contributes to oxidative stress. Inhibition of NADPH oxidase upregulation decreases oxidative stress, implying a major role for NADPH oxidase as a ROS source and a potential target for improving vascular function in the SCD mouse penis. © 2012 International Society for Sexual Medicine.

Neutrophils to the ROScue: Mechanisms of NADPH Oxidase Activation and Bacterial Resistance

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Giang T. Nguyen

2017-08-01

Full Text Available Reactive oxygen species (ROS generated by NADPH oxidase play an important role in antimicrobial host defense and inflammation. Their deficiency in humans results in recurrent and severe bacterial infections, while their unregulated release leads to pathology from excessive inflammation. The release of high concentrations of ROS aids in clearance of invading bacteria. Localization of ROS release to phagosomes containing pathogens limits tissue damage. Host immune cells, like neutrophils, also known as PMNs, will release large amounts of ROS at the site of infection following the activation of surface receptors. The binding of ligands to G-protein-coupled receptors (GPCRs, toll-like receptors, and cytokine receptors can prime PMNs for a more robust response if additional signals are encountered. Meanwhile, activation of Fc and integrin directly induces high levels of ROS production. Additionally, GPCRs that bind to the bacterial-peptide analog fMLP, a neutrophil chemoattractant, can both prime cells and trigger low levels of ROS production. Engagement of these receptors initiates intracellular signaling pathways, resulting in activation of downstream effector proteins, assembly of the NADPH oxidase complex, and ultimately, the production of ROS by this complex. Within PMNs, ROS released by the NADPH oxidase complex can activate granular proteases and induce the formation of neutrophil extracellular traps (NETs. Additionally, ROS can cross the membranes of bacterial pathogens and damage their nucleic acids, proteins, and cell membranes. Consequently, in order to establish infections, bacterial pathogens employ various strategies to prevent restriction by PMN-derived ROS or downstream consequences of ROS production. Some pathogens are able to directly prevent the oxidative burst of phagocytes using secreted effector proteins or toxins that interfere with translocation of the NADPH oxidase complex or signaling pathways needed for its activation

Improving Glyphosate Oxidation Activity of Glycine Oxidase from Bacillus cereus by Directed Evolution

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Zhan, Tao; Zhang, Kai; Chen, Yangyan; Lin, Yongjun; Wu, Gaobing; Zhang, Lili; Yao, Pei; Shao, Zongze; Liu, Ziduo

2013-01-01

Glyphosate, a broad spectrum herbicide widely used in agriculture all over the world, inhibits 5-enolpyruvylshikimate-3-phosphate synthase in the shikimate pathway, and glycine oxidase (GO) has been reported to be able to catalyze the oxidative deamination of various amines and cleave the C-N bond in glyphosate. Here, in an effort to improve the catalytic activity of the glycine oxidase that was cloned from a glyphosate-degrading marine strain of Bacillus cereus (BceGO), we used a bacteriophage T7 lysis-based method for high-throughput screening of oxidase activity and engineered the gene encoding BceGO by directed evolution. Six mutants exhibiting enhanced activity toward glyphosate were screened from two rounds of error-prone PCR combined with site directed mutagenesis, and the beneficial mutations of the six evolved variants were recombined by DNA shuffling. Four recombinants were generated and, when compared with the wild-type BceGO, the most active mutant B3S1 showed the highest activity, exhibiting a 160-fold increase in substrate affinity, a 326-fold enhancement in catalytic efficiency against glyphosate, with little difference between their pH and temperature stabilities. The role of these mutations was explored through structure modeling and molecular docking, revealing that the Arg51 mutation is near the active site and could be an important residue contributing to the stabilization of glyphosate binding, while the role of the remaining mutations is unclear. These results provide insight into the application of directed evolution in optimizing glycine oxidase function and have laid a foundation for the development of glyphosate-tolerant crops. PMID:24223901

Improving glyphosate oxidation activity of glycine oxidase from Bacillus cereus by directed evolution.

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Tao Zhan

Full Text Available Glyphosate, a broad spectrum herbicide widely used in agriculture all over the world, inhibits 5-enolpyruvylshikimate-3-phosphate synthase in the shikimate pathway, and glycine oxidase (GO has been reported to be able to catalyze the oxidative deamination of various amines and cleave the C-N bond in glyphosate. Here, in an effort to improve the catalytic activity of the glycine oxidase that was cloned from a glyphosate-degrading marine strain of Bacillus cereus (BceGO, we used a bacteriophage T7 lysis-based method for high-throughput screening of oxidase activity and engineered the gene encoding BceGO by directed evolution. Six mutants exhibiting enhanced activity toward glyphosate were screened from two rounds of error-prone PCR combined with site directed mutagenesis, and the beneficial mutations of the six evolved variants were recombined by DNA shuffling. Four recombinants were generated and, when compared with the wild-type BceGO, the most active mutant B3S1 showed the highest activity, exhibiting a 160-fold increase in substrate affinity, a 326-fold enhancement in catalytic efficiency against glyphosate, with little difference between their pH and temperature stabilities. The role of these mutations was explored through structure modeling and molecular docking, revealing that the Arg(51 mutation is near the active site and could be an important residue contributing to the stabilization of glyphosate binding, while the role of the remaining mutations is unclear. These results provide insight into the application of directed evolution in optimizing glycine oxidase function and have laid a foundation for the development of glyphosate-tolerant crops.

Cyanobacterial Lactate Oxidases Serve as Essential Partners in N2 Fixation and Evolved into Photorespiratory Glycolate Oxidases in Plants[w

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Hackenberg, Claudia; Kern, Ramona; Hüge, Jan; Stal, Lucas J.; Tsuji, Yoshinori; Kopka, Joachim; Shiraiwa, Yoshihiro; Bauwe, Hermann; Hagemann, Martin

2011-01-01

Glycolate oxidase (GOX) is an essential enzyme involved in photorespiratory metabolism in plants. In cyanobacteria and green algae, the corresponding reaction is catalyzed by glycolate dehydrogenases (GlcD). The genomes of N2-fixing cyanobacteria, such as Nostoc PCC 7120 and green algae, appear to harbor genes for both GlcD and GOX proteins. The GOX-like proteins from Nostoc (No-LOX) and from Chlamydomonas reinhardtii showed high l-lactate oxidase (LOX) and low GOX activities, whereas glycolate was the preferred substrate of the phylogenetically related At-GOX2 from Arabidopsis thaliana. Changing the active site of No-LOX to that of At-GOX2 by site-specific mutagenesis reversed the LOX/GOX activity ratio of No-LOX. Despite its low GOX activity, No-LOX overexpression decreased the accumulation of toxic glycolate in a cyanobacterial photorespiratory mutant and restored its ability to grow in air. A LOX-deficient Nostoc mutant grew normally in nitrate-containing medium but died under N2-fixing conditions. Cultivation under low oxygen rescued this lethal phenotype, indicating that N2 fixation was more sensitive to O2 in the Δlox Nostoc mutant than in the wild type. We propose that LOX primarily serves as an O2-scavenging enzyme to protect nitrogenase in extant N2-fixing cyanobacteria, whereas in plants it has evolved into GOX, responsible for glycolate oxidation during photorespiration. PMID:21828292

Key role of alternative oxidase in lovastatin solid-state fermentation.

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Pérez-Sánchez, Ailed; Uribe-Carvajal, Salvador; Cabrera-Orefice, Alfredo; Barrios-González, Javier

2017-10-01

Lovastatin is a commercially important secondary metabolite produced by Aspergillus terreus, either by solid-state fermentation or by submerged fermentation. In a previous work, we showed that reactive oxygen species (ROS) accumulation in idiophase positively regulates lovastatin biosynthetic genes. In addition, it has been found that lovastatin-specific production decreases with aeration in solid-state fermentation (SSF). To study this phenomenon, we determined ROS accumulation during lovastatin SSF, under high and low aeration conditions. Paradoxically, high aeration caused lower ROS accumulation, and this was the underlying reason of the aeration effect on lovastatin production. Looking for a mechanism that is lowering ROS production under those conditions, we studied alternative respiration. The alternative oxidase provides an alternative route for electrons passing through the electron transport chain to reduce oxygen. Here, we showed that an alternative oxidase (AOX) is expressed in SSF, and only during idiophase. It was shown that higher aeration induces higher alternative respiration (AOX activity), and this is a mechanism that limits ROS generation and keeps them within healthy limits and adequate signaling limits for lovastatin production. Indeed, the aox gene was induced in idiophase, i.e., at the time of ROS accumulation. Moreover, exogenous ROS (H 2 O 2 ), added to lovastatin solid-state fermentation, induced higher AOX activity. This suggests that high O 2 availability in SSF generates dangerously high ROS, so alternative respiration is induced in SSF, indirectly favoring lovastatin production. Conversely, alternative respiration was not detected in lovastatin-submerged fermentation (SmF), although exogenous ROS also induced relatively low AOX activity in SmF.

Engineering glucose oxidase to minimize the influence of oxygen on sensor response

International Nuclear Information System (INIS)

Horaguchi, Yohei; Saito, Shoko; Kojima, Katsuhiro; Tsugawa, Wakako; Ferri, Stefano; Sode, Koji

2014-01-01

Glucose oxidase (GOx) is an important industrial enzyme and is recognized as the gold standard for monitoring blood glucose. However, due to its inherent oxidase property, the presence of oxygen affects electrochemical measurements of venous blood glucose employing artificial electron mediators. We therefore attempted to engineer Penicillium amagasakiense-derived GOx into a dehydrogenase by focusing on the amino acid residues predicted to interact with oxygen. Our rational amino acid substitution approach resulted in the construction of the Ser114Ala/Phe355Leu mutant, which has an 11-fold decrease in oxidase activity and 2.8-fold increase in dehydrogenase activity compared with wild-type GOx. As a result, the dehydrogenase/oxidase activity ratio of the engineered enzyme was 32-fold greater than that of the wild-type enzyme. The enzyme sensor constructed with Ser114Ala/Phe355Leu was considerably less affected by oxygen than the wild-type GOx-based sensor at lower glucose concentrations

Radio-isotopic determination of platelet monoamine oxidase and regulation of its activity by an indigenous drug

International Nuclear Information System (INIS)

Dubey, G.P.; Srivastava, V.K.; Agrawal, A.; Udupa, K.N.

1988-01-01

Platelet monoamine oxidase is a mitochondrial enzyme taking part in the deamination reaction of total catecholamine. Recent studies of monoamine oxidase inhibitors have gained its importance in the control of variety of psychosomatic disorders like mental depression, arterial hypertension and anxiety neurosis. 30 apparently normal individuals and 42 diagnosed cases of essential hypertension were selected for the present study. The platelet monoamine oxidase activity was measured by using 14 C-tryptamine bisuccinate. Comparatively low activity of platelet monoamine oxidase was noticed in hypertension cases than in the normal. After oral administration of an indigenous drug 'Geriforte' for three months, a significant rise in platelet monoamine oxidase activity was noticed in hypertension cases. It can be concluded that this indigenous formulation has the capacity to regulate the monoamine oxidase activity, as such, it may provide an alternative remedy in the management of psychosomatic disorders. (author). 11 refs

The substrate oxidation mechanism of pyranose 2-oxidase and other related enzymes in the glucose-methanol-choline superfamily.

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Wongnate, Thanyaporn; Chaiyen, Pimchai

2013-07-01

Enzymes in the glucose-methanol-choline (GMC) oxidoreductase superfamily catalyze the oxidation of an alcohol moiety to the corresponding aldehyde. In this review, the current understanding of the sugar oxidation mechanism in the reaction of pyranose 2-oxidase (P2O) is highlighted and compared with that of other enzymes in the GMC family for which structural and mechanistic information is available, including glucose oxidase, choline oxidase, cholesterol oxidase, cellobiose dehydrogenase, aryl-alcohol oxidase, and pyridoxine 4-oxidase. Other enzymes in the family that have been newly discovered or for which less information is available are also discussed. A large primary kinetic isotope effect was observed for the flavin reduction when 2-d-D-glucose was used as a substrate, but no solvent kinetic isotope effect was detected for the flavin reduction step. The reaction of P2O is consistent with a hydride transfer mechanism in which there is stepwise formation of d-glucose alkoxide prior to the hydride transfer. Site-directed mutagenesis of P2O and pH-dependence studies indicated that His548 is a catalytic base that facilitates the deprotonation of C2-OH in D-glucose. This finding agrees with the current mechanistic model for aryl-alcohol oxidase, glucose oxidase, cellobiose dehydrogenase, methanol oxidase, and pyridoxine 4-oxidase, but is different from that of cholesterol oxidase and choline oxidase. Although all of the GMC enzymes share similar structural folding and use the hydride transfer mechanism for flavin reduction, they appear to have subtle differences in the fine-tuned details of how they catalyze substrate oxidation. © 2013 The Authors Journal compilation © 2013 FEBS.

The tau positron-emission tomography tracer AV-1451 binds with similar affinities to tau fibrils and monoamine oxidases.

Science.gov (United States)

Vermeiren, Céline; Motte, Philippe; Viot, Delphine; Mairet-Coello, Georges; Courade, Jean-Philippe; Citron, Martin; Mercier, Joël; Hannestad, Jonas; Gillard, Michel

2018-02-01

Lilly/Avid's AV-1451 is one of the most advanced tau PET tracers in the clinic. Although results obtained in Alzheimer's disease patients are compelling, discrimination of tracer uptake in healthy individuals and patients with supranuclear palsy (PSP) is less clear as there is substantial overlap of signal in multiple brain regions. Moreover, accurate quantification of [ 18 F]AV-1451 uptake in Alzheimer's disease may not be possible. The aim of the present study was to characterize the in vitro binding of AV-1451 to understand and identify potential off-target binding that could explain the poor discrimination observed in PSP patients. [ 3 H]AV-1451 and AV-1451 were characterized in in vitro binding assays using recombinant and native proteins/tissues from postmortem samples of controls and Alzheimer's disease and PSP patients. [ 3 H]AV-1451 binds to multiple sites with nanomolar affinities in brain homogenates and to tau fibrils isolated from Alzheimer's disease or PSP patients. [ 3 H]AV-1451 also binds with similarly high affinities in brain homogenates devoid of tau pathology. This unexpected binding was demonstrated to be because of nanomolar affinities of [ 3 H]AV-1451 for monoamine oxidase A and B enzymes. High affinity of AV-1451 for monoamine oxidase proteins may limit its utility as a tau PET tracer in PSP and Alzheimer's disease because of high levels of monoamine oxidase expression in brain regions also affected by tau deposition, especially if monoamine oxidase levels change over time or with a treatment intervention. © 2017 International Parkinson and Movement Disorder Society. © 2017 International Parkinson and Movement Disorder Society.

Direct comparison of gluco-oligosaccharide oxidase variants and glucose oxidase: substrate range and H2O2 stability.

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Vuong, Thu V; Foumani, Maryam; MacCormick, Benjamin; Kwan, Rachel; Master, Emma R

2016-11-21

Glucose oxidase (GO) activity is generally restricted to glucose and is susceptible to inactivation by H 2 O 2 . By comparison, the Y300A variant of gluco-oligosaccharide oxidase (GOOX) from Sarocladium strictum showed broader substrate range and higher H 2 O 2 stability. Specifically, Y300A exhibited up to 40 times higher activity on all tested sugars except glucose, compared to GO. Moreover, fusion of the Y300A variant to a family 22 carbohydrate binding module from Clostridium thermocellum (CtCBM22A) nearly doubled its catalytic efficiency on glucose, while retaining significant activity on oligosaccharides. In the presence of 200 mM of H 2 O 2 , the recombinant CtCBM22A_Y300A retained 80% of activity on glucose and 100% of activity on cellobiose, the preferred substrate for this enzyme. By contrast, a commercial glucose oxidase reported to contain ≤0.1 units catalase/ mg protein, retained 60% activity on glucose under the same conditions. GOOX variants appear to undergo a different mechanism of inactivation, as a loss of histidine instead of methionine was observed after H 2 O 2 incubation. The addition of CtCBM22A also promoted functional binding of the fusion enzyme to xylan, facilitating its simultaneous purification and immobilization using edible oat spelt xylan, which might benefit the usage of this enzyme preparation in food and baking applications.

The antioxidant properties, cytotoxicity and monoamine oxidase ...

African Journals Online (AJOL)

ajl yemi

2011-11-28

Nov 28, 2011 ... Department of Pharmaceutical Chemistry, North-West University, Private Bag X6001, Potchefstroom 2520, ..... on the inhibition of the catabolism of serotonin, .... Structure of human monoamine oxidase B, a drug target for.

Investigation of antihemolytic, xanthine oxidase inhibition ...

African Journals Online (AJOL)

Abbreviations: SVEs: Salvia Verbenaca L. aerial part Extracts; CrE: Crud Extract; ChE: Chloroform Extract ; EAE: Ethyl Acetate Extract; AqE : Aqueous Extract ; ROS: Reactive Oxygen Spices; AAPH : 2,2, -Azobis (2-AmidinoPropane) Dihydrochloride ; DPPH: DiPhenyl- Picryl-Hydrazyl; XO: Xanthine Oxidase; Gen: Gentamicin ...

Gold-Coated M13 Bacteriophage as a Template for Glucose Oxidase Biofuel Cells with Direct Electron Transfer.

Science.gov (United States)

Blaik, Rita A; Lan, Esther; Huang, Yu; Dunn, Bruce

2016-01-26

Glucose oxidase-based biofuel cells are a promising source of alternative energy for small device applications, but still face the challenge of achieving robust electrical contact between the redox enzymes and the current collector. This paper reports on the design of an electrode consisting of glucose oxidase covalently attached to gold nanoparticles that are assembled onto a genetically engineered M13 bacteriophage using EDC-NHS chemistry. The engineered phage is modified at the pIII protein to attach onto a gold substrate and serves as a high-surface-area template. The resulting "nanomesh" architecture exhibits direct electron transfer (DET) and achieves a higher peak current per unit area of 1.2 mA/cm(2) compared to most other DET attachment schemes. The final enzyme surface coverage on the electrode was calculated to be approximately 4.74 × 10(-8) mol/cm(2), which is a significant improvement over most current glucose oxidase (GOx) DET attachment methods.

Biphenyl Modulates the Expression and Function of Respiratory Oxidases in the Polychlorinated-Biphenyls Degrader Pseudomonas pseudoalcaligenes KF707

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Federica Sandri

2017-06-01

Full Text Available Pseudomonas pseudoalcaligenes KF707 is a soil bacterium which is known for its capacity to aerobically degrade harmful organic compounds such as polychlorinated biphenyls (PCBs using biphenyl as co-metabolite. Here we provide the first genetic and functional analysis of the KF707 respiratory terminal oxidases in cells grown with two different carbon sources: glucose and biphenyl. We identified five terminal oxidases in KF707: two c(caa3 type oxidases (Caa3 and Ccaa3, two cbb3 type oxidases (Cbb31 and Cbb32, and one bd type cyanide-insensitive quinol oxidase (CIO. While the activity and expression of both Cbb31 and Cbb32 oxidases was prevalent in glucose grown cells as compared to the other oxidases, the activity and expression of the Caa3 oxidase increased considerably only when biphenyl was used as carbon source in contrast to the Cbb32 oxidase which was repressed. Further, the respiratory activity and expression of CIO was up-regulated in a Cbb31 deletion strain as compared to W.T. whereas the CIO up-regulation was not present in Cbb32 and C(caa3 deletion mutants. These results, together, reveal that both function and expression of cbb3 and caa3 type oxidases in KF707 are modulated by biphenyl which is the co-metabolite needed for the activation of the PCBs-degradation pathway.

Steady-state kinetics of substrate binding and iron release in tomato ACC oxidase.

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Thrower, J S; Blalock, R; Klinman, J P

2001-08-14

1-Aminocyclopropane-1-carboxylate oxidase (ACC oxidase) catalyzes the last step in the biosynthetic pathway of the plant hormone, ethylene. This unusual reaction results in the oxidative ring cleavage of 1-aminocyclopropane carboxylate (ACC) into ethylene, cyanide, and CO2 and requires ferrous ion, ascorbate, and molecular oxygen for catalysis. A new purification procedure and assay method have been developed for tomato ACC oxidase that result in greatly increased enzymatic activity. This method allowed us to determine the rate of iron release from the enzyme and the effect of the activator, CO2, on this rate. Initial velocity studies support an ordered kinetic mechanism where ACC binds first followed by O2; ascorbate can bind after O2 or possibly before ACC. This kinetic mechanism differs from one recently proposed for the ACC oxidase from avocado.

Expression Studies of Gibberellin Oxidases in Developing Pumpkin Seeds1

Science.gov (United States)

Frisse, Andrea; Pimenta, Maria João; Lange, Theo

2003-01-01

Two cDNA clones, 3-ox and 2-ox, have been isolated from developing pumpkin (Cucurbita maxima) embryos that show significant amino acid homology to gibberellin (GA) 3-oxidases and 2-oxidases, respectively. Recombinant fusion protein of clone 3-ox converted GA12-aldehyde, GA12, GA15, GA24, GA25, and GA9 to GA14-aldehyde, GA14, GA37, GA36, GA13, and GA4, respectively. Recombinant 2-ox protein oxidized GA9, GA4, and GA1 to GA51, GA34, and GA8, respectively. Previously cloned GA 7-oxidase revealed additional 3β-hydroxylation activity of GA12. Transcripts of this gene were identified in endosperm and embryo of the developing seed by quantitative reverse transcriptase-polymerase chain reaction and localized in protoderm, root apical meristem, and quiescent center by in situ hybridization. mRNA of the previously cloned GA 20-oxidase from pumpkin seeds was localized in endosperm and in tissues of protoderm, ground meristem, and cotyledons of the embryo. However, transcripts of the recently cloned GA 20-oxidase from pumpkin seedlings were found all over the embryo, and in tissues of the inner seed coat at the micropylar end. Previously cloned GA 2β,3β-hydroxylase mRNA molecules were specifically identified in endosperm tissue. Finally, mRNA molecules of the 3-ox and 2-ox genes were found in the embryo only. 3-ox transcripts were localized in tissues of cotyledons, protoderm, and inner cell layers of the root apical meristem, and 2-ox transcripts were found in all tissues of the embryo except the root tips. These results indicate tissue-specific GA-biosynthetic pathways operating within the developing seed. PMID:12644672

Davallialactone reduces inflammation and repairs dentinogenesis on glucose oxidase-induced stress in dental pulp cells.

Science.gov (United States)

Lee, Young-Hee; Kim, Go-Eun; Song, Yong-Beom; Paudel, Usha; Lee, Nan-Hee; Yun, Bong-Sik; Yu, Mi-Kyung; Yi, Ho-Keun

2013-11-01

The chronic nature of diabetes mellitus (DM) raises the risk of oral complication diseases. In general, DM causes oxidative stress to organs. This study aimed to evaluate the cellular change of dental pulp cells against glucose oxidative stress by glucose oxidase with a high glucose state. The purpose of this study was to test the antioxidant character of davallialactone and to reduce the pathogenesis of dental pulp cells against glucose oxidative stress. The glucose oxidase with a high glucose concentration was tested for hydroxy peroxide (H2O2) production, cellular toxicity, reactive oxygen species (ROS) formation, induction of inflammatory molecules and disturbance of dentin mineralization in human dental pulp cells. The anti-oxidant effect of Davallilactone was investigated to restore dental pulp cells' vitality and dentin mineralization via reduction of H2O2 production, cellular toxicity, ROS formation and inflammatory molecules. The treatment of glucose oxidase with a high glucose concentration increased H2O2 production, cellular toxicity, and inflammatory molecules and disturbed dentin mineralization by reducing pulp cell activity. However, davallialactone reduced H2O2 production, cellular toxicity, ROS formation, inflammatory molecules, and dentin mineralization disturbances even with a long-term glucose oxidative stress state. The results of this study imply that the development of oral complications is related to the irreversible damage of dental pulp cells by DM-induced oxidative stress. Davallialactone, a natural antioxidant, may be useful to treat complicated oral disease, representing an improvement for pulp vital therapy. Copyright © 2013 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Biogenic manganese oxide nanoparticle formation by a multimeric multicopper oxidase Mnx.

Science.gov (United States)

Romano, Christine A; Zhou, Mowei; Song, Yang; Wysocki, Vicki H; Dohnalkova, Alice C; Kovarik, Libor; Paša-Tolić, Ljiljana; Tebo, Bradley M

2017-09-29

Bacteria that produce Mn oxides are extraordinarily skilled engineers of nanomaterials that contribute significantly to global biogeochemical cycles. Their enzyme-based reaction mechanisms may be genetically tailored for environmental remediation applications or bioenergy production. However, significant challenges exist for structural characterization of the enzymes responsible for biomineralization. The active Mn oxidase in Bacillus sp. PL-12, Mnx, is a complex composed of a multicopper oxidase (MCO), MnxG, and two accessory proteins, MnxE and MnxF. MnxG shares sequence similarity with other, structurally characterized MCOs. MnxE and MnxF have no similarity to any characterized proteins. The ~200 kDa complex has been recalcitrant to crystallization, so its structure is unknown. Here, we show that native mass spectrometry defines the subunit topology and copper binding of Mnx, while high-resolution electron microscopy visualizes the protein and nascent Mn oxide minerals. These data provide critical structural information for understanding Mn biomineralization by such unexplored enzymes.Significant challenges exist for structural characterization of enzymes responsible for biomineralization. Here the authors show that native mass spectrometry and high resolution electron microscopy can define the subunit topology and copper binding of a manganese oxidizing complex, and describe early stage formation of its mineral products.

The conserved baculovirus protein p33 (Ac92) is a flavin adenine dinucleotide-linked sulfhydryl oxidase

International Nuclear Information System (INIS)

Long, C.M.; Rohrmann, G.F.; Merrill, G.F.

2009-01-01

Open reading frame 92 of the Autographa californica baculovirus (Ac92) is one of about 30 core genes present in all sequenced baculovirus genomes. Computer analyses predicted that the Ac92 encoded protein (called p33) and several of its baculovirus orthologs were related to a family of flavin adenine dinucleotide (FAD)-linked sulfhydryl oxidases. Alignment of these proteins indicated that, although they were highly diverse, a number of amino acids in common with the Erv1p/Alrp family of sulfhydryl oxidases are present. Some of these conserved amino acids are predicted to stack against the isoalloxazine and adenine components of FAD, whereas others are involved in electron transfer. To investigate this relationship, Ac92 was expressed in bacteria as a His-tagged fusion protein, purified, and characterized both spectrophotometrically and for its enzymatic activity. The purified protein was found to have the color (yellow) and absorption spectrum consistent with it being a FAD-containing protein. Furthermore, it was demonstrated to have sulfhydryl oxidase activity using dithiothreitol and thioredoxin as substrates.

Increased xanthine oxidase during labour--implications for oxidative stress.

Science.gov (United States)

Many, A; Roberts, J M

1997-11-01

Xanthine dehydrogenase/oxidase (XDH/XO) produces uric acid. When in the oxidase form, this production is coupled with the generation of free radicals. Hypoxia-reperfusion enhances conversion of XDH to XO. Since the placenta is exposed to short periods of hypoxia reperfusion during labour, 17 placentae of pregnancy terminated by elective caesarean section and five placentae of pregnancies terminated by caesarean section during labour were examined for XDH/XO activity. It was found that XO activity was higher in the placentae of labouring women (P = 0.003), which suggests that labour enhances conversion of XDH to XO, facilitating free radical production.

Production of a new D-amino acid oxidase from the fungus Fusarium oxysporum.

Science.gov (United States)

Gabler, M; Fischer, L

1999-08-01

The fungus Fusarium oxysporum produced a D-amino acid oxidase (EC 1. 4.3.3) in a medium containing glucose as the carbon and energy source and ammonium sulfate as the nitrogen source. The specific D-amino acid oxidase activity was increased up to 12.5-fold with various D-amino acids or their corresponding derivatives as inducers. The best inducers were D-alanine (2.7 microkat/g of dry biomass) and D-3-aminobutyric acid (2.6 microkat/g of dry biomass). The addition of zinc ions was necessary to permit the induction of peroxisomal D-amino acid oxidase. Bioreactor cultivations were performed on a 50-liter scale, yielding a volumetric D-amino acid oxidase activity of 17 microkat liter(-1) with D-alanine as an inducer. Under oxygen limitation, the volumetric activity was increased threefold to 54 microkat liter(-1) (3,240 U liter(-1)).

Mediation of glycosylated and partially-deglycosylated glucose oxidase of Aspergillus niger by a ferrocene-derivatised detergent.

Science.gov (United States)

Fraser, D M; Zakeeruddin, S M; Grätzel, M

1992-01-30

A ferrocene-derivatised detergent, (11-ferrocenylundecyl) trimethylammonium bromide (FTMAB), when oxidised to the corresponding ferricinium ion, was found by electrochemical studies to be an effective electron acceptor for reduced glucose oxidase of Aspergillus niger (EC 1.13.4) and thus acts as a electron-transfer mediator between glucose oxidase and a working electrode held at a potential sufficiently positive to reoxidise reduced FTMAB. An increase in mediating activity was produced when FTMAB was present in concentrations above its critical micelle concentration. An 'enzyme electrode' was formed by adsorption of glucose oxidase and FTMAB surfactant on a graphite rod. The electrode functioned as an amperometric biosensor for glucose in phosphate-buffered saline solution. A mixed micelle of glucose oxidase and FTMAB, probably adsorbed on the electrode surface, appears to be advantageous for the amperometric determination of glucose. Additionally, glucose oxidase was treated with alpha-mannosidase. When this partially-deglycosylated glucose oxidase was incorporated in an enzyme electrode, a 100-fold increase in the second-order rate constant (k) for electron transfer between the enzyme and FTMAB was observed, together with increased current densities, with respect to the equivalent values for FTMAB and commercial glucose oxidase. The use of deglycosylated enzymes in biosensors is suggested.

Feasibility of converting lactic acid to ethanol in food waste fermentation by immobilized lactate oxidase

International Nuclear Information System (INIS)

Ma, Hong-zhi; Xing, Yi; Yu, Miao; Wang, Qunhui

2014-01-01

Highlights: • Residue lactic acid in food waste could be converted to pyruvic acid. • Calcium alginate immobilized the lactate oxidase with high pH and thermal stability. • Immobilized enzyme could convert 70% lactic acid to pyruvic acid. • Ethanol yield could be increased by 20% with lactate oxidase added. - Abstract: Adoption of lactic acid bacteria (LAB) into ethanol fermentation from food waste can replace the sterilization process. However, LAB inoculation will convert part of the substrate into lactic acid (LA), not ethanol. This study adopted lactate oxidase to convert the produced LA to pyruvate, and then ethanol fermentation was carried out. The immobilization enzyme was utilized, and corresponding optimum conditions were determined. Results showed that calcium alginate could successfully immobilize the enzyme and improve pH and thermal stability. The optimum pH and temperature were 6.2 and 55 °C, respectively. The utilization of immobilized enzyme with catalytic time of 5 h could convert 70% LA to pyruvate, and the addition of enzyme increased the ethanol yield by 20% more than that of the control. The process could be applied in food waste storage and can help in reducing carbon source consumption

Surface Induced Dissociation Coupled with High Resolution Mass Spectrometry Unveils Heterogeneity of a 211 kDa Multicopper Oxidase Protein Complex

Science.gov (United States)

Zhou, Mowei; Yan, Jing; Romano, Christine A.; Tebo, Bradley M.; Wysocki, Vicki H.; Paša-Tolić, Ljiljana

2018-01-01

Manganese oxidation is an important biogeochemical process that is largely regulated by bacteria through enzymatic reactions. However, the detailed mechanism is poorly understood due to challenges in isolating and characterizing these unknown enzymes. A manganese oxidase, Mnx, from Bacillus sp. PL-12 has been successfully overexpressed in active form as a protein complex with a molecular mass of 211 kDa. We have recently used surface induced dissociation (SID) and ion mobility-mass spectrometry (IM-MS) to release and detect folded subcomplexes for determining subunit connectivity and quaternary structure. The data from the native mass spectrometry experiments led to a plausible structural model of this multicopper oxidase, which has been difficult to study by conventional structural biology methods. It was also revealed that each Mnx subunit binds a variable number of copper ions. Becasue of the heterogeneity of the protein and limited mass resolution, ambiguities in assigning some of the observed peaks remained as a barrier to fully understanding the role of metals and potential unknown ligands in Mnx. In this study, we performed SID in a modified Fourier transform-ion cyclotron resonance (FTICR) mass spectrometer. The high mass accuracy and resolution offered by FTICR unveiled unexpected artificial modifications on the protein that had been previously thought to be iron bound species based on lower resolution spectra. Additionally, isotopically resolved spectra of the released subcomplexes revealed the metal binding stoichiometry at different structural levels. This method holds great potential for in-depth characterization of metalloproteins and protein-ligand complexes. [Figure not available: see fulltext.

Cell Wall Amine Oxidases: New Players in Root Xylem Differentiation under Stress Conditions

Directory of Open Access Journals (Sweden)

Sandip A. Ghuge

2015-07-01

Full Text Available Polyamines (PAs are aliphatic polycations present in all living organisms. A growing body of evidence reveals their involvement as regulators in a variety of physiological and pathological events. They are oxidatively deaminated by amine oxidases (AOs, including copper amine oxidases (CuAOs and flavin adenine dinucleotide (FAD-dependent polyamine oxidases (PAOs. The biologically-active hydrogen peroxide (H2O2 is a shared compound in all of the AO-catalyzed reactions, and it has been reported to play important roles in PA-mediated developmental and stress-induced processes. In particular, the AO-driven H2O2 biosynthesis in the cell wall is well known to be involved in plant wound healing and pathogen attack responses by both triggering peroxidase-mediated wall-stiffening events and signaling modulation of defense gene expression. Extensive investigation by a variety of methodological approaches revealed high levels of expression of cell wall-localized AOs in root xylem tissues and vascular parenchyma of different plant species. Here, the recent progresses in understanding the role of cell wall-localized AOs as mediators of root xylem differentiation during development and/or under stress conditions are reviewed. A number of experimental pieces of evidence supports the involvement of apoplastic H2O2 derived from PA oxidation in xylem tissue maturation under stress-simulated conditions.

Paraquat and maneb co-exposure induces noradrenergic locus coeruleus neurodegeneration through NADPH oxidase-mediated microglial activation

International Nuclear Information System (INIS)

Hou, Liyan; Zhang, Cong; Wang, Ke; Liu, Xiaofang; Wang, Hongwei; Che, Yuning; Sun, Fuqiang; Zhou, Xueying; Zhao, Xiulan; Wang, Qingshan

2017-01-01

Highlights: • Microglial activation induced by paraquat and maneb precedes noradrenergic neurodegeneration in locus coeruleus. • NADPH oxidase activation contributes to microglia-mediated neuroinflammation and related noradrenergic neurodegeneration. • Inhibition of NADPH oxidase by apocynin protects noradrenergic neurons against paraquat and maneb-induced toxicity. - Abstract: Co-exposure to paraquat (PQ) and maneb (Mb) has been shown to increase the risk of Parkinson’s disease (PD) and dopaminergic (DA) neurodegeneration in the substantia nigra pars compacta (SNpc) is observed in PQ and Mb-treated experimental animals. The loss of noradrenergic locus coeruleus (LC/NE) neurons in brainstem is a common feature shared by multiple neurodegenerative diseases, including PD. However, whether PQ and Mb is able to damage LC/NE neurons remains undefined. In this study, mice treated with combined PQ and Mb displayed progressive LC/NE neurodegeneration. Time course studies revealed that the activation of microglia preceded LC/NE neurodegeneration. Mechanistically, the activation of NADPH oxidase contributed to microglial activation and subsequent LC/NE neurodegeneration. We found that PQ and Mb co-exposure induced activation of NADPH oxidase as shown by increased superoxide production and membrane translocation of p47 phox , a cytosolic subunit of NADPH oxidase. Inhibition of NADPH oxidase by apocynin, a widely used NADPH oxidase inhibitor, suppressed microglial activation and gene expressions of proinflammatory factors. Furthermore, reduced activation of nuclear factor-κB (NF-κB) pathway was observed in apocynin-treated mice. More importantly, inhibition of NADPH oxidase by apocynin afforded LC/NE neuroprotection against PQ and Mb-induced neurotoxicity. Thus, our findings revealed the critical role NADPH oxidase-mediated microglial activation in driving LC/NE neurodegeneration induced by PQ and Mb, providing new insights into the pathogenesis of environmental

Intramolecular electron transfer in ascorbate oxidase is enhanced in the presence of oxygen

DEFF Research Database (Denmark)

Farver, O; Wherland, S; Pecht, I

1994-01-01

Intramolecular electron transfer from the type 1 copper center to the type 3 copper(II) pair is induced in the multi-copper enzyme, ascorbate oxidase, following pulse radiolytic reduction of the type 1 Cu(II) ion. In the presence of a slight excess of dioxygen over ascorbate oxidase, interaction...... between the trinuclear copper center and O2 is observed even with singly reduced ascorbate oxidase molecules. Under these conditions, the rate constant for intramolecular electron transfer from type 1 Cu(I) to type 3 Cu(II) increases 5-fold to 1100 +/- 300 s-1 (20 degrees C, pH 5.8) as compared...

Reengineered glucose oxidase for amperometric glucose determination in diabetes analytics.

Science.gov (United States)

Arango Gutierrez, Erik; Mundhada, Hemanshu; Meier, Thomas; Duefel, Hartmut; Bocola, Marco; Schwaneberg, Ulrich

2013-12-15

Glucose oxidase is an oxidoreductase exhibiting a high β-D-glucose specificity and high stability which renders glucose oxidase well-suited for applications in diabetes care. Nevertheless, GOx activity is highly oxygen dependent which can lead to inaccuracies in amperometric β-D-glucose determinations. Therefore a directed evolution campaign with two rounds of random mutagenesis (SeSaM followed by epPCR), site saturation mutagenesis studies on individual positions, and one simultaneous site saturation library (OmniChange; 4 positions) was performed. A diabetes care well suited mediator (quinone diimine) was selected and the GOx variant (T30V I94V) served as starting point. For directed GOx evolution a microtiter plate detection system based on the quinone diimine mediator was developed and the well-known ABTS-assay was applied in microtiter plate format to validate oxygen independency of improved GOx variants. Two iterative rounds of random diversity generation and screening yielded to two subsets of amino acid positions which mainly improved activity (A173, A332) and oxygen independency (F414, V560). Simultaneous site saturation of all four positions with a reduced subset of amino acids using the OmniChange method yielded finally variant V7 with a 37-fold decreased oxygen dependency (mediator activity: 7.4 U/mg WT, 47.5 U/mg V7; oxygen activity: 172.3 U/mg WT, 30.1 U/mg V7). V7 is still highly β-D-glucose specific, highly active with the quinone diimine mediator and thermal resistance is retained (prerequisite for GOx coating of diabetes test stripes). The latter properties and V7's oxygen insensitivity make V7 a very promising candidate to replace standard GOx in diabetes care applications. Copyright © 2013 Elsevier B.V. All rights reserved.

Characterization of a Flavoprotein Oxidase from Opium Poppy Catalyzing the Final Steps in Sanguinarine and Papaverine Biosynthesis*

Science.gov (United States)

Hagel, Jillian M.; Beaudoin, Guillaume A. W.; Fossati, Elena; Ekins, Andrew; Martin, Vincent J. J.; Facchini, Peter J.

2012-01-01

Benzylisoquinoline alkaloids are a diverse class of plant specialized metabolites that includes the analgesic morphine, the antimicrobials sanguinarine and berberine, and the vasodilator papaverine. The two-electron oxidation of dihydrosanguinarine catalyzed by dihydrobenzophenanthridine oxidase (DBOX) is the final step in sanguinarine biosynthesis. The formation of the fully conjugated ring system in sanguinarine is similar to the four-electron oxidations of (S)-canadine to berberine and (S)-tetrahydropapaverine to papaverine. We report the isolation and functional characterization of an opium poppy (Papaver somniferum) cDNA encoding DBOX, a flavoprotein oxidase with homology to (S)-tetrahydroprotoberberine oxidase and the berberine bridge enzyme. A query of translated opium poppy stem transcriptome databases using berberine bridge enzyme yielded several candidate genes, including an (S)-tetrahydroprotoberberine oxidase-like sequence selected for heterologous expression in Pichia pastoris. The recombinant enzyme preferentially catalyzed the oxidation of dihydrosanguinarine to sanguinarine but also converted (RS)-tetrahydropapaverine to papaverine and several protoberberine alkaloids to oxidized forms, including (RS)-canadine to berberine. The Km values of 201 and 146 μm for dihydrosanguinarine and the protoberberine alkaloid (S)-scoulerine, respectively, suggested high concentrations of these substrates in the plant. Virus-induced gene silencing to reduce DBOX transcript levels resulted in a corresponding reduction in sanguinarine, dihydrosanguinarine, and papaverine accumulation in opium poppy roots in support of DBOX as a multifunctional oxidative enzyme in BIA metabolism. PMID:23118227

Characterization of a flavoprotein oxidase from opium poppy catalyzing the final steps in sanguinarine and papaverine biosynthesis.

Science.gov (United States)

Hagel, Jillian M; Beaudoin, Guillaume A W; Fossati, Elena; Ekins, Andrew; Martin, Vincent J J; Facchini, Peter J

2012-12-14

Benzylisoquinoline alkaloids are a diverse class of plant specialized metabolites that includes the analgesic morphine, the antimicrobials sanguinarine and berberine, and the vasodilator papaverine. The two-electron oxidation of dihydrosanguinarine catalyzed by dihydrobenzophenanthridine oxidase (DBOX) is the final step in sanguinarine biosynthesis. The formation of the fully conjugated ring system in sanguinarine is similar to the four-electron oxidations of (S)-canadine to berberine and (S)-tetrahydropapaverine to papaverine. We report the isolation and functional characterization of an opium poppy (Papaver somniferum) cDNA encoding DBOX, a flavoprotein oxidase with homology to (S)-tetrahydroprotoberberine oxidase and the berberine bridge enzyme. A query of translated opium poppy stem transcriptome databases using berberine bridge enzyme yielded several candidate genes, including an (S)-tetrahydroprotoberberine oxidase-like sequence selected for heterologous expression in Pichia pastoris. The recombinant enzyme preferentially catalyzed the oxidation of dihydrosanguinarine to sanguinarine but also converted (RS)-tetrahydropapaverine to papaverine and several protoberberine alkaloids to oxidized forms, including (RS)-canadine to berberine. The K(m) values of 201 and 146 μm for dihydrosanguinarine and the protoberberine alkaloid (S)-scoulerine, respectively, suggested high concentrations of these substrates in the plant. Virus-induced gene silencing to reduce DBOX transcript levels resulted in a corresponding reduction in sanguinarine, dihydrosanguinarine, and papaverine accumulation in opium poppy roots in support of DBOX as a multifunctional oxidative enzyme in BIA metabolism.

Ratiometric glucose sensing based on fluorescent oxygen films and glucose oxidase

Directory of Open Access Journals (Sweden)

Fengyu Su

2017-06-01

Full Text Available A new two-layer sensor film was constructed for sensing glucose based on glucose oxidase and oxygen sensing material. The first layer of film containing the oxygen sensor and intra-reference material was polymerized, then the second layer of glucose oxidase and glutaraldehyde was formed on the oxygen sensor layer. The two-layer sensor film has a resolution up to 0.05 mM and a detection range from 0 to 5 mM to glucose. The effects of pH and temperature on the sensing performance were systematically investigated. The selective detection of glucose among other monosaccharides, such as fructose, mannose and galactose indicated that the sensing film has excellent selectivity. The prepared sensor was successfully applied for glucose sample detection of glucose concentration in artificial tears. Keywords: Glucose sensor, Glucose oxidase, Fluorescence, Oxygen film, Diabetes

Glucose oxidase-functionalized fluorescent gold nanoclusters as probes for glucose

International Nuclear Information System (INIS)

Xia, Xiaodong; Long, Yunfei; Wang, Jianxiu

2013-01-01

Highlights: ► A glucose oxidase/gold nanocluster conjugates formed by etching chemistry. ► Integration of the bioactivities and fluorescence properties within a single unit. ► These conjugates serve as novel fluorescent probe for glucose. -- Abstract: Creation and application of noble metal nanoclusters have received continuous attention. By integrating enzyme activity and fluorescence for potential applications, enzyme-capped metal clusters are more desirable. This work demonstrated a glucose oxidase (an enzyme for glucose)-functionalized gold cluster as probe for glucose. Under physiological conditions, such bioconjugate was successfully prepared by an etching reaction, where tetrakis (hydroxylmethyl) phosphonium-protected gold nanoparticle and thioctic acid-modified glucose oxidase were used as precursor and etchant, respectively. These bioconjugates showed unique fluorescence spectra (λ em max = 650 nm, λ ex max = 507 nm) with an acceptable quantum yield (ca. 7%). Moreover, the conjugated glucose oxidase remained active and catalyzed reaction of glucose and dissolved O 2 to produce H 2 O 2 , which quenched quantitatively the fluorescence of gold clusters and laid a foundation of glucose detection. A linear range of 2.0 × 10 −6 –140 × 10 −6 M and a detection limit of 0.7 × 10 −6 M (S/N = 3) were obtained. Also, another horseradish peroxidase/gold cluster bioconjugate was produced by such general synthesis method. Such enzyme/metal cluster bioconjugates represented a promising class of biosensors for biologically important targets in organelles or cells

Ultrafine carbon particles promote rotenone-induced dopamine neuronal loss through activating microglial NADPH oxidase

Energy Technology Data Exchange (ETDEWEB)

Wang, Yinxi; Liu, Dan; Zhang, Huifeng; Wang, Yixin [Department of Occupational and Environmental Health Sciences, School of Public Health, Peking University, 100191 (China); Wei, Ling [Beijing Center for Physical & Chemical Analysis, Beijing 100089 (China); Liu, Yutong [School of Life Science, Beijing Normal University, Beijing 100875 (China); Liao, Jieying [Department of Translational Medicine, Xiamen Institute of Rare Earth Materials, Chinese Academy of Sciences, Xiamen 361024 (China); Gao, Hui-Ming [Model Animal Research Center of Nanjing University, Nanjing 211800 (China); Zhou, Hui, E-mail: hardhui@gmail.com [Department of Occupational and Environmental Health Sciences, School of Public Health, Peking University, 100191 (China)

2017-05-01

Background: Atmospheric ultrafine particles (UFPs) and pesticide rotenone were considered as potential environmental risk factors for Parkinson's disease (PD). However, whether and how UFPs alone and in combination with rotenone affect the pathogenesis of PD remains largely unknown. Methods: Ultrafine carbon black (ufCB, a surrogate of UFPs) and rotenone were used individually or in combination to determine their roles in chronic dopaminergic (DA) loss in neuron-glia, and neuron-enriched, mix-glia cultures. Immunochemistry using antibody against tyrosine hydroxylase was performed to detect DA neuronal loss. Measurement of extracellular superoxide and intracellular reactive oxygen species (ROS) were performed to examine activation of NADPH oxidase. Genetic deletion and pharmacological inhibition of NADPH oxidase and MAC-1 receptor in microglia were employed to examine their role in DA neuronal loss triggered by ufCB and rotenone. Results: In rodent midbrain neuron-glia cultures, ufCB and rotenone alone caused neuronal death in a dose-dependent manner. In particularly, ufCB at doses of 50 and 100 μg/cm{sup 2} induced significant loss of DA neurons. More importantly, nontoxic doses of ufCB (10 μg/cm{sup 2}) and rotenone (2 nM) induced synergistic toxicity to DA neurons. Microglial activation was essential in this process. Furthermore, superoxide production from microglial NADPH oxidase was critical in ufCB/rotenone-induced neurotoxicity. Studies in mix-glia cultures showed that ufCB treatment activated microglial NADPH oxidase to induce superoxide production. Firstly, ufCB enhanced the expression of NADPH oxidase subunits (gp91{sup phox}, p47{sup phox} and p40{sup phox}); secondly, ufCB was recognized by microglial surface MAC-1 receptor and consequently promoted rotenone-induced p47{sup phox} and p67{sup phox} translocation assembling active NADPH oxidase. Conclusion: ufCB and rotenone worked in synergy to activate NADPH oxidase in microglia, leading to

Ultrafine carbon particles promote rotenone-induced dopamine neuronal loss through activating microglial NADPH oxidase

International Nuclear Information System (INIS)

Wang, Yinxi; Liu, Dan; Zhang, Huifeng; Wang, Yixin; Wei, Ling; Liu, Yutong; Liao, Jieying; Gao, Hui-Ming; Zhou, Hui

2017-01-01

Background: Atmospheric ultrafine particles (UFPs) and pesticide rotenone were considered as potential environmental risk factors for Parkinson's disease (PD). However, whether and how UFPs alone and in combination with rotenone affect the pathogenesis of PD remains largely unknown. Methods: Ultrafine carbon black (ufCB, a surrogate of UFPs) and rotenone were used individually or in combination to determine their roles in chronic dopaminergic (DA) loss in neuron-glia, and neuron-enriched, mix-glia cultures. Immunochemistry using antibody against tyrosine hydroxylase was performed to detect DA neuronal loss. Measurement of extracellular superoxide and intracellular reactive oxygen species (ROS) were performed to examine activation of NADPH oxidase. Genetic deletion and pharmacological inhibition of NADPH oxidase and MAC-1 receptor in microglia were employed to examine their role in DA neuronal loss triggered by ufCB and rotenone. Results: In rodent midbrain neuron-glia cultures, ufCB and rotenone alone caused neuronal death in a dose-dependent manner. In particularly, ufCB at doses of 50 and 100 μg/cm 2 induced significant loss of DA neurons. More importantly, nontoxic doses of ufCB (10 μg/cm 2 ) and rotenone (2 nM) induced synergistic toxicity to DA neurons. Microglial activation was essential in this process. Furthermore, superoxide production from microglial NADPH oxidase was critical in ufCB/rotenone-induced neurotoxicity. Studies in mix-glia cultures showed that ufCB treatment activated microglial NADPH oxidase to induce superoxide production. Firstly, ufCB enhanced the expression of NADPH oxidase subunits (gp91 phox , p47 phox and p40 phox ); secondly, ufCB was recognized by microglial surface MAC-1 receptor and consequently promoted rotenone-induced p47 phox and p67 phox translocation assembling active NADPH oxidase. Conclusion: ufCB and rotenone worked in synergy to activate NADPH oxidase in microglia, leading to oxidative damage to DA neurons. Our

Analysis of cellulase and polyphenol oxidase production by southern pine beetle associated fungi

Science.gov (United States)

Abduvali Valiev; Zumrut B. Ogel; Dier D. Klepzig

2009-01-01

In this study, the production of extracellular enzymes by fungi associated with southern pine beetle was investigated for the first time. Cellulase and polyphenol oxidase production were analyzed for three beetle associated fungi. Only the mutualistic symbiont Entomocorticium sp. A was found to produce cellulases and polyphenol oxidase....

Ozone affects pollen viability and NAD(P)H oxidase release from Ambrosia artemisiifolia pollen

International Nuclear Information System (INIS)

Pasqualini, Stefania; Tedeschini, Emma; Frenguelli, Giuseppe; Wopfner, Nicole; Ferreira, Fatima; D'Amato, Gennaro; Ederli, Luisa

2011-01-01

Air pollution is frequently proposed as a cause of the increased incidence of allergy in industrialised countries. We investigated the impact of ozone (O 3 ) on reactive oxygen species (ROS) and allergen content of ragweed pollen (Ambrosia artemisiifolia). Pollen was exposed to acute O 3 fumigation, with analysis of pollen viability, ROS and nitric oxide (NO) content, activity of nicotinamide adenine dinucleotide phosphate (NAD[P]H) oxidase, and expression of major allergens. There was decreased pollen viability after O 3 fumigation, which indicates damage to the pollen membrane system, although the ROS and NO contents were not changed or were only slightly induced, respectively. Ozone exposure induced a significant enhancement of the ROS-generating enzyme NAD(P)H oxidase. The expression of the allergen Amb a 1 was not affected by O 3 , determined from the mRNA levels of the major allergens. We conclude that O 3 can increase ragweed pollen allergenicity through stimulation of ROS-generating NAD(P)H oxidase. - Highlights: → O 3 reduces the viability of ragweed pollen. → ROS and allergens of ragweed pollen were not affected by O 3 exposure. → O 3 enhances the activity of the ROS-generating enzyme NAD(P)H oxidase. → O 3 increases ragweed pollen allergenicity through NAD(P)H-oxidase stimulation. - This study focuses on the effects of the atmospheric pollutant ozone on ROS content and NAD(P)H oxidase activity of ragweed pollen grains.

Ozone affects pollen viability and NAD(P)H oxidase release from Ambrosia artemisiifolia pollen

Energy Technology Data Exchange (ETDEWEB)

Pasqualini, Stefania, E-mail: spas@unipg.it [Department of Applied Biology, University of Perugia, Perugia (Italy); Tedeschini, Emma; Frenguelli, Giuseppe [Department of Applied Biology, University of Perugia, Perugia (Italy); Wopfner, Nicole; Ferreira, Fatima [Department of Molecular Biology, CD Laboratory for Allergy Diagnosis and Therapy, University of Salzburg, Salzburg (Austria); D' Amato, Gennaro [Division of Respiratory and Allergic Diseases, ' A. Cardarelli' High Speciality Hospital, Naples (Italy); Ederli, Luisa [Department of Applied Biology, University of Perugia, Perugia (Italy)

2011-10-15

Air pollution is frequently proposed as a cause of the increased incidence of allergy in industrialised countries. We investigated the impact of ozone (O{sub 3}) on reactive oxygen species (ROS) and allergen content of ragweed pollen (Ambrosia artemisiifolia). Pollen was exposed to acute O{sub 3} fumigation, with analysis of pollen viability, ROS and nitric oxide (NO) content, activity of nicotinamide adenine dinucleotide phosphate (NAD[P]H) oxidase, and expression of major allergens. There was decreased pollen viability after O{sub 3} fumigation, which indicates damage to the pollen membrane system, although the ROS and NO contents were not changed or were only slightly induced, respectively. Ozone exposure induced a significant enhancement of the ROS-generating enzyme NAD(P)H oxidase. The expression of the allergen Amb a 1 was not affected by O{sub 3}, determined from the mRNA levels of the major allergens. We conclude that O{sub 3} can increase ragweed pollen allergenicity through stimulation of ROS-generating NAD(P)H oxidase. - Highlights: > O{sub 3} reduces the viability of ragweed pollen. > ROS and allergens of ragweed pollen were not affected by O{sub 3} exposure. > O{sub 3} enhances the activity of the ROS-generating enzyme NAD(P)H oxidase. > O{sub 3} increases ragweed pollen allergenicity through NAD(P)H-oxidase stimulation. - This study focuses on the effects of the atmospheric pollutant ozone on ROS content and NAD(P)H oxidase activity of ragweed pollen grains.

Adaptive evolution of cytochrome c oxidase: Infrastructure for a carnivorous plant radiation

OpenAIRE

Jobson, Richard W.; Nielsen, Rasmus; Laakkonen, Liisa; Wikström, Mårten; Albert, Victor A.

2004-01-01

Much recent attention in the study of adaptation of organismal form has centered on developmental regulation. As such, the highly conserved respiratory machinery of eukaryotic cells might seem an unlikely target for selection supporting novel morphologies. We demonstrate that a dramatic molecular evolutionary rate increase in subunit I of cytochrome c oxidase (COX) from an active-trapping lineage of carnivorous plants is caused by positive Darwinian selection. Bladderworts (Utricularia) trap ...

Fabrication of Cu2 O-based Materials for Lithium-Ion Batteries.

Science.gov (United States)

Zhang, Li; Li, Qinyuan; Xue, Huaiguo; Pang, Huan

2018-05-25

The improvement of the performance of advanced batteries has played a key role in the energy research community since its inception. Therefore, it is necessary to explore high-performance materials for applications in advanced batteries. Among the variety of materials applied in batteries, much research has been dedicated to examine cuprous oxide materials as working electrodes in lithium cells to check their suitability as anodes for Li-ion cells and this has revealed great working capacities because of their specific characteristics (polymorphic forms, controllable structure, high cycling capacity, etc.). Thus, cuprous oxide and its composites will be fully introduced in this Review for their applications in advanced batteries. It is believed that, in the future, both the study and the impact of cuprous oxide and its composites will be much more profound and lasting. © 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.

A broad distribution of the alternative oxidase in microsporidian parasites.

Directory of Open Access Journals (Sweden)

Bryony A P Williams

2010-02-01

Full Text Available Microsporidia are a group of obligate intracellular parasitic eukaryotes that were considered to be amitochondriate until the recent discovery of highly reduced mitochondrial organelles called mitosomes. Analysis of the complete genome of Encephalitozoon cuniculi revealed a highly reduced set of proteins in the organelle, mostly related to the assembly of iron-sulphur clusters. Oxidative phosphorylation and the Krebs cycle proteins were absent, in keeping with the notion that the microsporidia and their mitosomes are anaerobic, as is the case for other mitosome bearing eukaryotes, such as Giardia. Here we provide evidence opening the possibility that mitosomes in a number of microsporidian lineages are not completely anaerobic. Specifically, we have identified and characterized a gene encoding the alternative oxidase (AOX, a typically mitochondrial terminal oxidase in eukaryotes, in the genomes of several distantly related microsporidian species, even though this gene is absent from the complete genome of E. cuniculi. In order to confirm that these genes encode functional proteins, AOX genes from both A. locustae and T. hominis were over-expressed in E. coli and AOX activity measured spectrophotometrically using ubiquinol-1 (UQ-1 as substrate. Both A. locustae and T. hominis AOX proteins reduced UQ-1 in a cyanide and antimycin-resistant manner that was sensitive to ascofuranone, a potent inhibitor of the trypanosomal AOX. The physiological role of AOX microsporidia may be to reoxidise reducing equivalents produced by glycolysis, in a manner comparable to that observed in trypanosomes.

A novel application of the CuI thin film for preparing thin copper nanowires

International Nuclear Information System (INIS)

Shi Shuo; Sun Jialin; Zhang Jianhong; Cao Yang

2005-01-01

We present a novel application of the CuI thin film for preparing thin copper nanowires under a direct current electric field (DCEF). The CuI thin film was used as a medium for transmitting cuprous ions during the growing process of copper nanowires. As electrodes are the source of cuprous ions, high-purity copper films were deposited on both ends of the CuI thin film. At 353 K, under whole solid condition, without any templates, and having applied a DCEF of 1.5x10 4 V/m, cuprous ions were generated at the anode and migrated towards the cathode through the CuI film. At the edge of the cathode, cuprous ions obtained electrons and congregated to form a disordered thin copper nanowires bundle. The SEM images showed that these copper nanowires were from 10 to 20 nm in diameter and several hundred nanometers in length. The effect of the electric field intensity and the growth temperature on the diameter of the nanowires was also studied

Integration of a highly ordered gold nanowires array with glucose oxidase for ultra-sensitive glucose detection

Energy Technology Data Exchange (ETDEWEB)

Cui, Jiewu [NanoScience and Sensor Technology Research Group, School of Applied Sciences and Engineering, Monash University, Gippsland Campus, Churchill 3842, VIC Australia (Australia); Laboratory of Functional Nanomaterials and Devices, School of Materials Science and Engineering, Hefei University of Technology, Hefei 230009, Anhui (China); Adeloju, Samuel B., E-mail: sam.adeloju@monash.edu [NanoScience and Sensor Technology Research Group, School of Applied Sciences and Engineering, Monash University, Gippsland Campus, Churchill 3842, VIC Australia (Australia); Wu, Yucheng, E-mail: ycwu@hfut.edu.cn [Laboratory of Functional Nanomaterials and Devices, School of Materials Science and Engineering, Hefei University of Technology, Hefei 230009, Anhui (China)

2014-01-27

Graphical abstract: -- Highlights: •Successfully synthesised highly-ordered gold nanowires array with an AAO template. •Fabricated an ultra-sensitive glucose nanobiosensor with the gold nanowires array. •Achieved sensitivity as high as 379.0 μA cm{sup −2} mM{sup −1} and detection limit as low as 50 nM. •Achieved excellent anti-interference with aid of Nafion membrane towards UA and AA. •Enabled successful detection and quantification of glucose in human blood serum. -- Abstract: A highly sensitive amperometric nanobiosensor has been developed by integration of glucose oxidase (GO{sub x}) with a gold nanowires array (AuNWA) by cross-linking with a mixture of glutaraldehyde (GLA) and bovine serum albumin (BSA). An initial investigation of the morphology of the synthesized AuNWA by field emission scanning electron microscopy (FESEM) and field emission transmission electron microscopy (FETEM) revealed that the nanowires array was highly ordered with rough surface, and the electrochemical features of the AuNWA with/without modification were also investigated. The integrated AuNWA–BSA–GLA–GO{sub x} nanobiosensor with Nafion membrane gave a very high sensitivity of 298.2 μA cm{sup −2} mM{sup −1} for amperometric detection of glucose, while also achieving a low detection limit of 0.1 μM, and a wide linear range of 5–6000 μM. Furthermore, the nanobiosensor exhibited excellent anti-interference ability towards uric acid (UA) and ascorbic acid (AA) with the aid of Nafion membrane, and the results obtained for the analysis of human blood serum indicated that the device is capable of glucose detection in real samples.

A decade of crystallization drops: crystallization of the cbb3 cytochrome c oxidase from Pseudomonas stutzeri.

Science.gov (United States)

Buschmann, Sabine; Richers, Sebastian; Ermler, Ulrich; Michel, Hartmut

2014-04-01

The cbb3 cytochrome c oxidases are distant members of the superfamily of heme copper oxidases. These terminal oxidases couple O2 reduction with proton transport across the plasma membrane and, as a part of the respiratory chain, contribute to the generation of an electrochemical proton gradient. Compared with other structurally characterized members of the heme copper oxidases, the recently determined cbb3 oxidase structure at 3.2 Å resolution revealed significant differences in the electron supply system, the proton conducting pathways and the coupling of O2 reduction to proton translocation. In this paper, we present a detailed report on the key steps for structure determination. Improvement of the protein quality was achieved by optimization of the number of lipids attached to the protein as well as the separation of two cbb3 oxidase isoenzymes. The exchange of n-dodecyl-β-D-maltoside for a precisely defined mixture of two α-maltosides and decanoylsucrose as well as the choice of the crystallization method had a most profound impact on crystal quality. This report highlights problems frequently encountered in membrane protein crystallization and offers meaningful approaches to improve crystal quality. © 2014 The Protein Society.

Age-related ultrastructural and monoamine oxidase changes in the rat optic nerve.

Science.gov (United States)

Taurone, S; Ripandelli, G; Minni, A; Lattanzi, R; Miglietta, S; Pepe, N; Fumagalli, L; Micera, A; Pastore, F S; Artico, M

2016-01-01

The aim of this paper is to study the morphology and the distribution of the monoamine oxidase enzymatic system in the optic nerve of 4 month-old Wistar (young) and 28 month-old Wistar (old) rats. The optic nerve was harvested from 20 young and old rats. The segment of optic nerve was divided longitudinally into two pieces, each 0.1 mm in length. The first piece was used for transmission electron microscopy. The second piece was stained with histochemical reaction for monoamine oxidase. The agerelated changes in the optic nerve of rats include micro-anatomical details, ultrastructure and monoamine oxidase histochemical staining. A strong decrease of the thin nerve fibers and a swelling of the thick ones can be observed in optic nerve fibers of old rats. Increased monoamine oxidase histochemical staining of the optic nerve of aged rats is well demonstrated. The increase of meningeal shealth and the decrease of thin nerve fibers of the optic nerve in old rats are well documented. Morphological, ultrastructural and histochemical changes observed in optic nerve fibers of the old rats show a close relation with aging.

The antioxidant properties, cytotoxicity and monoamine oxidase ...

African Journals Online (AJOL)

Tarchonanthus camphoratus (camphor bush) has been widely used for numerous medicinal purposes. The aim of the present study was to evaluate the antioxidant properties, cytotoxicity and monoamine oxidase inhibition activities of the crude dichloromethane leaf extract of T. camphoratus. The antioxidant activities were ...

Glycosylation site-targeted PEGylation of glucose oxidase retains native enzymatic activity.

Science.gov (United States)

Ritter, Dustin W; Roberts, Jason R; McShane, Michael J

2013-04-10

Targeted PEGylation of glucose oxidase at its glycosylation sites was investigated to determine the effect on enzymatic activity, as well as the bioconjugate's potential in an optical biosensing assay. Methoxy-poly(ethylene glycol)-hydrazide (4.5kDa) was covalently coupled to periodate-oxidized glycosylation sites of glucose oxidase from Aspergillus niger. The bioconjugate was characterized using gel electrophoresis, liquid chromatography, mass spectrometry, and dynamic light scattering. Gel electrophoresis data showed that the PEGylation protocol resulted in a drastic increase (ca. 100kDa) in the apparent molecular mass of the protein subunit, with complete conversion to the bioconjugate; liquid chromatography data corroborated this large increase in molecular size. Mass spectrometry data proved that the extent of PEGylation was six poly(ethylene glycol) chains per glucose oxidase dimer. Dynamic light scattering data indicated the absence of higher-order oligomers in the PEGylated GOx sample. To assess stability, enzymatic activity assays were performed in triplicate at multiple time points over the course of 29 days in the absence of glucose, as well as before and after exposure to 5% w/v glucose for 24h. At a confidence level of 95%, the bioconjugate's performance was statistically equivalent to native glucose oxidase in terms of activity retention over the 29 day time period, as well as following the 24h glucose exposure. Finally, the bioconjugate was entrapped within a poly(2-hydroxyethyl methacrylate) hydrogel containing an oxygen-sensitive phosphor, and the construct was shown to respond approximately linearly with a 220±73% signal change (n=4, 95% confidence interval) over the physiologically-relevant glucose range (i.e., 0-400mg/dL); to our knowledge, this represents the first demonstration of PEGylated glucose oxidase incorporated into an optical biosensing assay. Copyright © 2013 Elsevier Inc. All rights reserved.

Gene cloning and heterologous expression of pyranose 2-oxidase from the brown-rot fungus, Gloeophyllum trabeum

Science.gov (United States)

Diane Dietrich; Casey Crooks

2009-01-01

A pyranose 2-oxidase gene from the brown-rot basidiomycete Gloeophyllum trabeum was isolated using homology-based degenerate PCR. The gene structure was determined and compared to that of several pyranose 2-oxidases cloned from white-rot fungi. The G. trabeum pyranose 2-oxidase gene consists of 16 coding exons with canonical promoter CAAT and TATA elements in the 5’UTR...

Cyanobacterial lactate oxidases serve as essential partners in N2-fixation and evolved into photorespiratory glycolate oxidases in plants.

NARCIS (Netherlands)

Hackenberg, C.; Kern, R.; Hüge, J; Stal, L.J.; Tsuji, Y.; Kopka, J.; Shiraiwa, Y.; Bauwe, H.; Hagemann, M.

2011-01-01

Glycolate oxidase (GOX) is an essential enzyme involved in photorespiratory metabolism in plants. In cyanobacteria and green algae, the corresponding reaction is catalyzed by glycolate dehydrogenases (GlcD). The genomes of N2-fixing cyanobacteria, such as Nostoc PCC 7120 and green algae, appear to

Cyanobacterial lactate oxidases serve as essential partners of N2-fixation and evolved to photorespiratory glycolate oxidases in plants

NARCIS (Netherlands)

Hackenberg, C.; Kern, R.; Hüge, J.; Stal, L.J.; Tsuji, Y.; Kopka, J.; Shiraiwa, Y.; Bauwe, H.; Hagemann, M.

2011-01-01

Glycolate oxidase (GOX) is an essential enzyme involved in photorespiratory metabolism in plants. In cyanobacteria and green algae, the corresponding reaction is catalyzed by glycolate dehydrogenases (GlcD). The genomes of N2-fixing cyanobacteria, such as Nostoc PCC 7120 and green algae, appear to

Oxoferryl-Porphyrin Radical Catalytic Intermediate in Cytochrome bd Oxidases Protects Cells from Formation of Reactive Oxygen Species*

Science.gov (United States)

Paulus, Angela; Rossius, Sebastiaan Gijsbertus Hendrik; Dijk, Madelon; de Vries, Simon

2012-01-01

The quinol-linked cytochrome bd oxidases are terminal oxidases in respiration. These oxidases harbor a low spin heme b558 that donates electrons to a binuclear heme b595/heme d center. The reaction with O2 and subsequent catalytic steps of the Escherichia coli cytochrome bd-I oxidase were investigated by means of ultra-fast freeze-quench trapping followed by EPR and UV-visible spectroscopy. After the initial binding of O2, the O–O bond is heterolytically cleaved to yield a kinetically competent heme d oxoferryl porphyrin π-cation radical intermediate (compound I) magnetically interacting with heme b595. Compound I accumulates to 0.75–0.85 per enzyme in agreement with its much higher rate of formation (∼20,000 s−1) compared with its rate of decay (∼1,900 s−1). Compound I is next converted to a short lived heme d oxoferryl intermediate (compound II) in a phase kinetically matched to the oxidation of heme b558 before completion of the reaction. The results indicate that cytochrome bd oxidases like the heme-copper oxidases break the O–O bond in a single four-electron transfer without a peroxide intermediate. However, in cytochrome bd oxidases, the fourth electron is donated by the porphyrin moiety rather than by a nearby amino acid. The production of reactive oxygen species by the cytochrome bd oxidase was below the detection level of 1 per 1000 turnovers. We propose that the two classes of terminal oxidases have mechanistically converged to enzymes in which the O–O bond is broken in a single four-electron transfer reaction to safeguard the cell from the formation of reactive oxygen species. PMID:22287551

Peroxidasin-mediated crosslinking of collagen IV is independent of NADPH oxidases

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Gábor Sirokmány

2018-06-01

Full Text Available Collagen IV is a major component of the basement membrane in epithelial tissues. The NC1 domains of collagen IV protomers are covalently linked together through sulfilimine bonds, the formation of which is catalyzed by peroxidasin. Although hydrogen peroxide is essential for this reaction, the exact source of the oxidant remains elusive. Members of the NOX/DUOX NADPH oxidase family are specifically devoted to the production of superoxide and hydrogen peroxide. Our aim in this study was to find out if NADPH oxidases contribute in vivo to the formation of collagen IV sulfilimine crosslinks. We used multiple genetically modified in vivo model systems to provide a detailed assessment of this question. Our data indicate that in various peroxidasin-expressing tissues sulfilimine crosslinks between the NC1 domains of collagen IV can be readily detected in the absence of functioning NADPH oxidases. We also analyzed how subatmospheric oxygen levels influence the collagen IV network in collagen-producing cultured cells with rapid matrix turnover. We showed that collagen IV crosslinks remain intact even under strongly hypoxic conditions. Our hypothesis is that during collagen IV network formation PXDN cooperates with a NOX/DUOX-independent H2O2 source that is functional also at very low ambient oxygen levels. Keywords: Peroxidasin, NADPH oxidase, Hydrogen peroxide, Collagen IV, Sulfilimine

Biological Effects of Potato Plants Transformation with Glucose Oxidase Gene and their Resistance to Hyperthermia

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O.I. Grabelnych

2017-02-01

Full Text Available It is known that regulation of plant tolerance to adverse environmental factors is connected with short term increase of the concentration of endogenous reactive oxygen species (ROS, which are signalling molecules for the induction of protective mechanisms. Introduction and expression of heterologous gox gene, which encodes glucose oxidase enzyme in plant genome, induce constantly higher content of hydrogen peroxide in plant tissues. It is not known how the introduction of native or modified gox gene affects the plant resistance to high-temperature stress, one of the most commonly used model for the study of stress response and thermal tolerance. In this study, we investigated biological effects of transformation and evaluated the resistance to temperature stress of potato plants with altered levels of glucose oxidase expression. Transformation of potato plants by gox gene led to the more early coming out from tuber dormancy of transformed plants and slower growth rate. Transformants containing the glucose oxidase gene were more sensitive to lethal thermal shock (50 °C, 90 min than the transformant with the empty vector (pBI or untransformed plants (CK. Pre-heating of plants at 37 °C significantly weakened the damaging effect of lethal thermal shock. This attenuation was more significant in the non-transformed plants.

A Biocatalytic One-Pot Approach for the Preparation of Lignin Oligomers Using an Oxidase/Peroxidase Cascade Enzyme System

NARCIS (Netherlands)

Habib, Mohamed H. M.; Deuss, Peter J.; Loncar, Nikola; Trajkovic, Milos; Fraaije, Marco W.

2017-01-01

Synthetic lignin was prepared biocatalytically in a one-pot, two-step reaction using an oxidase/peroxidase cascade enzyme system. Using eugenol in combination with eugenol oxidase and a peroxidase, lignin-like material was produced. The cascade reaction takes advantage of the ability of the oxidase

Identification and characterization of an antennae-specific aldehyde oxidase from the navel orangeworm.

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Young-Moo Choo

Full Text Available Antennae-specific odorant-degrading enzymes (ODEs are postulated to inactivate odorant molecules after they convey their signal. Different classes of insect ODEs are specific to esters, alcohols, and aldehydes--the major functional groups of female-produced, hydrophobic sex pheromones from moth species. Esterases that rapidly inactive acetate and other esters have been well-studied, but less is known about aldehyde oxidases (AOXs. Here we report cloning of an aldehyde oxidase, AtraAOX2, from the antennae of the navel orangeworm (NOW, Amyelois transitella, and the first activity characterization of a recombinant insect AOX. AtraAOX2 gene spans 3,813 bp and encodes a protein with 1,270 amino acid residues. AtraAOX2 cDNA was expressed in baculovirus-infected insect Sf21 cells as a ≈280 kDa homodimer with 140 kDa subunits. Recombinant AtraAOX2 degraded Z11Z13-16Ald and plant volatile aldehydes as substrates. However, as expected for aldehyde oxidases, recombinant AtraAOX2 did not show specificity for Z11Z13-16Ald, the main constituent of the sex pheromone, but showed high activity for plant volatile aldehydes. Our data suggest AtraAOX2 might be involved in degradation of a diversity of aldehydes including sex pheromones, plant-derived semiochemicals, and chemical cues for oviposition sites. Additionally, AtraAOX2 could protect the insect's olfactory system from xenobiotics, including pesticides that might reach the sensillar lymph surrounding the olfactory receptor neurons.

ENHANCING DIRECT ELECTRON TRANSFER OF GLUCOSE OXIDASE USING A GOLD NANOPARTICLE |TITANATE NANOTUBE NANOCOMPOSITE ON A BIOSENSOR

International Nuclear Information System (INIS)

Zhao, Ruoxia; Liu, Xiaoqiang; Zhang, Jiamei; Zhu, Jie; Wong, Danny K.Y.

2015-01-01

ABSTRACT: In this paper, we have developed a gold nanoparticle (GNP) decorated titanate nanotubes (TNT) nanocomposite that aids in the direct electron transfer of a large enzyme, such as glucose oxidase (GOD), in which the electroactive site of flavin adenine dinucleotide is deeply buried within the enzyme. The ionic liquid, brominated 1-decyl-3-methyl imidazole, was used to immobilise the nanocomposite and the enzyme on a glassy carbon electrode to further aid in the electron transfer between GOD and the electrode surface. Nafion was also added to anchor the biosensor scaffold. Initially, the tubiform geometry of titanate nanomaterials and the GNP-TNT nanocomposite was confirmed by microscopic and spectroscopic techniques before glucose oxidase was entrapped in the nanocomposite. Based on voltammetric results, this biosensor showed a strong electrocatalytic capability towards glucose (with a heterogeneous electron transfer rate constant of 7.1 s −1 at 180 mV s −1 ) and the calibration for glucose exhibited a high sensitivity (5.1 μA mM −1 ) and a wide linear range (0.01–1.2 mM). These results demonstrated superior analytical performance of our biosensor over others fabricated using bulkier TiO 2 nanoparticles or nanobundles, which could be attributed to a high degree of biocompatibility to glucose oxidase and electrical conductivity of the nanocomposite

Oxidized low-density lipoproteins upregulate proline oxidase to initiate ROS-dependent autophagy

OpenAIRE

Zabirnyk, Olga; Liu, Wei; Khalil, Shadi; Sharma, Anit; Phang, James M.

2009-01-01

Epidemiological studies showed that high levels of oxidized low-density lipoproteins (oxLDLs) are associated with increased cancer risk. We examined the direct effect of physiologic concentrations oxLDL on cancer cells. OxLDLs were cytotoxic and activate both apoptosis and autophagy. OxLDLs have ligands for peroxisome proliferator-activated receptor gamma and upregulated proline oxidase (POX) through this nuclear receptor. We identified 7-ketocholesterol (7KC) as a main component responsible ...

Glucose oxidase-functionalized fluorescent gold nanoclusters as probes for glucose

Energy Technology Data Exchange (ETDEWEB)

Xia, Xiaodong [College of Chemistry and Chemical Engineering, Central South University, Changsha 410083 (China); School of Chemistry and Chemical Engineering, Hunan University of Science and Technology, Xiangtan 411201 (China); Long, Yunfei, E-mail: l_yunfei927@163.com [School of Chemistry and Chemical Engineering, Hunan University of Science and Technology, Xiangtan 411201 (China); Wang, Jianxiu, E-mail: jxiuwang@csu.edu.cn [College of Chemistry and Chemical Engineering, Central South University, Changsha 410083 (China)

2013-04-15

Highlights: ► A glucose oxidase/gold nanocluster conjugates formed by etching chemistry. ► Integration of the bioactivities and fluorescence properties within a single unit. ► These conjugates serve as novel fluorescent probe for glucose. -- Abstract: Creation and application of noble metal nanoclusters have received continuous attention. By integrating enzyme activity and fluorescence for potential applications, enzyme-capped metal clusters are more desirable. This work demonstrated a glucose oxidase (an enzyme for glucose)-functionalized gold cluster as probe for glucose. Under physiological conditions, such bioconjugate was successfully prepared by an etching reaction, where tetrakis (hydroxylmethyl) phosphonium-protected gold nanoparticle and thioctic acid-modified glucose oxidase were used as precursor and etchant, respectively. These bioconjugates showed unique fluorescence spectra (λ{sub em} {sub max} = 650 nm, λ{sub ex} {sub max} = 507 nm) with an acceptable quantum yield (ca. 7%). Moreover, the conjugated glucose oxidase remained active and catalyzed reaction of glucose and dissolved O{sub 2} to produce H{sub 2}O{sub 2}, which quenched quantitatively the fluorescence of gold clusters and laid a foundation of glucose detection. A linear range of 2.0 × 10{sup −6}–140 × 10{sup −6} M and a detection limit of 0.7 × 10{sup −6} M (S/N = 3) were obtained. Also, another horseradish peroxidase/gold cluster bioconjugate was produced by such general synthesis method. Such enzyme/metal cluster bioconjugates represented a promising class of biosensors for biologically important targets in organelles or cells.

Biochemical Conservation and Evolution of Germacrene A Oxidase in Asteraceae*

Science.gov (United States)

Nguyen, Don Trinh; Göpfert, Jens Christian; Ikezawa, Nobuhiro; MacNevin, Gillian; Kathiresan, Meena; Conrad, Jürgen; Spring, Otmar; Ro, Dae-Kyun

2010-01-01

Sesquiterpene lactones are characteristic natural products in Asteraceae, which constitutes ∼8% of all plant species. Despite their physiological and pharmaceutical importance, the biochemistry and evolution of sesquiterpene lactones remain unexplored. Here we show that germacrene A oxidase (GAO), evolutionarily conserved in all major subfamilies of Asteraceae, catalyzes three consecutive oxidations of germacrene A to yield germacrene A acid. Furthermore, it is also capable of oxidizing non-natural substrate amorphadiene. Co-expression of lettuce GAO with germacrene synthase in engineered yeast synthesized aberrant products, costic acids and ilicic acid, in an acidic condition. However, cultivation in a neutral condition allowed the de novo synthesis of a single novel compound that was identified as germacrene A acid by gas and liquid chromatography and NMR analyses. To trace the evolutionary lineage of GAO in Asteraceae, homologous genes were further isolated from the representative species of three major subfamilies of Asteraceae (sunflower, chicory, and costus from Asteroideae, Cichorioideae, and Carduoideae, respectively) and also from the phylogenetically basal species, Barnadesia spinosa, from Barnadesioideae. The recombinant GAOs from these genes clearly showed germacrene A oxidase activities, suggesting that GAO activity is widely conserved in Asteraceae including the basal lineage. All GAOs could catalyze the three-step oxidation of non-natural substrate amorphadiene to artemisinic acid, whereas amorphadiene oxidase diverged from GAO displayed negligible activity for germacrene A oxidation. The observed amorphadiene oxidase activity in GAOs suggests that the catalytic plasticity is embedded in ancestral GAO enzymes that may contribute to the chemical and catalytic diversity in nature. PMID:20351109

Gene cloning and characterization of NADH oxidase from ...

African Journals Online (AJOL)

use

2011-12-07

Dec 7, 2011 ... potent inhibitors of NADH oxidases, silver nitrate and potassium cyanide did not show any significant ... anaerobes, a class of organisms that have not been ... DNA and amino acid sequence analyses were performed using.

Dithiocarbamates are teratogenic to developing zebrafish through inhibition of lysyl oxidase activity

International Nuclear Information System (INIS)

Boxtel, Antonius L. van; Kamstra, Jorke H.; Fluitsma, Donna M.; Legler, Juliette

2010-01-01

Dithiocarbamates (DTCs) are a class of compounds that are extensively used in agriculture as pesticides. As such, humans and wildlife are undoubtedly exposed to these chemicals. Although DTCs are thought to be relatively safe due to their short half lives, it is well established that they are teratogenic to vertebrates, especially to fish. In zebrafish, these teratogenic effects are characterized by distorted notochord development and shortened anterior to posterior axis. DTCs are known copper (Cu) chelators but this does not fully explain the observed teratogenic effects. We show here that DTCs cause malformations in zebrafish that highly resemble teratogenic effects observed by direct inhibition of a group of cuproenzymes termed lysyl oxidases (LOX). Additionally, we demonstrate that partial knockdown of three LOX genes, lox, loxl1 and loxl5b, sensitizes the developing embryo to DTC exposure. Finally, we show that DTCs directly inhibit zebrafish LOX activity in an ex vivo amine oxidase assay. Taken together, these results provide the first evidence that DTC induced teratogenic effects are, at least in part, caused by direct inhibition of LOX activity.

Time-to-positivity, type of culture media and oxidase test performed on positive blood culture vials to predict Pseudomonas aeruginosa in patients with Gram-negative bacilli bacteraemia.

Science.gov (United States)

Cobos-Triguero, N; Zboromyrska, Y; Morata, L; Alejo, I; De La Calle, C; Vergara, A; Cardozo, C; Arcas, M P; Soriano, A; Marco, F; Mensa, J; Almela, M; Martínez, J A

2017-02-01

The aim of this study was to determine the usefulness of oxidase test and time-to-positivity (TTP) in aerobic and anaerobic blood culture vials to detect the presence of Pseudomonas aeruginosa in patients with Gram-negative bacilli (GNB) bacteraemia. TTP was recorded for each aerobic and anaerobic blood culture vial of monomicrobial bacteraemia due to GNB. Oxidase test was performed in a pellet of the centrifuged content of the positive blood culture. An algorithm was developed in order to perform the oxidase test efficiently taking into account TTP and type of vial. A total of 341 episodes of GNB bacteraemia were analysed. Sensitivity, specificity, positive predictive value and negative predictive value of the oxidase test performed on positive vials with GNB to predict P. aeruginosa were 95%, 99%, 91%, and 99%, respectively. When growth was first or exclusively detected in anaerobic vials, P. aeruginosa was never identified hence the performance of the oxidase test could be avoided. When growth was only or first detected in aerobic vials, a TTP≥8h predicted P. aeruginosa in 37% or cases (63 of 169), therefore oxidase test is highly recommended. Oxidase test performed onto positive blood culture vials previously selected by TTP and type of vials is an easy and inexpensive way to predict P. aeruginosa. In most cases, this can lead to optimization of treatment in less than 24 hours.

Low-temperature kinetic measurements of microsecond freeze-hyperquench (MHQ) cytochrome oxidase monitored by UV-visible spectroscopy with a newly designed cuvette.

Science.gov (United States)

Wiertz, F G M; de Vries, S

2006-02-01

A special cuvette was designed to measure optical changes of MHQ (microsecond freeze-hyperquench) powder samples at temperatures below approx. 250 K. Reduced cytochrome c oxidase from Paracoccus denitrificans was reacted with O(2) for 100 micros, frozen as a powder and transferred to the cuvette. Subsequently, cytochrome oxidase was allowed to react further following stepwise increments of the temperature from 100 K up to 250 K while recording spectra between 300 and 700 nm. The temperature was raised only when no further changes in the spectra could be detected. The experiment yielded spectra of the A, P(M), F and O intermediate states. This demonstrated that the catalytic cycle of cytochrome oxidase at low temperature is similar to that at room temperature and so verifies the suitability of this method for the study of enzymes with high catalytic-centre activity.

Conformational lock and dissociative thermal inactivation of lentil seedling amine oxidase.

Science.gov (United States)

Moosavi-Nejad, S Zahra; Moosavi-Movahedi, Ali-Akbar; Rezaei-Tavirani, Mostafa; Floris, Giovanni; Medda, Rosaria

2003-03-31

The kinetics of thermal inactivation of copper-containing amine oxidase from lentil seedlings were studied in a 100 mM potassium phosphate buffer, pH 7, using putrescine as the substrate. The temperature range was between 47-60 degrees C. The thermal inactivation curves were not linear at 52 and 57 degrees C; three linear phases were shown. The first phase gave some information about the number of dimeric forms of the enzyme that were induced by the higher temperatures using the "conformational lock" pertaining theory to oligomeric enzyme. The "conformational lock" caused two additional dimeric forms of the enzyme when the temperature increased to 57 degrees C. The second and third phases were interpreted according to a dissociative thermal inactivation model. These phases showed that lentil amine oxidase was reversibly-dissociated before the irreversible thermal inactivation. Although lentil amine oxidase is not a thermostable enzyme, its dimeric structure can form "conformational lock," conferring a structural tolerance to the enzyme against heat stress.

The glucose oxidase-peroxidase assay for glucose

Science.gov (United States)

The glucose oxidase-peroxidase assay for glucose has served as a very specific, sensitive, and repeatable assay for detection of glucose in biological samples. It has been used successfully for analysis of glucose in samples from blood and urine, to analysis of glucose released from starch or glycog...

[The X+ chronic granulomatous disease as a fabulous model to study the NADPH oxidase complex activation].

Science.gov (United States)

Stasia, Marie-José

2007-05-01

Chronic granulomatous disease (CGD) is a rare inherited disorder in which phagocytes lack NADPH oxidase activity. Patients with CGD suffer from recurrent bacterial and fungal infections because of the absence of superoxide anions (O2- degrees ) generatingsystem. The NADPH oxidase complex is composed of a membranous cytochrome b558, cytosolic proteins p67phox, p47phox, p40phox and two small GTPases Rac2 and Rap1A. Cytochrome b558 consists of two sub-units gp91phox and p22phox. The most common form of CGD is due to mutations in CYBB gene encoding gp91phox. In some rare cases, the mutated gp91phox is normally expressed but is devoided of oxidase activity. These variants called X+ CGD, have provided interesting informations about oxidase activation mechanisms. However modelization of such variants is necessary to obtain enough biological material for studies at the molecular level. A cellular model (knock-out PLB-985 cells) has been developed for expressing recombinant mutated gp91phox for functional analysis of the oxidase complex. Recent works demonstrated that this cell line genetically deficient in gp91phox is a powerful tool for functional analysis of the NADPH oxidase complex activation.

Interrupted reperfusion reduces the activation of NADPH oxidase after cerebral I/R injury.

Science.gov (United States)

Shen, Jia; Bai, Xiao-Yin; Qin, Yuan; Jin, Wei-Wei; Zhou, Jing-Yin; Zhou, Ji-Ping; Yan, Ying-Gang; Wang, Qiong; Bruce, Iain C; Chen, Jiang-Hua; Xia, Qiang

2011-06-15

Interrupted reperfusion reduces ischemia/reperfusion (I/R) injury. This study was designed to determine whether NADPH oxidase participates in the neural protection against global I/R injury after interrupted reperfusion. Mice were randomly divided into five groups: sham (sham-operated), I/R (20-min global I/R), RR (I/R+interrupted reperfusion), Apo (I/R+apocynin administration), and RR+Apo. Behavioral tests (pole test, beam walking, and Morris water maze) and Nissl staining were undertaken in all five groups; superoxide levels, expression of gp91(phox) and p47(phox), p47(phox) translocation, and Rac1 activation were measured in the sham, I/R, and RR groups. The motor coordination, bradykinesia, and spatial learning and memory, as well as the neuron survival rates, were better in the RR, Apo, and RR+Apo groups than in the I/R group. The NADPH oxidase-dependent superoxide levels, p47(phox) and gp91(phox) expression, p47(phox) translocation, and Rac1 activation were lower in the RR group than in the I/R group. In conclusion, the neural protective effect of interrupted reperfusion is at least partly mediated by decreasing the expression and assembly of NADPH oxidase and the levels of NADPH oxidase-derived superoxide. The most striking reduction Rac1-GTP in the RR group suggests that interrupted reperfusion also acts on the activation of assembled NADPH oxidase by reducing the availability of Rac1-GTP. Copyright © 2011 Elsevier Inc. All rights reserved.

Production of recombinant cholesterol oxidase containing covalently bound FAD in Escherichia coli

Directory of Open Access Journals (Sweden)

Molla Gianluca

2010-04-01

Full Text Available Abstract Background Cholesterol oxidase is an alcohol dehydrogenase/oxidase flavoprotein that catalyzes the dehydrogenation of C(3-OH of cholesterol. It has two major biotechnological applications, i.e. in the determination of serum (and food cholesterol levels and as biocatalyst providing valuable intermediates for industrial steroid drug production. Cholesterol oxidases of type I are those containing the FAD cofactor tightly but not covalently bound to the protein moiety, whereas type II members contain covalently bound FAD. This is the first report on the over-expression in Escherichia coli of type II cholesterol oxidase from Brevibacterium sterolicum (BCO. Results Design of the plasmid construct encoding the mature BCO, optimization of medium composition and identification of the best cultivation/induction conditions for growing and expressing the active protein in recombinant E. coli cells, concurred to achieve a valuable improvement: BCO volumetric productivity was increased from ~500 up to ~25000 U/L and its crude extract specific activity from 0.5 up to 7.0 U/mg protein. Interestingly, under optimal expression conditions, nearly 55% of the soluble recombinant BCO is produced as covalently FAD bound form, whereas the protein containing non-covalently bound FAD is preferentially accumulated in insoluble inclusion bodies. Conclusions Comparison of our results with those published on non-covalent (type I COs expressed in recombinant form (either in E. coli or Streptomyces spp., shows that the fully active type II BCO can be produced in E. coli at valuable expression levels. The improved over-production of the FAD-bound cholesterol oxidase will support its development as a novel biotool to be exploited in biotechnological applications.

A preliminary neutron diffraction study of rasburicase, a recombinant urate oxidase enzyme, complexed with 8-azaxanthin

Energy Technology Data Exchange (ETDEWEB)

Budayova-Spano, Monika, E-mail: spano@embl-grenoble.fr [European Molecular Biology Laboratory Grenoble Outstation, 6 Rue Jules Horowitz, 38042 Grenoble (France); Institut Laue-Langevin, 6 Rue Jules Horowitz, BP 156, 38042 Grenoble (France); Bonneté, Françoise; Ferté, Natalie [Centre de Recherche en Matière Condensée et Nanosciences, Campus de Luminy, Case 913, 13288 Marseille (France); El Hajji, Mohamed [Sanofi-Aventis, 371 Rue du Professeur Blayac, 34184 Montpellier (France); Meilleur, Flora [Institut Laue-Langevin, 6 Rue Jules Horowitz, BP 156, 38042 Grenoble (France); Blakeley, Matthew Paul [European Molecular Biology Laboratory Grenoble Outstation, 6 Rue Jules Horowitz, 38042 Grenoble (France); Castro, Bertrand [Sanofi-Aventis, 371 Rue du Professeur Blayac, 34184 Montpellier (France); European Molecular Biology Laboratory Grenoble Outstation, 6 Rue Jules Horowitz, 38042 Grenoble (France)

2006-03-01

Neutron diffraction data of hydrogenated recombinant urate oxidase enzyme (Rasburicase), complexed with a purine-type inhibitor 8-azaxanthin, was collected to 2.1 Å resolution from a crystal grown in D{sub 2}O by careful control and optimization of crystallization conditions via knowledge of the phase diagram. Deuterium atoms were clearly seen in the neutron-scattering density map. Crystallization and preliminary neutron diffraction measurements of rasburicase, a recombinant urate oxidase enzyme expressed by a genetically modified Saccharomyces cerevisiae strain, complexed with a purine-type inhibitor (8-azaxanthin) are reported. Neutron Laue diffraction data were collected to 2.1 Å resolution using the LADI instrument from a crystal (grown in D{sub 2}O) with volume 1.8 mm{sup 3}. The aim of this neutron diffraction study is to determine the protonation states of the inhibitor and residues within the active site. This will lead to improved comprehension of the enzymatic mechanism of this important enzyme, which is used as a protein drug to reduce toxic uric acid accumulation during chemotherapy. This paper illustrates the high quality of the neutron diffraction data collected, which are suitable for high-resolution structural analysis. In comparison with other neutron protein crystallography studies to date in which a hydrogenated protein has been used, the volume of the crystal was relatively small and yet the data still extend to high resolution. Furthermore, urate oxidase has one of the largest primitive unit-cell volumes (space group I222, unit-cell parameters a = 80, b = 96, c = 106 Å) and molecular weights (135 kDa for the homotetramer) so far successfully studied with neutrons.

Surface reconstitution of glucose oxidase onto a norbornylogous bridge self-assembled monolayer

International Nuclear Information System (INIS)

Liu Jingquan; Paddon-Row, Michael N.; Gooding, J. Justin

2006-01-01

An electrode construct was fabricated in which a self-assembled monolayer containing a novel norbornylogous bridge was covalently attached to flavin adenine dinucleotide (FAD), the redox active centre of several oxidase enzymes. The electrochemistry of the construct was investigated before and after the reconstitution of glucose oxidase around the surface bound FAD. Rapid rates of electron transfer were observed both before and after the reconstitution of biocatalytically active enzyme. However, no biocatalytic activity was observed under anaerobic conditions suggesting the a lack of enzyme turnover through direct electron transfer. It is proposed that a decrease in the electronic coupling between the redox active FAD and the electrode following reconstitution of the glucose oxidase - a probable consequence of the FAD being immersed in a protein environment - was responsible for the inability of the enzyme to be turned over under anaerobic conditions

Glucose oxidase probe as a surface-enhanced Raman scattering sensor for glucose.

Science.gov (United States)

Qi, Guohua; Wang, Yi; Zhang, Biying; Sun, Dan; Fu, Cuicui; Xu, Weiqing; Xu, Shuping

2016-10-01

Glucose oxidase (GOx) possessing a Raman-active chromophore (flavin adenine dinucleotide) is used as a signal reporter for constructing a highly specific "turn off" surface-enhanced Raman scattering (SERS) sensor for glucose. This sensing chip is made by the electrostatic assembly of GOx over silver nanoparticle (Ag NP)-functionalized SERS substrate through a positively charged polyelectrolyte linker under the pH of 6.86. To trace glucose in blood serum, owing to the reduced pH value caused by the production of gluconic acid in the GOx-catalyzed oxidation reaction, the bonding force between GOx and polyelectrolyte weakens, making GOx drop off from the sensing chip. As a result, the SERS intensity of GOx on the chip decreases along with the concentration of glucose. This glucose SERS sensor exhibits excellent selectivity based on the specific GOx/glucose catalysis reaction and high sensitivity to 1.0 μM. The linear sensing range is 2.0-14.0 mM, which also meets the requirement on the working range of the human blood glucose detection. Using GOx as a probe shows superiority over other organic probes because GOx almost has no toxicity to the biological system. This sensing mechanism can be applied for intracellular in vivo SERS monitoring of glucose in the future. Graphical abstract Glucose oxidase is used as a Raman signal reporter for constructing a highly specific glucose surface-enhanced Raman scattering (SERS) sensor.

Downregulation of Lysyl Oxidase Protects Retinal Endothelial Cells From High Glucose-Induced Apoptosis.

Science.gov (United States)

Kim, Dongjoon; Mecham, Robert P; Trackman, Philip C; Roy, Sayon

2017-05-01

To investigate the effect of reducing high glucose (HG)-induced lysyl oxidase (LOX) overexpression and increased activity on retinal endothelial cell apoptosis. Rat retinal endothelial cells (RRECs) were grown in normal (N) or HG (30 mM glucose) medium for 7 days. In parallel, RRECs were grown in HG medium and transfected with LOX small interfering RNA (siRNA), scrambled siRNA as control, or exposed to β-aminopropionitrile (BAPN), a LOX inhibitor. LOX expression, AKT activation, and caspase-3 activity were determined by Western blot (WB) analysis and apoptosis by differential dye staining assay. Moreover, to determine whether diabetes-induced LOX overexpression alters AKT activation and promotes apoptosis, changes in LOX expression, AKT phosphorylation, caspase-3 activation, and Bax expression were assessed in retinas of streptozotocin (STZ)-induced diabetic mice and LOX heterozygous knockout (LOX+/-) mice. WB analysis indicated significant LOX overexpression and reduced AKT activation under HG condition in RRECs. Interestingly, when cells grown in HG were transfected with LOX siRNA or exposed to BAPN, the number of apoptotic cells was significantly decreased concomitant with increased AKT phosphorylation. Diabetic mouse retinas exhibited LOX overexpression, decreased AKT phosphorylation, and increased Bax and caspase-3 activation compared to values in nondiabetic mice. In LOX+/- mice, reduced LOX levels were observed with increased AKT activity, and reduced Bax and caspase-3 activity. Furthermore, decreased levels of LOX in the LOX+/- mice was protective against diabetes-induced apoptosis. Findings from this study indicate that preventing LOX overexpression may be protective against HG-induced apoptosis in retinal vascular cells associated with diabetic retinopathy.

Glucose oxidase stabilization against thermal inactivation using high hydrostatic pressure and hydrophobic modification.

Science.gov (United States)

Halalipour, Ali; Duff, Michael R; Howell, Elizabeth E; Reyes-De-Corcuera, José I

2017-03-01

High hydrostatic pressure (HHP) stabilized glucose oxidase (GOx) against thermal inactivation. The apparent first-order kinetics of inactivation of GOx were investigated at 0.1-300 MPa and 58.8-80.0°C. At 240 MPa and 74.5°C, GOx inactivated at a rate 50 times slower than at atmospheric pressure at the same temperature. The apparent activation energy of inactivation at 300 MPa was 281.0 ± 17.4 kJ mol -1 or 1.3-fold smaller than for the inactivation at atmospheric pressure (378.1 ± 25.6 kJ mol -1 ). The stabilizing effect of HHP was greatest at 74.5°C, where the activation volume of 57.0 ± 12.0 cm 3  mol -1 was highest compared to all other studied temperatures. Positive apparent activation volumes for all the treatment temperatures confirmed that HHP favors GOx stabilization. A second approach to increase GOx stability involved crosslinking with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC) and either aniline or benzoate. The modified enzyme remained fully active with only slight increases in K M (1.3-1.9-fold increases for aniline and benzoate modification, respectively). The thermal stability of GOx increased by 8°C with aniline modification, while it decreased by 0.9°C upon modification with benzoate. Biotechnol. Bioeng. 2017;114: 516-525. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Genetic defects of cytochrome c oxidase assembly

Czech Academy of Sciences Publication Activity Database

Pecina, Petr; Houšťková, H.; Hansíková, H.; Zeman, J.; Houštěk, Josef

2004-01-01

Roč. 53, Suppl. 1 (2004), s. S213-S223 ISSN 0862-8408 R&D Projects: GA ČR GA303/03/0749 Institutional research plan: CEZ:AV0Z5011922 Keywords : cytochrome c oxidase * mitochondrial disorders Subject RIV: FB - Endocrinology, Diabetology, Metabolism, Nutrition Impact factor: 1.140, year: 2004

High-content screening of Aspergillus niger with both increased production and high secretion rate of glucose oxidase.

Science.gov (United States)

Zhu, Xudong; Sun, Jingchun; Chu, Ju

2018-01-01

To develop a rapid, dual-parameter, plate-based screening process to improve production and secretion rate of glucose oxidase simultaneously in Aspergillus niger. A morphology engineering based on CaCO 3 was implemented, where the yield of GOD by A. niger was increased by up to 50%. Analysis of extracellular GOD activity was achieved in 96-well plates. There was a close negative correlation between the total GOD activity and its residual glucose of the fermentation broth. Based on this, a rapid, plate-based, qualitative analysis method of the total GOD activity was developed. Compared with the conventional analysis method using o-dianisidine, a correlation coefficient of -0.92 by statistical analysis was obtained. Using this dual-parameter screening method, we acquired a strain with GOD activity of 3126 U l -1 , which was 146% higher than the original strain. Its secretion rate of GOD was 83, 32% higher than the original strain.

Direct and indirect effects of RNA interference against pyridoxal kinase and pyridoxine 5'-phosphate oxidase genes in Bombyx mori.

Science.gov (United States)

Huang, ShuoHao; Yao, LiLi; Zhang, JianYun; Huang, LongQuan

2016-08-01

Vitamin B6 comprises six interconvertible pyridine compounds (vitamers), among which pyridoxal 5'-phosphate is a coenzyme involved in a high diversity of biochemical reactions. Humans and animals obtain B6 vitamers from diet, and synthesize pyridoxal 5'-phosphate by pyridoxal kinase and pyridoxine 5'-phosphate oxidase. Currently, little is known on how pyridoxal 5'-phosphate biosynthesis is regulated, and pyridoxal 5'-phosphate is supplied to meet their requirement in terms of cofactor. Bombyx mori is a large silk-secreting insect, in which protein metabolism is most active, and the vitamin B6 demand is high. In this study, we successfully down-regulated the gene expression of pyridoxal kinase and pyridoxine 5'-phosphate oxidase by body cavity injection of synthesized double-stranded small interfering RNA to 5th instar larvae of Bombyx mori, and analyzed the gene transcription levels of pyridoxal 5'-phosphate dependent enzymes, phosphoserine aminotransferase and glutamic-oxaloacetic transaminase. Results show that the gene expression of pyridoxal kinase and pyridoxine 5'-phosphate oxidase has a greater impact on the gene transcription of enzymes using pyridoxal 5'-phosphate as a cofactor in Bombyx mori. Our study suggests that pyridoxal 5'-phosphate biosynthesis and dynamic balance may be regulated by genetic networks. Copyright © 2016 Elsevier B.V. All rights reserved.

Combined effect of loss of the caa3 oxidase and Crp regulation drives Shewanella to thrive in redox-stratified environments.

Science.gov (United States)

Zhou, Guangqi; Yin, Jianhua; Chen, Haijiang; Hua, Yijie; Sun, Linlin; Gao, Haichun

2013-09-01

Shewanella species are a group of facultative Gram-negative microorganisms with remarkable respiration abilities that allow the use of a diverse array of terminal electron acceptors (EA). Like most bacteria, S. oneidensis possesses multiple terminal oxidases, including two heme-copper oxidases (caa3- and cbb3-type) and a bd-type quinol oxidase. As aerobic respiration is energetically favored, mechanisms underlying the fact that these microorganisms thrive in redox-stratified environments remain vastly unexplored. In this work, we discovered that the cbb3-type oxidase is the predominant system for respiration of oxygen (O2), especially when O2 is abundant. Under microaerobic conditions, the bd-type quinol oxidase has a significant role in addition to the cbb3-type oxidase. In contrast, multiple lines of evidence suggest that under test conditions the caa3-type oxidase, an analog to the mitochondrial enzyme, has no physiological significance, likely because of its extremely low expression. In addition, expression of both cbb3- and bd-type oxidases is under direct control of Crp (cAMP receptor protein) but not the well-established redox regulator Fnr (fumarate nitrate regulator) of canonical systems typified in Escherichia coli. These data, collectively, suggest that adaptation of S. oneidensis to redox-stratified environments is likely due to functional loss of the caa3-type oxidase and switch of the regulatory system for respiration.

Identification and statistical optimization of fermentation conditions for a newly isolated extracellular cholesterol oxidase-producing Streptomyces cavourensis strain NEAE-42

OpenAIRE

El-Naggar, Noura El-Ahmady; El-Shweihy, Nancy M.; El-Ewasy, Sara M.

2016-01-01

Background Due to broad range of clinical and industrial applications of cholesterol oxidase, isolation and screening of bacterial strains producing extracellular form of cholesterol oxidase is of great importance. Results One hundred and thirty actinomycete isolates were screened for their cholesterol oxidase activity. Among them, a potential culture, strain NEAE-42 is displayed the highest extracellular cholesterol oxidase activity. It was selected and identified as Streptomyces cavourensis...

Effective immobilization of glucose oxidase on chitosan submicron particles from gladius of Todarodes pacificus for glucose sensing.

Science.gov (United States)

Anusha, J R; Fleming, Albin T; Kim, Hee-Je; Kim, Byung Chul; Yu, Kook-Hyun; Raj, C Justin

2015-08-01

An effective enzymatic glucose biosensor was developed by immobilizing glucose oxidase on chitosan submicron particles synthesized from the gladius of Todarodes pacificus (GCSP). The chemically synthesized chitosan from gladius was pulverized to submicron particles by ball milling technique, which was further characterized and compared with the standard chitosan (SCS). The degree of deacetylation of GCSP was determined using FTIR spectroscopy which was comparable to the value of standard chitosan. The glucose oxidase (GOx) was immobilized over GCSP on porous zinc oxide/platinum nanoparticle (ZnO/Pt) based electrode. The morphological and structural properties of the electrodes were analyzed using scanning electron microscopy and X-ray diffraction analysis. The glucose sensing behavior of electrode was estimated using electrochemical analysis and showed an excellent analytical performance. The electrode ZnO/Pt/GCSP conjugated with GOx displayed high sensitivity (88.76 μA mM(-1) cm(-2)) with low detection limit in short response time. In addition, the very low value of Michaelis-Menten constant for GCSP based electrode contributes a better affinity of the electrode surface towards glucose oxidase. Copyright © 2015 Elsevier B.V. All rights reserved.

Effect of temperature stress on polyphenol oxidase activity in grains of some wheat cultivars

International Nuclear Information System (INIS)

Kayani, W.K.

2011-01-01

Color is a key quality trait of wheat-based products and polyphenol oxidase (PPO) is implicated to play a significant role in their undesirable darkening. Polyphenol oxidase catalyzes the oxidation of phenols to quinines, which auto oxidize and polymerize with amino acid of cellular proteins resulting brown and black pigmentation propounding reduced nutritional values. In present study, the PPO activity in 50 different Pakistani wheat cultivars was investigated and grouped into three categories viz; low, medium and high PPO activity cultivars. PPO is a heat labile enzyme. To investigate effect of heat stress, nine cultivars from each category were chosen for treatment at 30, 40 and 50 deg. C for 30, 60, and 120 minutes each. A substantial change was experienced in PPO activity as compared to room temperature. Two wheat cultivar Wafaq-2001 and AS-2002 showed a compromising attitude of minimum PPO activity at 30 deg. C for a period of 30 and 60 minutes of incubation. In general, an incubation of 30 deg. C or 60 deg. C (low or high) for a period of 30 minutes can be recommended for suppressing PPO activity. (author)

NADPH oxidase and redox status in amygdala, hippocampus and cortex of male Wistar rats in an animal model of post-traumatic stress disorder.

Science.gov (United States)

Petrovic, Romana; Puskas, Laslo; Jevtic Dozudic, Gordana; Stojkovic, Tihomir; Velimirovic, Milica; Nikolic, Tatjana; Zivkovic, Milica; Djorovic, Djordje J; Nenadovic, Milutin; Petronijevic, Natasa

2018-05-26

Post-traumatic stress disorder (PTSD) is a highly prevalent and impairing disorder. Oxidative stress is implicated in its pathogenesis. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase is an important source of free radicals. The aim of the study was to assess oxidative stress parameters, activities of respiratory chain enzymes, and the expression of NADPH oxidase subunits (gp91phox, p22phox, and p67phox) in the single prolonged stress (SPS) animal model of PTSD. Twenty-four (12 controls; 12 subjected to SPS), 9-week-old, male Wistar rats were used. SPS included physical restraint, forced swimming, and ether exposure. The rats were euthanized seven days later. Cortex, hippocampus, amygdala, and thalamus were dissected. Malondialdehyde (MDA), reduced glutathione (GSH), superoxide dismutase (SOD), glutathione peroxidase (GPx), catalase (CAT), Complex I, and cytochrome C oxidase were measured using spectrophotometric methods, while the expression of NADPH oxidase subunits was determined by Western blot. Increased MDA and decreased GSH concentrations were found in the amygdala and hippocampus of the SPS rats. SOD activity was decreased in amygdala and GPx was decreased in hippocampus. Increased expression of the NADPH oxidase subunits was seen in amygdala, while mitochondrial respiratory chain enzyme expression was unchanged both in amygdala and hippocampus. In the cortex concentrations of MDA and GSH were unchanged despite increased Complex I and decreased GPx, while in the thalamus no change of any parameter was noticed. We conclude that oxidative stress is present in hippocampus and amygdala seven days after the SPS procedure. NADPH oxidase seems to be a main source of free radicals in the amygdala.

Crystallization of carbohydrate oxidase from Microdochium nivale

Czech Academy of Sciences Publication Activity Database

Dušková, Jarmila; Dohnálek, Jan; Skálová, Tereza; Ostergaard, L. H.; Fuglsang, C. C.; Kolenko, Petr; Štěpánková, Andrea; Hašek, Jindřich

2009-01-01

Roč. 65, č. 6 (2009), s. 638-640 ISSN 1744-3091 R&D Projects: GA AV ČR IAA500500701; GA ČR GA305/07/1073 Institutional research plan: CEZ:AV0Z40500505 Keywords : carbohydrate oxidase * crystallization * data processing Subject RIV: CD - Macromolecular Chemistry Impact factor: 0.551, year: 2009

Immunological and molecular comparison of polyphenol oxidase in Rosaceae fruit trees.

Science.gov (United States)

Haruta, M; Murata, M; Kadokura, H; Homma, S

1999-03-01

An antibody raised against apple polyphenol oxidase (PPO) cross-reacted with PPOs from Japanese pear (Pyrus pyrifolia), pear (Pyrus communis), peach (Prunus persica), Chinese quince (Pseudocydonia sinensis) and Japanese loquat (Eriobotrya japonica). Core fragments (681 bp) of the corresponding PPO genes were amplified and characterized. The deduced protein sequences showed identities of 85.3 to 97.5%. Chlorogenic acid oxidase activity of these PPOs showed higher activities when assayed at pH 4 than at pH 6. These results indicate that PPOs in Rosaceae plants are structurally and enzymatically similar.

Ratiometric glucose sensing based on fluorescent oxygen films and glucose oxidase

OpenAIRE

Fengyu Su; Liqiang Zhang; Xiangxing Kong; Fred Lee; Yanqing Tian; Deirdre R. Meldrum

2017-01-01

A new two-layer sensor film was constructed for sensing glucose based on glucose oxidase and oxygen sensing material. The first layer of film containing the oxygen sensor and intra-reference material was polymerized, then the second layer of glucose oxidase and glutaraldehyde was formed on the oxygen sensor layer. The two-layer sensor film has a resolution up to 0.05 mM and a detection range from 0 to 5 mM to glucose. The effects of pH and temperature on the sensing performance were systemati...

High Catalysis Activity of Cu₂O Microcrystals to the Electrochemiluminesence of Luminol and H₂O₂

DEFF Research Database (Denmark)

Zhang, Ling; Xu, Guobao; Zhang, Jingdong

Cuprous oxide (Cu2O) is a classical p-type semiconductor with a direct band gap of 2.17 eV, which is wildly used for solar energy conversion, CO oxidation, and photo catalytic water splitting for the low cost and environmental friendliness. For the energy band positions are favorable to the hydro......Cuprous oxide (Cu2O) is a classical p-type semiconductor with a direct band gap of 2.17 eV, which is wildly used for solar energy conversion, CO oxidation, and photo catalytic water splitting for the low cost and environmental friendliness. For the energy band positions are favorable...... to the hydrogen evolution and oxygen evolution potentials, Cu2O materials also catalyze the reduction of hydrogen peroxide (H2O2), which is an critical molecule in the bodies’ metabolism processes or the industrial catalysis reactions. To improve detection sensitivity of H2O2, people have composed Cu2O materials...... that Cu2O semiconductor microcrystal possess the good catalytic performance to this ECL reaction, which is important to develop the high-efficient and lowcost biosensors....

Extra virgin olive oil rich in polyphenols modulates VEGF-induced angiogenic responses by preventing NADPH oxidase activity and expression.

Science.gov (United States)

Calabriso, Nadia; Massaro, Marika; Scoditti, Egeria; D'Amore, Simona; Gnoni, Antonio; Pellegrino, Mariangela; Storelli, Carlo; De Caterina, Raffaele; Palasciano, Giuseppe; Carluccio, Maria Annunziata

2016-02-01

Previous studies have shown the antiinflammatory, antioxidant and antiangiogenic properties by pure olive oil polyphenols; however, the effects of olive oil phenolic fraction on the inflammatory angiogenesis are unknown. In this study, we investigated the effects of the phenolic fraction (olive oil polyphenolic extract, OOPE) from extra virgin olive oil and related circulating metabolites on the VEGF-induced angiogenic responses and NADPH oxidase activity and expression in human cultured endothelial cells. We found that OOPE (1-10 μg/ml), at concentrations achievable nutritionally, significantly reduced, in a concentration-dependent manner, the VEGF-induced cell migration, invasiveness and tube-like structure formation through the inhibition of MMP-2 and MMP-9. OOPE significantly (Pextra virgin olive oil, with high polyphenol content, decreased VEGF-induced NADPH oxidase activity and Nox4 expression, as well as, MMP-9 expression, as compared with fasting control serum. Overall, native polyphenols and serum metabolites of extra virgin olive oil rich in polyphenols are able to lower the VEGF-induced angiogenic responses by preventing endothelial NADPH oxidase activity and decreasing the expression of selective NADPH oxidase subunits. Our results provide an alternative mechanism by which the consumption of olive oil rich in polyphenols may account for a reduction of oxidative stress inflammatory-related sequelae associated with chronic degenerative diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

RhoA/ROCK downregulates FPR2-mediated NADPH oxidase activation in mouse bone marrow granulocytes.

Science.gov (United States)

Filina, Julia V; Gabdoulkhakova, Aida G; Safronova, Valentina G

2014-10-01

Polymorphonuclear neutrophils (PMNs) express the high and low affinity receptors to formylated peptides (mFPR1 and mFPR2 in mice, accordingly). RhoA/ROCK (Rho activated kinase) pathway is crucial for cell motility and oxidase activity regulated via FPRs. There are contradictory data on RhoA-mediated regulation of NADPH oxidase activity in phagocytes. We have shown divergent Rho GTPases signaling via mFPR1 and mFPR2 to NADPH oxidase in PMNs from inflammatory site. The present study was aimed to find out the role of RhoA/ROCK in the respiratory burst activated via mFPR1 and mFPR2 in the bone marrow PMNs. Different kinetics of RhoA activation were detected with 0.1μM fMLF and 1μM WKYMVM operating via mFPR1 and mFPR2, accordingly. RhoA was translocated in fMLF-activated cells towards the cell center and juxtamembrane space versus uniform allocation in the resting cells. Specific inhibition of RhoA by CT04, Rho inhibitor I, weakly depressed the respiratory burst induced via mFPR1, but significantly increased the one induced via mFPR2. Inhibition of ROCK, the main effector of RhoA, by Y27632 led to the same effect on the respiratory burst. Regulation of mFPR2-induced respiratory response by ROCK was impossible under the cytoskeleton disruption by cytochalasin D, whereas it persisted in the case of mFPR1 activation. Thus we suggest RhoA to be one of the regulatory and signal transduction components in the respiratory burst through FPRs in the mouse bone marrow PMNs. Both mFPR1 and mFPR2 binding with a ligand trigger the activation of RhoA. FPR1 signaling through RhoA/ROCK increases NADPH-oxidase activity. But in FPR2 action RhoA/ROCK together with cytoskeleton-linked systems down-regulates NADPH-oxidase. This mechanism could restrain the reactive oxygen species dependent damage of own tissues during the chemotaxis of PMNs and in the resting cells. Copyright © 2014 Elsevier Inc. All rights reserved.

Kinetic Resolution of sec-Thiols via Enantioselective Oxidation with Rationally Engineered 5-(Hydroxymethyl)furfural Oxidase

NARCIS (Netherlands)

Pickl, Mathias; Swoboda, Alexander; Romero, Elvira; Winkler, Christoph; Binda, Claudia; Mattevi, Andrea; Faber, Kurt; Fraaije, Marco

2018-01-01

Various flavoprotein oxidases were recently shown to oxidize prim-thiols. Here we extend this reactivity towards sec-thiols via structure-guided engineering of 5-(hydroxymethyl)furfural oxidase (HMFO). The variants obtained were employed for the oxidative kinetic resolution of rac-sec-thiols

Alternative oxidase in the branched mitochondrial respiratory network: an overview on structure, function, regulation, and role

Directory of Open Access Journals (Sweden)

Sluse F.E.

1998-01-01

Full Text Available Plants and some other organisms including protists possess a complex branched respiratory network in their mitochondria. Some pathways of this network are not energy-conserving and allow sites of energy conservation to be bypassed, leading to a decrease of the energy yield in the cells. It is a challenge to understand the regulation of the partitioning of electrons between the various energy-dissipating and -conserving pathways. This review is focused on the oxidase side of the respiratory chain that presents a cyanide-resistant energy-dissipating alternative oxidase (AOX besides the cytochrome pathway. The known structural properties of AOX are described including transmembrane topology, dimerization, and active sites. Regulation of the alternative oxidase activity is presented in detail because of its complexity. The alternative oxidase activity is dependent on substrate availability: total ubiquinone concentration and its redox state in the membrane and O2 concentration in the cell. The alternative oxidase activity can be long-term regulated (gene expression or short-term (post-translational modification, allosteric activation regulated. Electron distribution (partitioning between the alternative and cytochrome pathways during steady-state respiration is a crucial measurement to quantitatively analyze the effects of the various levels of regulation of the alternative oxidase. Three approaches are described with their specific domain of application and limitations: kinetic approach, oxygen isotope differential discrimination, and ADP/O method (thermokinetic approach. Lastly, the role of the alternative oxidase in non-thermogenic tissues is discussed in relation to the energy metabolism balance of the cell (supply in reducing equivalents/demand in energy and carbon and with harmful reactive oxygen species formation.

Lower growth temperature increases alternative pathway capacity and alternative oxidase protein in tobacco.

Science.gov (United States)

Vanlerberghe, G C; McIntosh, L

1992-09-01

Suspension cells of NT1 tobacco (Nicotiana tabacum L. cv bright yellow) have been used to study the effect of growth temperature on the CN-resistant, salicylhydroxamic acid-sensitive alternative pathway of respiration. Mitochondria isolated from cells maintained at 30 degrees C had a low capacity to oxidize succinate via the alternative pathway, whereas mitochondria isolated from cells 24 h after transfer to 18 degrees C displayed, on average, a 5-fold increase in this capacity (from 7 to 32 nanoatoms oxygen per milligram protein per minute). This represented an increase in alternative pathway capacity from 18 to 45% of the total capacity of electron transport. This increased capacity was lost upon transfer of cells back to 30 degrees C. A monoclonal antibody to the terminal oxidase of the alternative pathway (the alternative oxidase) from Sauromatum guttatum (T.E. Elthon, R.L. Nickels, L. McIntosh [1989] Plant Physiology 89: 1311-1317) recognized a 35-kilodalton mitochondrial protein in tobacco. There was an excellent correlation between the capacity of the alternative path in isolated tobacco mitochondria and the levels of this 35-kilodalton alternative oxidase protein. Cycloheximide could inhibit both the increased level of the 35-kilodalton alternative oxidase protein and the increased alternative pathway capacity normally seen upon transfer to 18 degrees C. We conclude that transfer of tobacco cells to the lower temperature increases the capacity of the alternative pathway due, at least in part, to de novo synthesis of the 35-kilodalton alternative oxidase protein.

Optimization of cholesterol oxidase production by Brevibacterium sp ...

African Journals Online (AJOL)

An ultrasound-assisted emulsification as a pretreatment for cholesterol oxidase production by submerge fermentation using Brevibacterium sp. in a batch system was studied. Medium improvement for the production employing response surface methodology (RSM) was optimized in this paper. The concentration of ...

Impact of high pressure processing on color, bioactive compounds, polyphenol oxidase activity, and microbiological attributes of pumpkin purée.

Science.gov (United States)

González-Cebrino, Francisco; Durán, Rocío; Delgado-Adámez, Jonathan; Contador, Rebeca; Bernabé, Rosario Ramírez

2016-04-01

Physicochemical parameters, bioactive compounds' content (carotenoids and total phenols), total antioxidant activity, and enzymatic activity of polyphenol oxidase (PPO) were evaluated after high pressure processing (HPP) on a pumpkin purée (cv. 'Butternut'). Three pressure levels (400, 500, and 600 MPa) were combined with three holding times (200, 400, and 600 s). The applied treatments reduced the levels of total aerobic mesophilic (TAM), total psychrophilic and psychrotrophic bacteria (TPP), and molds and yeasts (M&Y). All applied treatments did not affect enzymatic activity of PPO. Pressure level increased CIE L* values, which could enhance the lightness perception of high pressure (HP)-treated purées. No differences were found between the untreated and HP-treated purées regarding total phenols and carotenoids content (lutein, α-carotene, and β-carotene) and total antioxidant activity. HPP did not affect most quality parameters and maintained the levels of bioactive compounds. However, it did not achieve the complete inhibition of PPO, which could reduce the shelf-life of the pumpkin purée. © The Author(s) 2015.

Hydroxychavicol: a potent xanthine oxidase inhibitor obtained from the leaves of betel, Piper betle.

Science.gov (United States)

Murata, Kazuya; Nakao, Kikuyo; Hirata, Noriko; Namba, Kensuke; Nomi, Takao; Kitamura, Yoshihisa; Moriyama, Kenzo; Shintani, Takahiro; Iinuma, Munekazu; Matsuda, Hideaki

2009-07-01

The screening of Piperaceous plants for xanthine oxidase inhibitory activity revealed that the extract of the leaves of Piper betle possesses potent activity. Activity-guided purification led us to obtain hydroxychavicol as an active principle. Hydroxychavicol is a more potent xanthine oxidase inhibitor than allopurinol, which is clinically used for the treatment of hyperuricemia.

The Cytochrome bd Oxidase of Porphyromonas gingivalis Contributes to Oxidative Stress Resistance and Dioxygen Tolerance.

Directory of Open Access Journals (Sweden)

Julia Leclerc

Full Text Available Porphyromonas gingivalis is an etiologic agent of periodontal disease in humans. The disease is associated with the formation of a mixed oral biofilm which is exposed to oxygen and environmental stress, such as oxidative stress. To investigate possible roles for cytochrome bd oxidase in the growth and persistence of this anaerobic bacterium inside the oral biofilm, mutant strains deficient in cytochrome bd oxidase activity were characterized. This study demonstrated that the cytochrome bd oxidase of Porphyromonas gingivalis, encoded by cydAB, was able to catalyse O2 consumption and was involved in peroxide and superoxide resistance, and dioxygen tolerance.

Urate oxidase for the prevention and treatment of tumour lysis syndrome in children with cancer.

Science.gov (United States)

Cheuk, Daniel Kl; Chiang, Alan Ks; Chan, Godfrey Cf; Ha, Shau Yin

2017-03-08

% CI 1.29 to 64.00; P = 0.03).Another included RCT, with 30 participants, compared different doses of rasburicase (0.2 mg/kg versus 0.15 mg/kg). The primary outcome was not evaluated. No clear evidence of a difference in mortality (all-cause mortality (Fisher's exact test P = 1.0) and mortality due to TLS (no deaths in both groups)) and renal failure (no renal failure in both groups) was identified. It demonstrated no clear evidence of a difference in uric acid normalisation (RR 1.07, 95% CI 0.89 to 1.28; P = 0.49) and uric acid level at four hours (MD 8.10%, 95% CI -0.99% to 17.19%; P = 0.08). Common adverse events of urate oxidase included hypersensitivity, haemolysis, and anaemia, but no clear evidence of a difference between treatment groups was identified (RR 0.54, 95% CI 0.12 to 2.48; P = 0.42).The quality of evidence ranks from very low to low because of imprecise results, and all included trials were highly susceptible to biases. Although urate oxidase might be effective in reducing serum uric acid, it is unclear whether it reduces clinical TLS, renal failure, or mortality. Adverse effects might be more common for urate oxidase compared with allopurinol. Clinicians should weigh the potential benefits of reducing uric acid and uncertain benefits of preventing mortality or renal failure from TLS against the potential risk of adverse effects.

Quinolinic Carboxylic Acid Derivatives as Potential Multi-target Compounds for Neurodegeneration: Monoamine Oxidase and Cholinesterase Inhibition.

Science.gov (United States)

Khan, Nehal A; Khan, Imtiaz; Abid, Syed M A; Zaib, Sumera; Ibrar, Aliya; Andleeb, Hina; Hameed, Shahid; Iqbal, Jamshed

2018-01-01

Parkinson's disease (PD), a debilitating and progressive disorder, is among the most challenging and devastating neurodegenerative diseases predominantly affecting the people over 60 years of age. To confront PD, an advanced and operational strategy is to design single chemical functionality able to control more than one target instantaneously. In this endeavor, for the exploration of new and efficient inhibitors of Parkinson's disease, we synthesized a series of quinoline carboxylic acids (3a-j) and evaluated their in vitro monoamine oxidase and cholinesterase inhibitory activities. The molecular docking and in silico studies of the most potent inhibitors were performed to identify the probable binding modes in the active site of the monoamine oxidase enzymes. Moreover, molecular properties were calculated to evaluate the druglikeness of the compounds. The biological evaluation results revealed that the tested compounds were highly potent against monoamine oxidase (A & B), 3c targeted both the isoforms of MAO with IC50 values of 0.51 ± 0.12 and 0.51 ± 0.03 µM, respectively. The tested compounds also demonstrated high and completely selective inhibitory action against acetylcholinesterase (AChE) with IC50 values ranging from 4.36 to 89.24 µM. Among the examined derivatives, 3i was recognized as the most potent inhibitor of AChE with an IC50 value of 4.36 ± 0.12 ±µM. The compounds appear to be promising inhibitors and could be used for the future development of drugs targeting neurodegenerative disorders. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Antioxidant Activity of Some Plant Extracts Towards Xanthine Oxidase, Lipoxygenase and Tyrosinase

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Pi-Yu Chen

2009-08-01

Full Text Available Natural products have the potential to be developed into new drugs for the treatment of various diseases. The aim of the present study was to screen the antioxidant activities of some common edible fruits, garden plants and medicinal plants indigenous to Taiwan. This was performed by assessing the activities of lipoxygenase, xanthine oxidase and tyrosinase following incubation with extracts from these plants. A further aim was to use HPLC-DAD and tyrosinase to chromatographically identify the antioxidative constituents obtained from an extract exhibiting strong antioxidative properties. The acetone extracts of 27 cultivated plant species from Taiwan were tested for antioxidant activities towards xanthine oxidase, tyrosinase and lipoxygenase using spectrophotometric assays. Koelreuteria henryi, Prunus campanulata, and Rhodiola rosea showed the highest xanthine oxidase inhibitory activities. Camellia sinensis, Rhodiola rosea, and Koelreuteria henryi exhibited good tyrosinase inhibitory activities and potent anti-lipoxygenase activities. As Koelreuteria henryi had notable significant inhibitory activities towards xanthine oxidase, tyrosinase, and lipoxygenase, it was further tested with tyrosinase and HPLC-DAD. The results from this part of the study revealed that the more powerful the antioxidant capability of the extracted component, the greater the decrease in peak height obtained after reacting with tyrosinase. Additional studies are warranted to further characterize the compounds responsible for the antioxidant properties of the examined extracts.

Crystallization and preliminary X-ray analysis of Aspergillus oryzae catechol oxidase

International Nuclear Information System (INIS)

Kaljunen, Heidi; Gasparetti, Chiara; Kruus, Kristiina; Rouvinen, Juha; Hakulinen, Nina

2011-01-01

Catechol oxidase from A. oryzae was crystallized by the hanging-drop vapour-diffusion method. Catechol oxidase is an enzyme that catalyzes the oxidation of o-diphenols to the corresponding o-quinones. It is a copper-containing enzyme with a binuclear copper active site. Here, the crystallization and multiple-wavelength anomalous dispersion data collection of catechol oxidase from the mould fungus Aspergillus oryzae are described. During the purification, three forms of the enzyme (39.3, 40.5 and 44.3 kDa) were obtained. A mixture of these three forms was initially crystallized and gave crystals that diffracted to 2.5 Å resolution and belonged to space group P3 2 21, with unit-cell parameters a = b = 118.9, c = 84.5 Å, α = β = 90, γ = 120°. A preparation containing only the shorter form (39.3 kDa) produced crystals that diffracted to 2.9 Å resolution and belonged to space group P2 1 2 1 2 1 , with unit-cell parameters a = 51.8, b = 95.3, c = 139.5 Å, α = β = γ = 90°

In vitro oxidation of indoleacetic acid by soluble auxin-oxidases and peroxidases from maize roots

International Nuclear Information System (INIS)

Beffa, R.; Martin, H.V.; Pilet, P.E.

1990-01-01

Soluble auxin-oxidases were extracted from Zea mays L. cv LG11 apical root segments and partially separated from peroxidases (EC 1.11.1.7) by size-exclusion chromatography. Auxin-oxidases were resolved into one main peak corresponding to a molecular mass of 32.5 kilodaltons and a minor peak at 54.5 kilodaltons. Peroxidases were separated into at least four peaks, with molecular masses from 32.5 to 78 kilodaltons. In vitro activity of indoleacetic acid-oxidases was dependent on the presence of MnCl 2 and p-coumaric acid. Compound(s) present in the crude extract and several synthetic auxin transport inhibitors (including 2,3,5-triiodobenzoic acid and N-1-naphthylphthalamic acid) inhibited auxin-oxidase activity, but had no effect on peroxidases. The products resulting from the in vitro enzymatic oxidation of [ 3 H]indoleacetic acid were separated by HPLC and the major metabolite was found to cochromatograph with indol-3yl-methanol

Multiple controls affect arsenite oxidase gene expression in Herminiimonas arsenicoxydans

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Coppée Jean-Yves

2010-02-01

Full Text Available Abstract Background Both the speciation and toxicity of arsenic are affected by bacterial transformations, i.e. oxidation, reduction or methylation. These transformations have a major impact on environmental contamination and more particularly on arsenic contamination of drinking water. Herminiimonas arsenicoxydans has been isolated from an arsenic- contaminated environment and has developed various mechanisms for coping with arsenic, including the oxidation of As(III to As(V as a detoxification mechanism. Results In the present study, a differential transcriptome analysis was used to identify genes, including arsenite oxidase encoding genes, involved in the response of H. arsenicoxydans to As(III. To get insight into the molecular mechanisms of this enzyme activity, a Tn5 transposon mutagenesis was performed. Transposon insertions resulting in a lack of arsenite oxidase activity disrupted aoxR and aoxS genes, showing that the aox operon transcription is regulated by the AoxRS two-component system. Remarkably, transposon insertions were also identified in rpoN coding for the alternative N sigma factor (σ54 of RNA polymerase and in dnaJ coding for the Hsp70 co-chaperone. Western blotting with anti-AoxB antibodies and quantitative RT-PCR experiments allowed us to demonstrate that the rpoN and dnaJ gene products are involved in the control of arsenite oxidase gene expression. Finally, the transcriptional start site of the aoxAB operon was determined using rapid amplification of cDNA ends (RACE and a putative -12/-24 σ54-dependent promoter motif was identified upstream of aoxAB coding sequences. Conclusion These results reveal the existence of novel molecular regulatory processes governing arsenite oxidase expression in H. arsenicoxydans. These data are summarized in a model that functionally integrates arsenite oxidation in the adaptive response to As(III in this microorganism.

IAA oxidase activity in relation to adventitious root formation on stem cuttings of some forest tree species. [Salix tetrasperma, Populus Robusta, Hibiscus rosa-sinensis, Eucalyptus citriodora

Energy Technology Data Exchange (ETDEWEB)

Bansal, M.P.; Nanda, K.K.

1981-01-01

In rooting tests with stem cuttings, IAA oxidase activity was found to be very high in Salix tetrasperma and Populus 'Robusta' both of which rooted profusely, less in Hibiscus rosa-sinensis which rooted but weakly and insignificant in Eucalyptus citriodora, which did not root at all. Proteins extracted from the stem cuttings of E. citriodora inhibited IAA oxidase activity, and also root formation on hypocotyl cuttings of Phaseolus mungo.

Comparative study between dimethyl sulfoxide (dmso), allopurinol and urate oxidase administration in nephrotoxic rats induced with

International Nuclear Information System (INIS)

Heibashy, M.I.A.; El-Nahla, A.M.; Ibrahim, A.I.; Saleh, Sh.Y.A.

2010-01-01

This study was conducted to show whether DMSO, allopurinol and urate oxidase could offer ameliorating effects against abnormal alterations in kidney function tests in gentamicin (GM) induced nephrotoxic rats . Two experiments were carried out, the first one showed that daily injection of 80 mg GM/kg b. wt interapertonealy (I.P) for two weeks induced acute renal failure indicated by significant elevation in serum urea, creatinine, uric acid, potassium, inorganic phosphorus, TBARS and PTH and a significant decline in serum sodium, total and ionized calcium when compared with their corresponding values in saline injected rats. In the second experiment, comparisons were made between GM induced nephrotoxic rats and other nephrotoxic groups received daily pf I.P injection of DMSO (4 ml/kg b.wt), allopurinol (1.5 mg/100 g b.wt) and urate oxidase (10 mg/100 g b.wt) for 30 days after the incidence of nephrotoxicity. At all intervals, 10,20 and 30 days; serum urea, creatinine, uric acid, potassium, inorganic phosphorus, TBARS and PTH in DMSO, allopurinol and urate oxidase treated groups exhibited significant reduction than nephrotoxic untreated rats. During the same intervals, the levels of serum total and ionized calcium showed an opposite trend. serum sodium level did not show any significant difference between all treated groups except after 20 days , it was increased significantly in urate oxidase treated group and after 30 days in both allopurinol and urate oxidase treated groups. in term time intervals, a significant correction was recorded on the level of most measured parameters. in nephritic rats, the administration of DMSO, allopurinol or urate oxidase led to a significant amelioration effects in the kidney function tests and urate oxidase was the best protective.

A Xylenol Orange-Based Screening Assay for the Substrate Specificity of Flavin-Dependent para-Phenol Oxidases

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Tom A. Ewing

2018-01-01

Full Text Available Vanillyl alcohol oxidase (VAO and eugenol oxidase (EUGO are flavin-dependent enzymes that catalyse the oxidation of para-substituted phenols. This makes them potentially interesting biocatalysts for the conversion of lignin-derived aromatic monomers to value-added compounds. To facilitate their biocatalytic exploitation, it is important to develop methods by which variants of the enzymes can be rapidly screened for increased activity towards substrates of interest. Here, we present the development of a screening assay for the substrate specificity of para-phenol oxidases based on the detection of hydrogen peroxide using the ferric-xylenol orange complex method. The assay was used to screen the activity of VAO and EUGO towards a set of twenty-four potential substrates. This led to the identification of 4-cyclopentylphenol as a new substrate of VAO and EUGO and 4-cyclohexylphenol as a new substrate of VAO. Screening of a small library of VAO and EUGO active-site variants for alterations in their substrate specificity led to the identification of a VAO variant (T457Q with increased activity towards vanillyl alcohol (4-hydroxy-3-methoxybenzyl alcohol and a EUGO variant (V436I with increased activity towards chavicol (4-allylphenol and 4-cyclopentylphenol. This assay provides a quick and efficient method to screen the substrate specificity of para-phenol oxidases, facilitating the enzyme engineering of known para-phenol oxidases and the evaluation of the substrate specificity of novel para-phenol oxidases.

A Xylenol Orange-Based Screening Assay for the Substrate Specificity of Flavin-Dependent para-Phenol Oxidases.

Science.gov (United States)

Ewing, Tom A; van Noord, Aster; Paul, Caroline E; van Berkel, Willem J H

2018-01-14

Vanillyl alcohol oxidase (VAO) and eugenol oxidase (EUGO) are flavin-dependent enzymes that catalyse the oxidation of para -substituted phenols. This makes them potentially interesting biocatalysts for the conversion of lignin-derived aromatic monomers to value-added compounds. To facilitate their biocatalytic exploitation, it is important to develop methods by which variants of the enzymes can be rapidly screened for increased activity towards substrates of interest. Here, we present the development of a screening assay for the substrate specificity of para -phenol oxidases based on the detection of hydrogen peroxide using the ferric-xylenol orange complex method. The assay was used to screen the activity of VAO and EUGO towards a set of twenty-four potential substrates. This led to the identification of 4-cyclopentylphenol as a new substrate of VAO and EUGO and 4-cyclohexylphenol as a new substrate of VAO. Screening of a small library of VAO and EUGO active-site variants for alterations in their substrate specificity led to the identification of a VAO variant (T457Q) with increased activity towards vanillyl alcohol (4-hydroxy-3-methoxybenzyl alcohol) and a EUGO variant (V436I) with increased activity towards chavicol (4-allylphenol) and 4-cyclopentylphenol. This assay provides a quick and efficient method to screen the substrate specificity of para -phenol oxidases, facilitating the enzyme engineering of known para- phenol oxidases and the evaluation of the substrate specificity of novel para -phenol oxidases.

A Plastid Terminal Oxidase Associated with Carotenoid Desaturation during Chromoplast Differentiation1

Science.gov (United States)

Josse, Eve-Marie; Simkin, Andrew J.; Gaffé, Joël; Labouré, Anne-Marie; Kuntz, Marcel; Carol, Pierre

2000-01-01

The Arabidopsis IMMUTANS gene encodes a plastid homolog of the mitochondrial alternative oxidase, which is associated with phytoene desaturation. Upon expression in Escherichia coli, this protein confers a detectable cyanide-resistant electron transport to isolated membranes. In this assay this activity is sensitive to n-propyl-gallate, an inhibitor of the alternative oxidase. This protein appears to be a plastid terminal oxidase (PTOX) that is functionally equivalent to a quinol:oxygen oxidoreductase. This protein was immunodetected in achlorophyllous pepper (Capsicum annuum) chromoplast membranes, and a corresponding cDNA was cloned from pepper and tomato (Lycopersicum esculentum) fruits. Genomic analysis suggests the presence of a single gene in these organisms, the expression of which parallels phytoene desaturase and ζ-carotene desaturase gene expression during fruit ripening. Furthermore, this PTOX gene is impaired in the tomato ghost mutant, which accumulates phytoene in leaves and fruits. These data show that PTOX also participates in carotenoid desaturation in chromoplasts in addition to its role during early chloroplast development. PMID:10938359

Carbon fiber microelectrodes modified with carbon nanotubes as a new support for immobilization of glucose oxidase

International Nuclear Information System (INIS)

Wen, H.; Nallathambi, V.; Chakraborty, D.; Barton, S.C.

2011-01-01

Carboxylated carbon nanotubes were coated onto carbon microfiber electrodes to create a micron-scale bioelectrode. This material has a high surface area and can serve as a support for immobilization of enzymes such as glucose oxidase. A typical carbon nanotube loading of 13 μg cm -1 yields a coating thickness of 17 μm and a 2000-fold increase in surface capacitance. The modified electrode was further coated with a biocatalytic hydrogel composed of a conductive redox polymer, glucose oxidase, and a crosslinker to create a glucose bioelectrode. The current density on oxidation of glucose is 16.6 mA cm-2 at 0.5 V (vs. Ag/AgCl) in oxygen-free glucose solution. We consider this approach to be useful for designing and characterizing surface treatments for carbon mats and papers by mimicking their local microenvironment. (author)

Cytochrome oxidase as an indicator of ice storage and frozen storage

DEFF Research Database (Denmark)

Godiksen, Helene; Jessen, Flemming

2001-01-01

in different cods was 21%, and the coefficient of variation of different analyses on the same homogenate was 5%. It was shown that ice storage of muscle samples before they were frozen and thawed resulted in a major freezing-induced activation of cytochrome oxidase activity. The enzyme may therefore be used...... as an indicator of frozen fish to determine if the fish has been stored on ice before freezing. Cytochrome oxidase activity showed also potential as an indicator of frozen storage, as it was possible to distinguish between the frozen storage temperatures -9, -20, and -40 degreesC....

Dual inhibitors of cholinesterases and monoamine oxidases for Alzheimer's disease.

Science.gov (United States)

Knez, Damijan; Sova, Matej; Košak, Urban; Gobec, Stanislav

2017-05-01

Accumulating evidence indicates a solid relationship between several enzymes and Alzheimer's disease. Cholinesterases and monoamine oxidases are closely associated with the disease symptomatology and progression and have been tackled simultaneously using several multifunctional ligands. This design strategy offers great chances to alter the course of Alzheimer's disease, in addition to alleviation of the symptoms. More than 15 years of research has led to the identification of various dual cholinesterase/monoamine oxidase inhibitors, while some showing positive outcomes in clinical trials, thus giving rise to additional research efforts in the field. The aim of this review is to provide an update on the novel dual inhibitors identified recently and to shed light on their therapeutic potential.

On the Possibility of Uphill Intramolecular Electron Transfer in Multicopper Oxidases: Electrochemical and Quantum Chemical Study of Bilirubin Oxidase

Czech Academy of Sciences Publication Activity Database

Shleev, S.; Andoralov, V.; Falk, M.; Reimann, C. T.; Ruzgas, T.; Srnec, Martin; Ryde, U.; Rulíšek, Lubomír

2012-01-01

Roč. 24, č. 7 (2012), s. 1524-1540 ISSN 1040-0397 Grant - others:7th Framework Program(XE) NMP4-SL-2009-229255 Institutional research plan: CEZ:AV0Z40550506 Keywords : bilirubin oxidase * intramolecular electron transfer * rate-limiting catalytic step * reorganization energy * QM/MM calculations Subject RIV: CF - Physical ; Theoretical Chemistry Impact factor: 2.817, year: 2012

Gene cloning and characterization of NADH oxidase from ...

African Journals Online (AJOL)

The genome search of Thermococcus kodakarensis revealed three open reading frames, Tk0304, Tk1299 and Tk1392 annotated as nicotinamide adenine dinucleotide (NADH) oxidases. This study deals with cloning, and characterization of Tk0304. The gene, composed of 1320 nucleotides, encodes a protein of 439 ...

Mapping the primary structure of copper/topaquinone-containing methylamine oxidase from Aspergillus niger.

Science.gov (United States)

Lenobel, R; Sebela, M; Frébort, I

2005-01-01

The amino acid sequence of methylamine oxidase (MeAO) from the fungus Aspergillus niger was analyzed using mass spectrometry (MS). First, MeAO was characterized by an accurate molar mass of 72.4 kDa of the monomer measured using MALDI-TOF-MS and by a pI value of 5.8 determined by isoelectric focusing. MALDI-TOF-MS revealed a clear peptide mass fingerprint after tryptic digestion, which did not provide any relevant hit when searched against a nonredundant protein database and was different from that of A. niger amine oxidase AO-I. Tandem mass spectrometry with electrospray ionization coupled to liquid chromatography allowed unambiguous reading of six peptide sequences (11-19 amino acids) and seven sequence tags (4-15 amino acids), which were used for MS BLAST homology searching. MeAO was found to be largely homologous to a hypothetical protein AN7641.2 (EMBL/GenBank protein-accession code EAA61827) from Aspergillus nidulans FGSC A4 with a theoretical molar mass of 76.46 kDa and pI 6.14, which belongs to the superfamily of copper amine oxidases. The protein AN7641.2 is only little homologous to the amine oxidase AO-I (32% identity, 49 % similarity).

Monoamine oxidase inhibitors from Gentiana lutea.

Science.gov (United States)

Haraguchi, Hiroyuki; Tanaka, Yasumasa; Kabbash, Amal; Fujioka, Toshihiro; Ishizu, Takashi; Yagi, Akira

2004-08-01

Three monoamine oxidase (MAO) inhibitors were isolated from Gentiana lutea. Their structures were elucidated to be 3-3''linked-(2'-hydroxy-4-O-isoprenylchalcone)-(2'''-hydroxy-4''-O-isoprenyldihydrochalcone) (1), 2-methoxy-3-(1,1'-dimethylallyl)-6a,10a-dihydrobenzo(1,2-c)chroman-6-one and 5-hydroxyflavanone. These compounds, and the hydrolysis product of 1, displayed competitive inhibitory properties against MAO-B which was more effective than MAO-A.

Oxidase catalysis via aerobically generated hypervalent iodine intermediates

Science.gov (United States)

Maity, Asim; Hyun, Sung-Min; Powers, David C.

2018-02-01

The development of sustainable oxidation chemistry demands strategies to harness O2 as a terminal oxidant. Oxidase catalysis, in which O2 serves as a chemical oxidant without necessitating incorporation of oxygen into reaction products, would allow diverse substrate functionalization chemistry to be coupled to O2 reduction. Direct O2 utilization suffers from intrinsic challenges imposed by the triplet ground state of O2 and the disparate electron inventories of four-electron O2 reduction and two-electron substrate oxidation. Here, we generate hypervalent iodine reagents—a broadly useful class of selective two-electron oxidants—from O2. This is achieved by intercepting reactive intermediates of aldehyde autoxidation to aerobically generate hypervalent iodine reagents for a broad array of substrate oxidation reactions. The use of aryl iodides as mediators of aerobic oxidation underpins an oxidase catalysis platform that couples substrate oxidation directly to O2 reduction. We anticipate that aerobically generated hypervalent iodine reagents will expand the scope of aerobic oxidation chemistry in chemical synthesis.

A point mutation of valine-311 to methionine in Bacillus subtilis protoporphyrinogen oxidase does not greatly increase resistance to the diphenyl ether herbicide oxyfluorfen.

Science.gov (United States)

Jeong, Eunjoo; Houn, Thavrak; Kuk, Yongin; Kim, Eun-Seon; Chandru, Hema Kumar; Baik, Myunggi; Back, Kyoungwhan; Guh, Ja-Ock; Han, Oksoo

2003-10-01

In an effort to asses the effect of Val311Met point mutation of Bacillus subtilis protoporphyrinogen oxidase on the resistance to diphenyl ether herbicides, a Val311Met point mutant of B. subtilis protoporphyrinogen oxidase was prepared, heterologously expressed in Escherichia coli, and the purified recombinant Val311Met mutant protoporphyrinogen oxidase was kinetically characterized. The mutant protoporphyrinogen oxidase showed very similar kinetic patterns to wild type protoporphyrinogen oxidase, with slightly decreased activity dependent on pH and the concentrations of NaCl, Tween 20, and imidazole. When oxyfluorfen was used as a competitive inhibitor, the Val311Met mutant protoporphyrinogen oxidase showed an increased inhibition constant about 1.5 times that of wild type protoporphyrinogen oxidase. The marginal increase of the inhibition constant indicates that the Val311Met point mutation in B. subtilis protoporphyrinogen oxidase may not be an important determinant in the mechanism that protects protoporphyrinogen oxidase against diphenyl ether herbicides.

Biodegradation of phenolic compounds with oxidases from sorghum and non-defined mixed bacterium media

International Nuclear Information System (INIS)

Obame, C. E. L.; Savadogo, P. W.; Mamoudou, D. H.; Dembele, R. H.; Traore, A. S.

2009-01-01

The biodegradation of the phenolic compounds is performed using oxidative enzymes, e. g. polyphenol oxidases (PPOs) and peroxidases (POXs). These oxidases displaying a wide spectrum for the oxidation of phenolic compounds were isolated either from sorghum or mixed bacteria. Spectrophotometric methods were used to assess the monophenolase and diphenolase activities of PPOs as well as the hydrogen-dependant oxidation of POXs. (Author)

Biodegradation of phenolic compounds with oxidases from sorghum and non-defined mixed bacterium media

Energy Technology Data Exchange (ETDEWEB)

Obame, C. E. L.; Savadogo, P. W.; Mamoudou, D. H.; Dembele, R. H.; Traore, A. S.

2009-07-01

The biodegradation of the phenolic compounds is performed using oxidative enzymes, e. g. polyphenol oxidases (PPOs) and peroxidases (POXs). These oxidases displaying a wide spectrum for the oxidation of phenolic compounds were isolated either from sorghum or mixed bacteria. Spectrophotometric methods were used to assess the monophenolase and diphenolase activities of PPOs as well as the hydrogen-dependant oxidation of POXs. (Author)

Study on the preparation of immobilized glucose oxidase membrane and its application in clinic analysis

International Nuclear Information System (INIS)

Yu Ye; Cao Jin; Su Zongxian; Chen Zixiong

1990-01-01

The paper deals with the preparation of immobilized glucose oxidase membrane by using two steps irradiation (irradiation gratfting, irradiation entrapping). Some properties of membrane were discussed. The immobilized glucose oxidase membrane with oxygen electrode and oxygen analyser can be satisfied with the clinic analysis for the determination of serum glucose

Study on dioxygen reduction by mutational modifications of the hydrogen bond network leading from bulk water to the trinuclear copper center in bilirubin oxidase

International Nuclear Information System (INIS)

Morishita, Hirotoshi; Kurita, Daisuke; Kataoka, Kunishige; Sakurai, Takeshi

2014-01-01

Highlights: • Proton transport pathway in bilirubin oxidase was mutated. • Two intermediates in the dioxygen reduction steps were trapped and characterized. • A specific glutamate for dioxygen reduction by multicopper oxidases was identified. - Abstract: The hydrogen bond network leading from bulk water to the trinuclear copper center in bilirubin oxidase is constructed with Glu463 and water molecules to transport protons for the four-electron reduction of dioxygen. Substitutions of Glu463 with Gln or Ala were attributed to virtually complete loss or significant reduction in enzymatic activities due to an inhibition of the proton transfer steps to dioxygen. The single turnover reaction of the Glu463Gln mutant afforded the highly magnetically interacted intermediate II (native intermediate) with a broad g = 1.96 electron paramagnetic resonance signal detectable at cryogenic temperatures. Reactions of the double mutants, Cys457Ser/Glu463Gln and Cys457Ser/Glu463Ala afforded the intermediate I (peroxide intermediate) because the type I copper center to donate the fourth electron to dioxygen was vacant in addition to the interference of proton transport due to the mutation at Glu463. The intermediate I gave no electron paramagnetic resonance signal, but the type II copper signal became detectable with the decay of the intermediate I. Structural and functional similarities between multicopper oxidases are discussed based on the present mutation at Glu463 in bilirubin oxidase

Study on dioxygen reduction by mutational modifications of the hydrogen bond network leading from bulk water to the trinuclear copper center in bilirubin oxidase

Energy Technology Data Exchange (ETDEWEB)

Morishita, Hirotoshi; Kurita, Daisuke; Kataoka, Kunishige; Sakurai, Takeshi, E-mail: tsakurai@se.kanazawa-u.ac.jp

2014-07-18

Highlights: • Proton transport pathway in bilirubin oxidase was mutated. • Two intermediates in the dioxygen reduction steps were trapped and characterized. • A specific glutamate for dioxygen reduction by multicopper oxidases was identified. - Abstract: The hydrogen bond network leading from bulk water to the trinuclear copper center in bilirubin oxidase is constructed with Glu463 and water molecules to transport protons for the four-electron reduction of dioxygen. Substitutions of Glu463 with Gln or Ala were attributed to virtually complete loss or significant reduction in enzymatic activities due to an inhibition of the proton transfer steps to dioxygen. The single turnover reaction of the Glu463Gln mutant afforded the highly magnetically interacted intermediate II (native intermediate) with a broad g = 1.96 electron paramagnetic resonance signal detectable at cryogenic temperatures. Reactions of the double mutants, Cys457Ser/Glu463Gln and Cys457Ser/Glu463Ala afforded the intermediate I (peroxide intermediate) because the type I copper center to donate the fourth electron to dioxygen was vacant in addition to the interference of proton transport due to the mutation at Glu463. The intermediate I gave no electron paramagnetic resonance signal, but the type II copper signal became detectable with the decay of the intermediate I. Structural and functional similarities between multicopper oxidases are discussed based on the present mutation at Glu463 in bilirubin oxidase.

Activation of NADPH oxidase mediates increased endoplasmic reticulum stress and left ventricular remodeling after myocardial infarction in rabbits.

Science.gov (United States)

Li, Bao; Tian, Jing; Sun, Yi; Xu, Tao-Rui; Chi, Rui-Fang; Zhang, Xiao-Li; Hu, Xin-Ling; Zhang, Yue-An; Qin, Fu-Zhong; Zhang, Wei-Fang

2015-05-01

Nicotinamide adenine dinucleotide 3-phosphate (NADPH) oxidase activity and endoplasmic reticulum (ER) stress are increased after myocardial infarction (MI). In this study, we proposed to test whether activation of the NADPH oxidase in the remote non-infarcted myocardium mediates ER stress and left ventricular (LV) remodeling after MI. Rabbits with MI or sham operation were randomly assigned to orally receive an NADPH oxidase inhibitor apocynin or placebo for 30 days. The agents were administered beginning at 1 week after surgery. MI rabbits exhibited decreases in LV fractional shortening, LV ejection fraction and the first derivative of the LV pressure rise, which were abolished by apocynin treatment. NADPH oxidase Nox2 protein and mRNA expressions were increased in the remote non-infarcted myocardium after MI. Immunolabeling further revealed that Nox2 was increased in cardiac myocytes in the remote myocardium. The apocynin treatment prevented increases in the Nox2 expression, NADPH oxidase activity, oxidative stress, myocyte apoptosis and GRP78, CHOP and cleaved caspase 12 protein expression in the remote myocardium. The apocynin treatment also attenuated increases in myocyte diameter and cardiac fibrosis. In cultured H9C2 cardiomyocytes exposed to angiotensin II, an important stimulus for post-MI remodeling, Nox2 knockdown with siRNA significantly inhibited angiotensin II-induced NADPH oxidase activation, reactive oxygen species and GRP78 and CHOP protein expression. We conclude that NADPH oxidase inhibition attenuates increased ER stress in the remote non-infarcted myocardium and LV remodeling late after MI in rabbits. These findings suggest that the activation of NADPH oxidase in the remote non-infarcted myocardium mediates increased ER stress, contributing to myocyte apoptosis and LV remodeling after MI. Copyright © 2015 Elsevier B.V. All rights reserved.

Characterization of water-forming NADH oxidases for co-factor regeneration

DEFF Research Database (Denmark)

Rehn, Gustav; Pedersen, Asbjørn Toftgaard; J. Charnock, Simon

an environmentaland economic perspective [1]. Alcohol dehydrogenases (ADH) offer one such alternative. However, the reaction requires the oxidized nicotinamide co-factor (NAD+) that must be recycled due to its high cost contribution. One regeneration method that offers certain advantages is the oxidation of NADH......Traditional chemical methods for alcohol oxidation are often associated with issues such as high consumption of expensive oxidizing agents, generation of metal waste and the use of environmentally undesirable organic solvents. Developing green, selective catalysts is therefore important from...... using water forming NADH oxidases (NOX-2). The implementation of the ADH/NOX system for alcohol oxidation, however, requires consideration of several different issues. Enzyme activity and stability at relevant pH and temperature conditions, but also the tolerance to the substrates and products present...

Random mutagenesis of aspergillus niger and process optimization for enhanced production of glucose oxidase

International Nuclear Information System (INIS)

Haq, I.; Nawaz, A.; Mukhtar, A.N.H.; Mansoor, H.M.Z.; Ameer, S.M.

2014-01-01

The study deals with the improvement of wild strain Aspergillus niger IIB-31 through random mutagenesis using chemical mutagens. The main aim of the work was to enhance the glucose oxidase (GOX) yield of wild strain (24.57+-0.01 U/g of cell mass) through random mutagenesis and process optimization. The wild strain of Aspergillus niger IIB-31 was treated with chemical mutagens such as Ethyl methane sulphonate (EMS) and nitrous acid for this purpose. Mutagen treated 98 variants indicating the positive results were picked and screened for the glucose oxidase production using submerged fermentation. EMS treated E45 mutant strain gave the highest glucose oxidase production (69.47 + 0.01 U/g of cell mass), which was approximately 3-folds greater than the wild strain IIB-31. The preliminary cultural conditions for the production of glucose oxidase using submerged fermentation from strain E45 were also optimized. The highest yield of GOD was obtained using 8% glucose as carbon and 0.3% peptone as nitrogen source at a medium pH of 7.0 after an incubation period of 72 hrs at 30 degree. (author)

Presteady-state and steady-state kinetic properties of human cytochrome c oxidase. Identification of rate-limiting steps in mammalian cytochrome c oxidase

NARCIS (Netherlands)

van Kuilenburg, A. B.; Gorren, A. C.; Dekker, H. L.; Nieboer, P.; van Gelder, B. F.; Muijsers, A. O.

1992-01-01

Human cytochrome c oxidase was purified in a fully active form from heart and skeletal muscle. The enzyme was selectively solubilised with octylglucoside and KCl from submitochondrial particles followed by ammonium sulphate fractionation. The presteady-state and steady-state kinetic properties of

Molecular evolution of the polyamine oxidase gene family in Metazoa

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Polticelli Fabio

2012-06-01

Full Text Available Abstract Background Polyamine oxidase enzymes catalyze the oxidation of polyamines and acetylpolyamines. Since polyamines are basic regulators of cell growth and proliferation, their homeostasis is crucial for cell life. Members of the polyamine oxidase gene family have been identified in a wide variety of animals, including vertebrates, arthropodes, nematodes, placozoa, as well as in plants and fungi. Polyamine oxidases (PAOs from yeast can oxidize spermine, N1-acetylspermine, and N1-acetylspermidine, however, in vertebrates two different enzymes, namely spermine oxidase (SMO and acetylpolyamine oxidase (APAO, specifically catalyze the oxidation of spermine, and N1-acetylspermine/N1-acetylspermidine, respectively. Little is known about the molecular evolutionary history of these enzymes. However, since the yeast PAO is able to catalyze the oxidation of both acetylated and non acetylated polyamines, and in vertebrates these functions are addressed by two specialized polyamine oxidase subfamilies (APAO and SMO, it can be hypothesized an ancestral reference for the former enzyme from which the latter would have been derived. Results We analysed 36 SMO, 26 APAO, and 14 PAO homologue protein sequences from 54 taxa including various vertebrates and invertebrates. The analysis of the full-length sequences and the principal domains of vertebrate and invertebrate PAOs yielded consensus primary protein sequences for vertebrate SMOs and APAOs, and invertebrate PAOs. This analysis, coupled to molecular modeling techniques, also unveiled sequence regions that confer specific structural and functional properties, including substrate specificity, by the different PAO subfamilies. Molecular phylogenetic trees revealed a basal position of all the invertebrates PAO enzymes relative to vertebrate SMOs and APAOs. PAOs from insects constitute a monophyletic clade. Two PAO variants sampled in the amphioxus are basal to the dichotomy between two well supported

Lower Growth Temperature Increases Alternative Pathway Capacity and Alternative Oxidase Protein in Tobacco 1

Science.gov (United States)

Vanlerberghe, Greg C.; McIntosh, Lee

1992-01-01

Suspension cells of NT1 tobacco (Nicotiana tabacum L. cv bright yellow) have been used to study the effect of growth temperature on the CN-resistant, salicylhydroxamic acid-sensitive alternative pathway of respiration. Mitochondria isolated from cells maintained at 30°C had a low capacity to oxidize succinate via the alternative pathway, whereas mitochondria isolated from cells 24 h after transfer to 18°C displayed, on average, a 5-fold increase in this capacity (from 7 to 32 nanoatoms oxygen per milligram protein per minute). This represented an increase in alternative pathway capacity from 18 to 45% of the total capacity of electron transport. This increased capacity was lost upon transfer of cells back to 30°C. A monoclonal antibody to the terminal oxidase of the alternative pathway (the alternative oxidase) from Sauromatum guttatum (T.E. Elthon, R.L. Nickels, L. McIntosh [1989] Plant Physiology 89: 1311-1317) recognized a 35-kilodalton mitochondrial protein in tobacco. There was an excellent correlation between the capacity of the alternative path in isolated tobacco mitochondria and the levels of this 35-kilodalton alternative oxidase protein. Cycloheximide could inhibit both the increased level of the 35-kilodalton alternative oxidase protein and the increased alternative pathway capacity normally seen upon transfer to 18°C. We conclude that transfer of tobacco cells to the lower temperature increases the capacity of the alternative pathway due, at least in part, to de novo synthesis of the 35-kilodalton alternative oxidase protein. Images Figure 3 Figure 4 PMID:16652932

Characterization of a Highly Thermostable and Organic Solvent-Tolerant Copper-Containing Polyphenol Oxidase with Dye-Decolorizing Ability from Kurthia huakuii LAM0618T.

Directory of Open Access Journals (Sweden)

Xiang Guo

Full Text Available Laccases are green biocatalysts that possess attractive advantages for the treatment of resistant environmental pollutants and dye effluents. A putative laccase-like gene, laclK, encoding a protein of 29.3 kDa and belonging to the Cu-oxidase_4 superfamily, was cloned and overexpressed in Escherichia coli. The purified recombinant protein LaclK (LaclK was able to oxidize typical laccase substrates such as 2,6-dimethoxyphenol and l-dopamine. The characteristic adsorption maximums of typical laccases at 330 nm and 610 nm were not detected for LaclK. Cu2+ was essential for substrate oxidation, but the ratio of copper atoms/molecule of LaclK was determined to only be 1:1. Notably, the optimal temperature of LaclK was 85°C with 2,6-dimethoxyphenol as substrates, and the half-life approximately 3 days at 80°C. Furthermore, 10% (v/v organic solvents (methanol, ethanol, isopropyl alcohol, butyl alcohol, Triton x-100 or dimethyl sulfoxide could promote enzymatic activity. LaclK exhibited wide-spectrum decolorization ability towards triphenylmethane dyes, azo dyes and aromatic dyes, decolorizing 92% and 94% of Victoria Blue B (25 μM and Ethyl Violet (25 μM, respectively, at a concentration of 60 U/L after 1 h of incubation at 60°C. Overall, we characterized a novel thermostable and organic solvent-tolerant copper-containing polyphenol oxidase possessing dye-decolorizing ability. These unusual properties make LaclK an alternative for industrial applications, particularly processes that require high-temperature conditions.

Interferon gamma/NADPH oxidase defence system in immunity and cancer

Czech Academy of Sciences Publication Activity Database

Hodný, Zdeněk; Reiniš, Milan; Hubáčková, Soňa; Vašicová, Pavla; Bartek, Jiří

-, 01 Sep (2015) ISSN 2162-4011 Institutional support: RVO:68378050 ; RVO:61388971 Keywords : IFNγ * NADPH oxidase * immunity * cancer Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.266, year: 2014

Distinguishing the Roles of Thylakoid Respiratory Terminal Oxidases in the Cyanobacterium Synechocystis sp. PCC 6803.

Science.gov (United States)

Ermakova, Maria; Huokko, Tuomas; Richaud, Pierre; Bersanini, Luca; Howe, Christopher J; Lea-Smith, David J; Peltier, Gilles; Allahverdiyeva, Yagut

2016-06-01

Various oxygen-utilizing electron sinks, including the soluble flavodiiron proteins (Flv1/3), and the membrane-localized respiratory terminal oxidases (RTOs), cytochrome c oxidase (Cox) and cytochrome bd quinol oxidase (Cyd), are present in the photosynthetic electron transfer chain of Synechocystis sp. PCC 6803. However, the role of individual RTOs and their relative importance compared with other electron sinks are poorly understood, particularly under light. Via membrane inlet mass spectrometry gas exchange, chlorophyll a fluorescence, P700 analysis, and inhibitor treatment of the wild type and various mutants deficient in RTOs, Flv1/3, and photosystem I, we investigated the contribution of these complexes to the alleviation of excess electrons in the photosynthetic chain. To our knowledge, for the first time, we demonstrated the activity of Cyd in oxygen uptake under light, although it was detected only upon inhibition of electron transfer at the cytochrome b6f site and in ∆flv1/3 under fluctuating light conditions, where linear electron transfer was drastically inhibited due to impaired photosystem I activity. Cox is mostly responsible for dark respiration and competes with P700 for electrons under high light. Only the ∆cox/cyd double mutant, but not single mutants, demonstrated a highly reduced plastoquinone pool in darkness and impaired gross oxygen evolution under light, indicating that thylakoid-based RTOs are able to compensate partially for each other. Thus, both electron sinks contribute to the alleviation of excess electrons under illumination: RTOs continue to function under light, operating on slower time ranges and on a limited scale, whereas Flv1/3 responds rapidly as a light-induced component and has greater capacity. © 2016 American Society of Plant Biologists. All Rights Reserved.

Mitochondrial terminal alternative oxidase and its enhancement by thermal stress in the coral symbiont Symbiodinium

Science.gov (United States)

Oakley, Clinton A.; Hopkinson, Brian M.; Schmidt, Gregory W.

2014-06-01

A terminal electron acceptor alternative to mitochondrial cytochrome c oxidase (COX), mitochondrial alternative oxidase (AOX), is ubiquitous in higher plants and represented in nearly every algal taxon but is poorly documented in dinoflagellates. AOX competes for electrons with the conventional COX and has been hypothesized to function as a means of reducing oxidative stress in mitochondria, as well as a potential mechanism for ameliorating thermal and other physiological stressors. Here, the presence of an active AOX in cultured Symbiodinium was assayed by the response of oxygen consumption to the AOX inhibitor salicylhydroxamic acid (SHAM) and the COX inhibitor cyanide (CN). CN-insensitive, SHAM-sensitive oxygen consumption was found to account for a large portion (26 %) of Symbiodinium dark respiration and is consistent with high levels of AOX activity. This experimental evidence of the existence of a previously unreported terminal oxidase was further corroborated by analysis of publicly available Symbiodinium transcriptome data. The potential for enhanced AOX expression to play a compensatory role in mediating thermal stress was supported by inhibitor assays of cultured Symbiodinium at low (18 °C), moderate (26 °C), and high (32 °C) temperature conditions. Maximum capacity of the putative AOX pathway as a proportion of total dark oxygen consumption was found to increase from 26 % at 26 °C to 45 % and 53 % at 18 °C and 32 °C, respectively, when cells were acclimated to the treatment temperatures. Cells assayed at 18 and 32 °C without acclimation exhibited either the same or lower AOX capacity as controls, suggesting that the AOX protein is upregulated under temperature stress. The physiological implications for the presence of AOX in the coral/algal symbiosis and its potential role in response to many forms of biotic and abiotic stress, particularly oxidative stress, are discussed.

Erv2p: characterization of the redox behavior of a yeast sulfhydryl oxidase

DEFF Research Database (Denmark)

Wang, Wenzhong; Winther, Jakob R; Thorpe, Colin

2007-01-01

centers that facilitate the transfer of reducing equivalents from the dithiol substrates of these oxidases to the isoalloxazine ring where the reaction with molecular oxygen occurs. The present study examines yeast Erv2p and compares the redox behavior of this ER luminal protein with the augmenter...... and with unfolded proteins. Rapid reaction studies confirm that reduction of the flavin center of Erv2p is rate-limiting during turnover with molecular oxygen. This comparison of the redox properties between members of the ERV/ALR family of sulfhydryl oxidases provides insights into their likely roles in oxidative......The FAD prosthetic group of the ERV/ALR family of sulfhydryl oxidases is housed at the mouth of a 4-helix bundle and communicates with a pair of juxtaposed cysteine residues that form the proximal redox active disulfide. Most of these enzymes have one or more additional distal disulfide redox...

Crystallization of carbohydrate oxidase from Microdochium nivale

International Nuclear Information System (INIS)

Dušková, Jarmila; Dohnálek, Jan; Skálová, Tereza; Østergaard, Lars Henrik; Fuglsang, Claus Crone; Kolenko, Petr; Štěpánková, Andrea; Hašek, Jindřich

2009-01-01

Industrially used carbohydrate oxidase was successfully crystallized in several forms, diffraction data suitable for structural analysis were collected. Microdochium nivale carbohydrate oxidase was produced by heterologous recombinant expression in Aspergillus oryzae, purified and crystallized. The enzyme crystallizes with varying crystal morphologies depending on the crystallization conditions. Several different crystal forms were obtained using the hanging-drop vapour-diffusion method, two of which were used for diffraction measurements. Hexagon-shaped crystals (form I) diffracted to 2.66 Å resolution, with unit-cell parameters a = b = 55.7, c = 610.4 Å and apparent space group P6 2 22. Analysis of the data quality showed almost perfect twinning of the crystals. Attempts to solve the structure by molecular replacement did not give satisfactory results. Recently, clusters of rod-shaped crystals (form II) were grown in a solution containing PEG MME 550. These crystals belonged to the monoclinic system C2, with unit-cell parameters a = 132.9, b = 56.6, c = 86.5 Å, β = 95.7°. Data sets were collected to a resolution of 2.4 Å. The structure was solved by the molecular-replacement method. Model refinement is currently in progress

Carbon Nanotube Modified Screen Printed Electrodes: Pyranose Oxidase Immobilization Platform for Amperometric Enzyme Sensors

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Dilek ODACI DEMIRKOL

2017-03-01

Full Text Available Here, a novel enzymatic biosensor was developed using multiwalled carbon nanotube including screen printed electrodes (MWCNT-SPE. Pyranose oxidase (PyOx was immobilized on the electrode surface by way of gelatin membrane and then cross-linked using glutaraldehyde. Glucose was detected at -0.7 V (vs. Ag/AgCl by watching consumed oxygen in enzymatic reaction after addition substrate. After optimization of pH and enzyme loading, the linearity was found in the range of 0.1–1.0 mM of glucose. After that, the effect of MCNT on the current was tested. Also the enzymatic biosensor including glucose oxidase instead of pyranose oxidase was prepared and the biosensor response followed for glucose. Furthermore, this system was tested for glucose analysis in soft drinks.

Crosstalk between HDAC6 and Nox2-based NADPH oxidase mediates HIV-1 Tat-induced pro-inflammatory responses in astrocytes

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Gi Soo Youn

2017-08-01

Full Text Available Histone deacetylase 6 (HDAC6 likely is important in inflammatory diseases. However, how HDAC6 exerts its effect on inflammatory processes remains unclear. HIV-1 transactivator of transcription (Tat activates NADPH oxidase resulting in generation of reactive oxygen species (ROS, leading to extensive neuro-inflammation in the central nervous system. We investigated the correlation of HDAC6 and NADPH oxidase in HIV-1 Tat-stimulated astrocytes. HDAC6 knockdown attenuated HIV-1 Tat-induced ROS generation and NADPH oxidase activation. HDAC6 knockdown suppressed HIV-1 Tat-induced expression of NADPH oxidase subunits, such as Nox2, p47phox, and p22phox. Specific inhibition of HDAC6 using tubastatin A suppressed HIV-1 Tat-induced ROS generation and activation of NADPH oxidase. N-acetyl cysteine, diphenyl iodonium, and apocynin suppressed HIV-1 Tat-induced expression of HDAC6 and the pro-inflammatory chemokines CCL2, CXCL8, and CXCL10. Nox2 knockdown attenuated HIV-1 Tat-induced HDAC6 expression and subsequent expression of chemokines. The collective results point to the potential crosstalk between HDAC6 and NADPH oxidase, which could be a combined therapeutic target for relief of HIV-1 Tat-mediated neuro-inflammation. Keywords: HIV-1 Tat, HDAC6, NADPH oxidase, ROS, Inflammation, Astrocytes

MONOAMINE OXIDASE: RADIOTRACER DEVELOPMENT AND HUMAN STUDIES.

Energy Technology Data Exchange (ETDEWEB)

FOWLER,J.S.; LOGAN,J.; VOLKOW,N.D.; WANG,G.J.; MACGREGOR,R.R.; DING,Y.S.

2000-09-28

PET is uniquely capable of providing information on biochemical transformations in the living human body. Although most of the studies of monoamine oxidase (MAO) have focused on measurements in the brain, the role of peripheral MAO as a phase 1 enzyme for the metabolism of drugs and xenobiotics is gaining attention (Strolin Benedetti and Tipton, 1998; Castagnoli et al., 1997.). MAO is well suited for this role because its concentration in organs such as kidneys, liver and digestive organs is high sometimes exceeding that in the brain. Knowledge of the distribution of the MAO subtypes within different organs and different cells is important in determining which substrates (and which drugs and xenobiotics) have access to which MAO subtypes. The highly variable subtype distribution with different species makes human studies even more important. In addition, the deleterious side effects of combining MAO inhibitors with other drugs and with foodstuffs makes it important to know the MAO inhibitory potency of different drugs both in the brain and in peripheral organs (Ulus et al., 2000). Clearly PET can play a role in answering these questions, in drug research and development and in discovering some of the factors which contribute to the highly variable MAO levels in different individuals.

Enhanced production of glucose oxidase from UVmutant of ...

African Journals Online (AJOL)

UV rays were used as mutagen in wild type strain of Aspergillus niger for enhanced production of glucose oxidase. After mutangenization and selection, mutant A. niger strains, resistant to 2-deoxy-Dglucose were obtained. The mutants showed 1.57 and 1.98 fold increase in activities of extra and intra cellular glucose ...

in Escherichia coli with native cholesterol oxidase expressed

African Journals Online (AJOL)

The structure and bio-activity of an endogenous cholesterol oxidase from Brevibacterium sp. was compared to the same enzyme exogenously expressed in Escherichia coli BL21 (DE3) with and without N- or C-terminal his-tags. The different proteins were purified with affinity and subtractive protocols. The specific activity of ...

Novel thidiazuron-derived inhibitors of cytokinin oxidase/dehydrogenase

Czech Academy of Sciences Publication Activity Database

Nisler, Jaroslav; Kopečný, D.; Končitíková, R.; Zatloukal, Marek; Bazgier, Václav; Berka, K.; Zalabák, D.; Briozzo, P.; Strnad, Miroslav; Spíchal, Lukáš

2016-01-01

Roč. 92, 1-2 (2016), s. 235-248 ISSN 0167-4412 R&D Projects: GA MŠk(CZ) LO1204; GA ČR GA15-22322S Institutional support: RVO:61389030 Keywords : Cytokinin oxidase/dehydrogenase * Crystal structure * Molecular docking Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.356, year: 2016

Purification and characterization of amine oxidase from soybean seedlings.

Science.gov (United States)

Vianello, F; Di Paolo, M L; Stevanato, R; Gasparini, R; Rigo, A

1993-11-15

A simple and rapid procedure for purification of soybean seedling amine oxidase is reported. The crude enzyme, obtained by ammonium sulfate fractionation was purified by ion-exchange chromatography on a cellulose phosphate column and batch affinity chromatography on 6-aminohexyl-Sepharose. Cyclohexylamine, a competitive inhibitor, was utilized to elute the enzyme. A homogeneous enzyme was obtained with a yield higher than 25%, the content of minor components being lauryl sulfate-polyacrylamide gel electrophoresis. The enzyme is a dimer and contains two Cu2+ ion per molecule. Its EPR spectrum is typical of Cu2+ in a tetragonal symmetry. The enzyme oxidizes cadaverine at high rate, the specific activity being 4.3 mukat/mg. Molecular, spectroscopic, and kinetic properties of this enzyme are reported.

Thermodynamic properties of copper compounds with oxygen and hydrogen from first principles

Energy Technology Data Exchange (ETDEWEB)

Korzhavyi, P.A.; Johansson, B. (Applied Materials Physics, Dept. of Materials Science and Engineering, Royal Inst. of Technology, Stockholm (Sweden))

2010-02-15

We employ quantum-mechanical calculations (based on density functional theory and linear response theory) in order to test the mechanical and chemical stability of several solid-state configurations of Cu1+, Cu2+, O2-, H1-, and H1+ ions. We begin our analysis with cuprous oxide (Cu{sub 2}O, cuprite structure), cupric oxide (CuO, tenorite structure), and cuprous hydride (CuH, wurtzite and sphalerite structures) whose thermodynamic properties have been studied experimentally. In our calculations, all these compounds are found to be mechanically stable configurations. Their formation energies calculated at T = 0 K (including the energy of zero-point and thermal motion of the ions) and at room temperature are in good agreement with existing thermodynamic data. A search for other possible solid-state conformations of copper, hydrogen, and oxygen ions is then performed. Several candidate structures for solid phases of cuprous oxy-hydride (Cu{sub 4}H{sub 2}O) and cupric hydride (CuH{sub 2}) have been considered but found to be dynamically unstable. Cuprous oxy-hydride is found to be energetically unstable with respect to decomposition onto cuprous oxide and cuprous hydride. Metastability of cuprous hydroxide (CuOH) is established in our calculations. The free energy of CuOH is calculated to be some 50 kJ/mol higher than the average of the free energies of Cu{sub 2}O and water. Thus, cuprite Cu{sub 2}O is the most stable of the examined Cu(I) compounds

Thermodynamic properties of copper compounds with oxygen and hydrogen from first principles

International Nuclear Information System (INIS)

Korzhavyi, P.A.; Johansson, B.

2010-02-01

We employ quantum-mechanical calculations (based on density functional theory and linear response theory) in order to test the mechanical and chemical stability of several solid-state configurations of Cu 1+ , Cu 2+ , O 2- , H 1- , and H 1+ ions. We begin our analysis with cuprous oxide (Cu 2 O, cuprite structure), cupric oxide (CuO, tenorite structure), and cuprous hydride (CuH, wurtzite and sphalerite structures) whose thermodynamic properties have been studied experimentally. In our calculations, all these compounds are found to be mechanically stable configurations. Their formation energies calculated at T = 0 K (including the energy of zero-point and thermal motion of the ions) and at room temperature are in good agreement with existing thermodynamic data. A search for other possible solid-state conformations of copper, hydrogen, and oxygen ions is then performed. Several candidate structures for solid phases of cuprous oxy-hydride (Cu 4 H 2 O) and cupric hydride (CuH 2 ) have been considered but found to be dynamically unstable. Cuprous oxy-hydride is found to be energetically unstable with respect to decomposition onto cuprous oxide and cuprous hydride. Metastability of cuprous hydroxide (CuOH) is established in our calculations. The free energy of CuOH is calculated to be some 50 kJ/mol higher than the average of the free energies of Cu 2 O and water. Thus, cuprite Cu 2 O is the most stable of the examined Cu(I) compounds

Switching an O2 sensitive glucose oxidase bioelectrode into an almost insensitive one by cofactor redesign.

Science.gov (United States)

Tremey, Emilie; Suraniti, Emmanuel; Courjean, Olivier; Gounel, Sébastien; Stines-Chaumeil, Claire; Louerat, Frédéric; Mano, Nicolas

2014-06-04

In the 5-8 mM glucose concentration range, of particular interest for diabetes management, glucose oxidase bioelectrodes are O2 dependent, which decrease their efficiencies. By replacing the natural cofactor of glucose oxidase, we succeeded in turning an O2 sensitive bioelectrode into an almost insensitive one.

Nanoporous gold assembly of glucose oxidase for electrochemical biosensing

DEFF Research Database (Denmark)

Xiao, Xinxin; Ulstrup, Jens; Li, Hui

2014-01-01

Nanoporous gold (NPG) is composed of three-dimensional (3D) bicontinuous nanostructures with large surface area. Nano-channels inside NPG provide an ideal local environment for immobilization of enzyme molecules with expected stabilization of the protein molecules. In this work, glucose oxidase (...

Variations in epidermal cytochrome oxidase activity after local irradiation

International Nuclear Information System (INIS)

Itoiz, M.E.; Rey, B.M. de; Cabrini, R.L.

1982-01-01

Cytochrome oxidase activity was evaluated histochemically as an index of mitochondrial damage after local irradiation with X-rays. It was determined by microphotometry on the tail skin of newly born Wistar rats four days after irradiation with doses ranging from 2 to 16krad. The enzyme activity of the whole epidermis increased after irradiation, the increases being related to the increase in thickness of the epithelium which was observed as a response to irradiation injury. Within the dose range tested, the enzyme concentration (expressed per unit volume of tissue) decreased in relation to the dose applied. At the electron microscopy level, the cytochemical demonstration of cytochrome oxidase revealed an irregular reaction over the cristae, intramitochondrial vacuolization and partial homogenization of the matrix. Positive membrane fragments were seen around lipid droplets. This reaction confirms the mitochondrial origin of these previously observed radiation-induced vacuoles. (author)

NADPH oxidase is involved in regulation of gene expression and ROS overproduction in soybean (Glycine max L. seedlings exposed to cadmium

Directory of Open Access Journals (Sweden)

Jagna Chmielowska-Bąk

2017-06-01

Full Text Available Cadmium-induced oxidative burst is partially mediated by NADPH oxidase. The aim of the present research was to evaluate the role of NADPH oxidase in soybeans’ response to short-term cadmium stress. The application of an NADPH oxidase inhibitor, diphenyleneiodonium chloride (DPI, affected expression of two Cd-inducible genes, encoding DOF1 and MYBZ2 transcription factors. This effect was observed after 3 h of treatment. Interestingly, Cd-dependent increases in NADPH oxidase activity occurred only after a period of time ranging from 6 and 24 h of stress. Stimulation of the enzyme correlated in time with a significant accumulation of reactive oxygen species (ROS. Further analysis revealed that pharmacological inhibition of NADPH oxidase activity during 24 h of Cd stress does not affect Cd uptake, seedling growth, or the level of lipid peroxidation. The role of NADPH oxidase in the response of soybean seedlings to short-term Cd exposure is discussed.

Development of Galactose Biosensor Based on Functionalized ZnO Nanorods with Galactose Oxidase

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K. Khun

2012-01-01

Full Text Available The fabrication of galactose biosensor based on functionalised ZnO nanorods is described. The galactose biosensor was developed by immobilizing galactose oxidase on ZnO nanorods in conjunction with glutaraldehyde as a cross-linker molecule. The IRAS study provided evidence for the interaction of galactose oxidase with the surface of ZnO nanorods. The electromotive force (EMF response of the galactose biosensor was measured by potentiometric method. We observed that the proposed biosensor has a linear detection range over a concentration range from 10 mM to 200 mM with good sensitivity of 89.10±1.23 mV/decade. In addition, the proposed biosensor has shown fast time response of less than 10 s and a good selectivity towards galactose in the presence of common interferents such as ascorbic acid, uric acid, glucose, and magnesium ions. The galactose biosensor based on galactose oxidase immobilized ZnO nanorods has a shelf life more than four weeks.

Inducible bilirubin oxidase: A novel function for the mouse cytochrome P450 2A5

Energy Technology Data Exchange (ETDEWEB)

Abu-Bakar, A' edah, E-mail: a.abubakar@uq.edu.au [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Arthur, Dionne Maioha [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Cooperative Research Centre for Contamination Assessment and Remediation of the Environment, Adelaide (Australia); Aganovic, Simona [Department of Pharmaceutical Biosciences, Uppsala University, Biomedical Centre, Box 578, S-751 23 Uppsala (Sweden); Ng, Jack C. [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Cooperative Research Centre for Contamination Assessment and Remediation of the Environment, Adelaide (Australia); Lang, Matti A. [The University of Queensland, National Research Centre for Environmental Toxicology (Entox), 4072 Brisbane, Queensland (Australia); Department of Pharmaceutical Biosciences, Uppsala University, Biomedical Centre, Box 578, S-751 23 Uppsala (Sweden)

2011-11-15

We have previously shown that bilirubin (BR), a breakdown product of haem, is a strong inhibitor and a high affinity substrate of the mouse cytochrome P450 2A5 (CYP2A5). The antioxidant BR, which is cytotoxic at high concentrations, is potentially useful in cellular protection against oxygen radicals if its intracellular levels can be strictly controlled. The mechanisms that regulate cellular BR levels are still obscure. In this paper we provide preliminary evidence for a novel function of CYP2A5 as hepatic 'BR oxidase'. A high-performance liquid chromatography/electrospray ionisation mass spectrometry screening showed that recombinant yeast microsomes expressing the CYP2A5 oxidise BR to biliverdin, as the main metabolite, and to three other smaller products with m/z values of 301, 315 and 333. The metabolic profile is significantly different from that of chemical oxidation of BR. In chemical oxidation the smaller products were the main metabolites. This suggests that the enzymatic reaction is selective, towards biliverdin production. Bilirubin treatment of primary hepatocytes increased the CYP2A5 protein and activity levels with no effect on the corresponding mRNA. Co-treatment with cycloheximide (CHX), a protein synthesis inhibitor, resulted in increased half-life of the CYP2A5 compared to cells treated only with CHX. Collectively, the observations suggest that the CYP2A5 is potentially an inducible 'BR oxidase' where BR may accelerate its own metabolism through stabilization of the CYP2A5 protein. It is possible that this metabolic pathway is potentially part of the machinery controlling intracellular BR levels in transient oxidative stress situations, in which high amounts of BR are produced. -- Highlights: Black-Right-Pointing-Pointer CYP2A5 metabolizes bilirubin to biliverdin and dipyrroles. Black-Right-Pointing-Pointer Bilirubin increased the hepatic CYP2A5 protein and activity levels. Black-Right-Pointing-Pointer Bilirubin does not

Regulation of the NADPH Oxidase RBOHD During Plant Immunity.

Science.gov (United States)

Kadota, Yasuhiro; Shirasu, Ken; Zipfel, Cyril

2015-08-01

Pathogen recognition induces the production of reactive oxygen species (ROS) by NADPH oxidases in both plants and animals. ROS have direct antimicrobial properties, but also serve as signaling molecules to activate further immune outputs. However, ROS production has to be tightly controlled to avoid detrimental effects on host cells, but yet must be produced in the right amount, at the right place and at the right time upon pathogen perception. Plant NADPH oxidases belong to the respiratory burst oxidase homolog (RBOH) family, which contains 10 members in the model plant Arabidopsis thaliana. The perception of pathogen-associated molecular patterns (PAMPs) by pattern recognition receptors (PRRs) leads to a rapid, specific and strong production of ROS, which is dependent on RBOHD. RBOHD is mainly controlled by Ca(2+) via direct binding to EF-hand motifs and phosphorylation by Ca(2+)-dependent protein kinases. Recent studies have, however, revealed a critical role for a Ca(2+)-independent regulation of RBOHD. The plasma membrane-associated cytoplasmic kinase BIK1 (BOTRYTIS-INDUCED KINASE1), which is a direct substrate of the PRR complex, directly interacts with and phosphorylates RBOHD upon PAMP perception. Impairment of these phosphorylation events completely abolishes the function of RBOHD in immunity. These results suggest that RBOHD activity is tightly controlled by multilayered regulations. In this review, we summarize recent advances in our understanding of the regulatory mechanisms controlling RBOHD activation. © The Author 2015. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Potassium-doped carbon nanotubes toward the direct electrochemistry of cholesterol oxidase and its application in highly sensitive cholesterol biosensor

Energy Technology Data Exchange (ETDEWEB)

Li Xiaorong [State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093 (China); Xu Jingjuan, E-mail: xujj@nju.edu.cn [State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093 (China); Chen Hongyuan [State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093 (China)

2011-10-30

We demonstrate herein a newly developed serum total cholesterol biosensor by using the direct electron transfer of cholesterol oxidase (ChOx), which is based on the immobilization of cholesterol oxidase and cholesterol esterase (ChEt) on potassium-doped multi-walled carbon nanotubes (KMWNTs) modified electrodes. The KMWNTs accelerate the electron transfer from electrode surface to the immobilized ChOx, achieving the direct electrochemistry of ChOx and maintaining its bioactivity. As a new platform in cholesterol analysis, the resulting electrode (ChOx/KMWNTs/GCE) exhibits a sensitive response to free cholesterol, with a linear range of 0.050-16.0 {mu}mol L{sup -1} and a detection limit of 5.0 nmol L{sup -1} (S/N = 3). Coimmobilization of ChEt and ChOx (ChEt/ChOx/KMWNTs/GCE) allows the determination of both free cholesterol and esterified cholesterol. The resulting biosensor shows the same linear range of 0.050-16.0 {mu}mol L{sup -1} for free cholesterol and cholesteryl oleate, with the detection limit of 10.0 and 12.0 nmol L{sup -1} (S/N = 3), respectively. The concentrations of total (free and esterified) cholesterol in human serum samples, determined by using the techniques developed in the present study, are in good agreement with those determined by the well-established techniques using the spectrophotometry.

Identification, expression, and taxonomic distribution of alternative oxidases in non-angiosperm plants.

Science.gov (United States)

Neimanis, Karina; Staples, James F; Hüner, Norman P A; McDonald, Allison E

2013-09-10

Alternative oxidase (AOX) is a terminal ubiquinol oxidase present in the respiratory chain of all angiosperms investigated to date, but AOX distribution in other members of the Viridiplantae is less clear. We assessed the taxonomic distribution of AOX using bioinformatics. Multiple sequence alignments compared AOX proteins and examined amino acid residues involved in AOX catalytic function and post-translational regulation. Novel AOX sequences were found in both Chlorophytes and Streptophytes and we conclude that AOX is widespread in the Viridiplantae. AOX multigene families are common in non-angiosperm plants and the appearance of AOX1 and AOX2 subtypes pre-dates the divergence of the Coniferophyta and Magnoliophyta. Residues involved in AOX catalytic function are highly conserved between Chlorophytes and Streptophytes, while AOX post-translational regulation likely differs in these two lineages. We demonstrate experimentally that an AOX gene is present in the moss Physcomitrella patens and that the gene is transcribed. Our findings suggest that AOX will likely exert an influence on plant respiration and carbon metabolism in non-angiosperms such as green algae, bryophytes, liverworts, lycopods, ferns, gnetophytes, and gymnosperms and that further research in these systems is required. Copyright © 2013 Elsevier B.V. All rights reserved.

Acupuncture elicits neuroprotective effect by inhibiting NAPDH oxidase-mediated reactive oxygen species production in cerebral ischaemia.

Science.gov (United States)

Shi, Guang-Xia; Wang, Xue-Rui; Yan, Chao-Qun; He, Tian; Yang, Jing-Wen; Zeng, Xiang-Hong; Xu, Qian; Zhu, Wen; Du, Si-Qi; Liu, Cun-Zhi

2015-12-10

In the current study, we aimed to investigate whether NADPH oxidase, a major ROS-producing enzyme, was involved in the antioxidant effect of acupuncture on cognitive impairment after cerebral ischaemia. The cognitive function, infract size, neuron cell loss, level of superoxide anion and expression of NADPH oxidase subunit in hippocampus of two-vessel occlusion (2VO) rats were determined after 2-week acupuncture. Furthermore, the cognitive function and production of O2(-) were determined in the presence and absence of NADPH oxidase agonist (TBCA) and antagonist (Apocynin). The effect of acupuncture on cognitive function after cerebral ischaemia in gp91phox-KO mice was evaluated by Morris water maze. Acupuncture reduced infarct size, attenuated overproduction of O2(-), and reversed consequential cognitive impairment and neuron cell loss in 2VO rats. The elevations of gp91phox and p47phox after 2VO were significantly decreased after acupuncture treatment. However, no differences of gp91phox mRNA were found among any experimental groups. Furthermore, these beneficial effects were reversed by TBCA, whereas apocynin mimicked the effect of acupuncture by improving cognitive function and decreasing O2(-) generation. Acupuncture failed to improve the memory impairment in gp91phox KO mice. Full function of the NADPH oxidase enzyme plays an important role in neuroprotective effects against cognitive impairment via inhibition of NAPDH oxidase-mediated oxidative stress.

The cytochemical demonstration of catalase and D-amino acid oxidase in the microbodies of teleost kidney cells

NARCIS (Netherlands)

Veenhuis, M.; Wendelaar Bonga, S.D.

1977-01-01

The distribution of catalase and D-amino acid oxidase, marker enzymes for peroxisomes, was determined cytochemically in the kidney tubules of an euryhaline teleost, the three-spined stickleback. Catalase activity was localized with the diaminobenzidine technique. The presence of D-amino acid oxidase

The Effects of Polyphenol Oxidase and Cycloheximide on the Early Stage of Browning in Phalaenopsis Explants

Directory of Open Access Journals (Sweden)

Xu Chuanjun

2015-11-01

Full Text Available Explant browning is one of the major problems in the tissue culture process, and polyphenol oxidase (PPO, is the major proteases involved in plant tissue browning. We investigated the effects of polyphenol oxidase on the early stage of browning in explants of the orchid Phalaenopsis. Our results show that PPO activity was significantly higher in explants cultured for 3 d than in the 0 h control. The levels of PPO transcripts and PPO protein were significantly higher in explants cultured for 6 h compared to the 0 h control; these high expression levels were maintained over increasing cultivation time. Cycloheximide (CHX treatment reduced PPO transcript levels, PPO protein levels, and PPO enzyme activity. High levels of PPO mRNA and PPO protein were detected in the cytoplasm and vascular bundles of Phalaenopsis explants cultured for 6 h compared to explants cultured for 0 h, 24 h, and 3 d. CHX treatment did not significantly affect the distribution of PPO mRNA and PPO protein in explant tissues, but their levels were significantly lower than those of the untreated control.

Valencene oxidase CYP706M1 from Alaska cedar (Callitropsis nootkatensis).

Science.gov (United States)

Cankar, Katarina; van Houwelingen, Adèle; Goedbloed, Miriam; Renirie, Rokus; de Jong, René M; Bouwmeester, Harro; Bosch, Dirk; Sonke, Theo; Beekwilder, Jules

2014-03-18

(+)-Nootkatone is a natural sesquiterpene ketone used in grapefruit and citrus flavour compositions. It occurs in small amounts in grapefruit and is a major component of Alaska cedar (Callitropsis nootkatensis) heartwood essential oil. Upon co-expression of candidate cytochrome P450 enzymes from Alaska cedar in yeast with a valencene synthase, a C. nootkatensis valencene oxidase (CnVO) was identified to produce trans-nootkatol and (+)-nootkatone. Formation of (+)-nootkatone was detected at 144±10μg/L yeast culture. CnVO belongs to a new subfamily of the CYP706 family of cytochrome P450 oxidases. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Comparison of brain mitochondrial cytochrome c oxidase activity with cyanide LD(50) yields insight into the efficacy of prophylactics.

Science.gov (United States)

Marziaz, Mandy L; Frazier, Kathryn; Guidry, Paul B; Ruiz, Robyn A; Petrikovics, Ilona; Haines, Donovan C

2013-01-01

Cyanide inhibits cytochrome c oxidase, the terminal oxidase of the mitochondrial respiratory pathway, therefore inhibiting the cell oxygen utilization and resulting in the condition of histotoxic anoxia. The enzyme rhodanese detoxifies cyanide by utilizing sulfur donors to convert cyanide to thiocyanate, and new and improved sulfur donors are actively sought as researchers seek to improve cyanide prophylactics. We have determined brain cytochrome c oxidase activity as a marker for cyanide exposure for mice pre-treated with various cyanide poisoning prophylactics, including sulfur donors thiosulfate (TS) and thiotaurine (TT3). Brain mitochondria were isolated by differential centrifugation, the outer mitochondrial membrane was disrupted by a maltoside detergent, and the decrease in absorbance at 550 nm as horse heart ferrocytochrome c (generated by the dithiothreitol reduction of ferricytochrome c) was oxidized was monitored. Overall, the TS control prophylactic treatment provided significant protection of the cytochrome c oxidase activity. The TT3-treated mice showed reduced cytochrome c oxidase activity even in the absence of cyanide. In both treatment series, addition of exogenous Rh did not significantly enhance the prevention of cytochrome c oxidase inhibition, but the addition of sodium nitrite did. These findings can lead to a better understanding of the protection mechanism by various cyanide antidotal systems. Copyright © 2011 John Wiley & Sons, Ltd.

Secreted fungal sulfhydryl oxidases: sequence analysis and characterisation of a representative flavin-dependent enzyme from Aspergillus oryzae

Directory of Open Access Journals (Sweden)

Faccio Greta

2010-08-01

Full Text Available Abstract Background Sulfhydryl oxidases are flavin-dependent enzymes that catalyse the formation of de novo disulfide bonds from free thiol groups, with the reduction of molecular oxygen to hydrogen peroxide. Sulfhydryl oxidases have been investigated in the food industry to remove the burnt flavour of ultraheat-treated milk and are currently studied as potential crosslinking enzymes, aiming at strengthening wheat dough and improving the overall bread quality. Results In the present study, potential sulfhydryl oxidases were identified in the publicly available fungal genome sequences and their sequence characteristics were studied. A representative sulfhydryl oxidase from Aspergillus oryzae, AoSOX1, was expressed in the fungus Trichoderma reesei. AoSOX1 was produced in relatively good yields and was purified and biochemically characterised. The enzyme catalysed the oxidation of thiol-containing compounds like glutathione, D/L-cysteine, beta-mercaptoethanol and DTT. The enzyme had a melting temperature of 57°C, a pH optimum of 7.5 and its enzymatic activity was completely inhibited in the presence of 1 mM ZnSO4. Conclusions Eighteen potentially secreted sulfhydryl oxidases were detected in the publicly available fungal genomes analysed and a novel proline-tryptophan dipeptide in the characteristic motif CXXC, where X is any amino acid, was found. A representative protein, AoSOX1 from A. oryzae, was produced in T. reesei in an active form and had the characteristics of sulfhydryl oxidases. Further testing of the activity on thiol groups within larger peptides and on protein level will be needed to assess the application potential of this enzyme.

Secreted fungal sulfhydryl oxidases: sequence analysis and characterisation of a representative flavin-dependent enzyme from Aspergillus oryzae.

Science.gov (United States)

Faccio, Greta; Kruus, Kristiina; Buchert, Johanna; Saloheimo, Markku

2010-08-20

Sulfhydryl oxidases are flavin-dependent enzymes that catalyse the formation of de novo disulfide bonds from free thiol groups, with the reduction of molecular oxygen to hydrogen peroxide. Sulfhydryl oxidases have been investigated in the food industry to remove the burnt flavour of ultraheat-treated milk and are currently studied as potential crosslinking enzymes, aiming at strengthening wheat dough and improving the overall bread quality. In the present study, potential sulfhydryl oxidases were identified in the publicly available fungal genome sequences and their sequence characteristics were studied. A representative sulfhydryl oxidase from Aspergillus oryzae, AoSOX1, was expressed in the fungus Trichoderma reesei. AoSOX1 was produced in relatively good yields and was purified and biochemically characterised. The enzyme catalysed the oxidation of thiol-containing compounds like glutathione, D/L-cysteine, beta-mercaptoethanol and DTT. The enzyme had a melting temperature of 57°C, a pH optimum of 7.5 and its enzymatic activity was completely inhibited in the presence of 1 mM ZnSO4. Eighteen potentially secreted sulfhydryl oxidases were detected in the publicly available fungal genomes analysed and a novel proline-tryptophan dipeptide in the characteristic motif CXXC, where X is any amino acid, was found. A representative protein, AoSOX1 from A. oryzae, was produced in T. reesei in an active form and had the characteristics of sulfhydryl oxidases. Further testing of the activity on thiol groups within larger peptides and on protein level will be needed to assess the application potential of this enzyme.

Proton translocation stoichiometry of cytochrome oxidase: use of a fast-responding oxygen electrode.

Science.gov (United States)

Reynafarje, B; Alexandre, A; Davies, P; Lehninger, A L

1982-01-01

The mechanistic stoichiometry of vectorial H+ ejection coupled to electron transport from added ferrocytochrome c to oxygen by the cytochrome oxidase (EC 1.9.3.1) of rat liver mitoplasts was determined from measurements of the initial rates of electron flow and H+ ejection in the presence of K+ (with valinomycin). Three different methods of measuring electron flow were used: (a) dual-wavelength spectrophotometry of ferrocytochrome c oxidation, (b) uptake of scalar H+ for the reduction of O2 in the presence of a protonophore, and (c) a fast-responding membraneless oxygen electrode. The reliability of the rate measurements was first established against the known stoichiometry of the scalar reaction of cytochrome oxidase (2ferrocytochrome c + 2H+ + 1/2O2 leads to 2ferricytochrome c + H2O) in the presence of excess protonophore. With all three methods the directly observed vectorial H+/O ejection ratios in the presence of K+ + valinomycin significantly exceeded 3.0. However, because the rate of backflow of the ejected H+ into the mitoplasts is very high and increases with the increasing delta pH generated across the membrane, there is a very rapid decline in the observed H+/O ratio from the beginning of the reaction. Kinetic analysis of ferrocytochrome c oxidation by the mitoplasts, carried out with a fast-responding membraneless oxygen electrode, showed the reaction to be first order in O2 and allowed accurate extrapolation of the rates of O2 uptake and H+ ejection to zero time. At this point, at which there is zero delta pH across the membrane, the H+/O ejection ratio of the cytochrome oxidase reaction, obtained from the rates at zero time, is close to 4.0. PMID:6296824

Immobilization of glucose oxidase on sepharose by UV-initiated graft copolymerization

International Nuclear Information System (INIS)

D'Angiuro, L.; Cremonesi, P.

1982-01-01

The performance of a new method of enzyme immobilization based on photochemically initiated direct graft copolymerization was recently investigated. The immobilization reaction can be carried out in a simple way and by carefully selecting the reaction conditions, the enzyme-graft copolymer can be obtained as the main reaction product. Coupling efficiency of glucose oxidase has been found to depend only on the amount of photocatalyst (FeCl 3 ) fixed on Sepharose used as polysaccharide support. Small quantities of glycidylmethacrylate (GMA) (0.25 g/g dry Sepharose) are sufficient but necessary to achieve the best enzyme coupling efficiency (20-40%). Enzyme immobilization occurs very rapidly and the entire reaction occurs within 60 min. Reaction patterns and physicochemical characteristics of the obtained enzyme-graft copolymers exclude the glucose oxidase entrapment: therefore a covalent attachment mechanism may be proposed. The kinetic parameters of immobilized glucose oxidase (K/sub m/' = 2.0 x 10 -2 M) are quite similar to those of free enzyme (K/sub m/ = 1.93 x 10 -2 M), and no diffusion limitation phenomena are evidenced in samples having different enzyme or polymer content. Lyophilization, thermostability, and long-term continuous operation also have been investigated. The advantages of this method over that using vinylenzyme copolymerization are discussed

NADPH Oxidase Activity in Cerebral Arterioles Is a Key Mediator of Cerebral Small Vessel Disease-Implications for Prevention.

Science.gov (United States)

McCarty, Mark F

2015-04-15

Cerebral small vessel disease (SVD), a common feature of brain aging, is characterized by lacunar infarcts, microbleeds, leukoaraiosis, and a leaky blood-brain barrier. Functionally, it is associated with cognitive decline, dementia, depression, gait abnormalities, and increased risk for stroke. Cerebral arterioles in this syndrome tend to hypertrophy and lose their capacity for adaptive vasodilation. Rodent studies strongly suggest that activation of Nox2-dependent NADPH oxidase activity is a crucial driver of these structural and functional derangements of cerebral arterioles, in part owing to impairment of endothelial nitric oxide synthase (eNOS) activity. This oxidative stress may also contribute to the breakdown of the blood-brain barrier seen in SVD. Hypertension, aging, metabolic syndrome, smoking, hyperglycemia, and elevated homocysteine may promote activation of NADPH oxidase in cerebral arterioles. Inhibition of NADPH oxidase with phycocyanobilin from spirulina, as well as high-dose statin therapy, may have potential for prevention and control of SVD, and high-potassium diets merit study in this regard. Measures which support effective eNOS activity in other ways-exercise training, supplemental citrulline, certain dietary flavonoids (as in cocoa and green tea), and capsaicin, may also improve the function of cerebral arterioles. Asian epidemiology suggests that increased protein intakes may decrease risk for SVD; conceivably, arginine and/or cysteine-which boosts tissue glutathione synthesis, and can be administered as N-acetylcysteine-mediate this benefit. Ameliorating the risk factors for SVD-including hypertension, metabolic syndrome, hyperglycemia, smoking, and elevated homocysteine-also may help to prevent and control this syndrome, although few clinical trials have addressed this issue to date.

Low temperature (< 100 °C) deposited P-type cuprous oxide thin films: Importance of controlled oxygen and deposition energy

International Nuclear Information System (INIS)

Li, Flora M.; Waddingham, Rob; Milne, William I.; Flewitt, Andrew J.; Speakman, Stuart; Dutson, James; Wakeham, Steve; Thwaites, Mike

2011-01-01

With the emergence of transparent electronics, there has been considerable advancement in n-type transparent semiconducting oxide (TSO) materials, such as ZnO, InGaZnO, and InSnO. Comparatively, the availability of p-type TSO materials is more scarce and the available materials are less mature. The development of p-type semiconductors is one of the key technologies needed to push transparent electronics and systems to the next frontier, particularly for implementing p–n junctions for solar cells and p-type transistors for complementary logic/circuits applications. Cuprous oxide (Cu 2 O) is one of the most promising candidates for p-type TSO materials. This paper reports the deposition of Cu 2 O thin films without substrate heating using a high deposition rate reactive sputtering technique, called high target utilisation sputtering (HiTUS). This technique allows independent control of the remote plasma density and the ion energy, thus providing finer control of the film properties and microstructure as well as reducing film stress. The effect of deposition parameters, including oxygen flow rate, plasma power and target power, on the properties of Cu 2 O films are reported. It is known from previously published work that the formation of pure Cu 2 O film is often difficult, due to the more ready formation or co-formation of cupric oxide (CuO). From our investigation, we established two key concurrent criteria needed for attaining Cu 2 O thin films (as opposed to CuO or mixed phase CuO/Cu 2 O films). First, the oxygen flow rate must be kept low to avoid over-oxidation of Cu 2 O to CuO and to ensure a non-oxidised/non-poisoned metallic copper target in the reactive sputtering environment. Secondly, the energy of the sputtered copper species must be kept low as higher reaction energy tends to favour the formation of CuO. The unique design of the HiTUS system enables the provision of a high density of low energy sputtered copper radicals/ions, and when combined with a

Cytochrome c oxidase loses catalytic activity and structural integrity during the aging process in Drosophila melanogaster

Energy Technology Data Exchange (ETDEWEB)

Ren, Jian-Ching; Rebrin, Igor [Department of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, CA 90033 (United States); Klichko, Vladimir; Orr, William C. [Department of Biological Sciences, Southern Methodist University, Dallas, TX 75275 (United States); Sohal, Rajindar S., E-mail: sohal@usc.edu [Department of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, CA 90033 (United States)

2010-10-08

Research highlights: {yields} Cytochrome c oxidase loses catalytic activity during the aging process. {yields} Abundance of seven nuclear-encoded subunits of cytochrome c oxidase decreased with age in Drosophila. {yields} Cytochrome c oxidase is specific intra-mitochondrial site of age-related deterioration. -- Abstract: The hypothesis, that structural deterioration of cytochrome c oxidase (CcO) is a causal factor in the age-related decline in mitochondrial respiratory activity and an increase in H{sub 2}O{sub 2} generation, was tested in Drosophila melanogaster. CcO activity and the levels of seven different nuclear DNA-encoded CcO subunits were determined at three different stages of adult life, namely, young-, middle-, and old-age. CcO activity declined progressively with age by 33%. Western blot analysis, using antibodies specific to Drosophila CcO subunits IV, Va, Vb, VIb, VIc, VIIc, and VIII, indicated that the abundance these polypeptides decreased, ranging from 11% to 40%, during aging. These and previous results suggest that CcO is a specific intra-mitochondrial site of age-related deterioration, which may have a broad impact on mitochondrial physiology.

Cytochrome c oxidase loses catalytic activity and structural integrity during the aging process in Drosophila melanogaster

International Nuclear Information System (INIS)

Ren, Jian-Ching; Rebrin, Igor; Klichko, Vladimir; Orr, William C.; Sohal, Rajindar S.

2010-01-01

Research highlights: → Cytochrome c oxidase loses catalytic activity during the aging process. → Abundance of seven nuclear-encoded subunits of cytochrome c oxidase decreased with age in Drosophila. → Cytochrome c oxidase is specific intra-mitochondrial site of age-related deterioration. -- Abstract: The hypothesis, that structural deterioration of cytochrome c oxidase (CcO) is a causal factor in the age-related decline in mitochondrial respiratory activity and an increase in H 2 O 2 generation, was tested in Drosophila melanogaster. CcO activity and the levels of seven different nuclear DNA-encoded CcO subunits were determined at three different stages of adult life, namely, young-, middle-, and old-age. CcO activity declined progressively with age by 33%. Western blot analysis, using antibodies specific to Drosophila CcO subunits IV, Va, Vb, VIb, VIc, VIIc, and VIII, indicated that the abundance these polypeptides decreased, ranging from 11% to 40%, during aging. These and previous results suggest that CcO is a specific intra-mitochondrial site of age-related deterioration, which may have a broad impact on mitochondrial physiology.

Heterologous expression of the Crassostrea gigas (Pacific oyster) alternative oxidase in the yeast Saccharomyces cerevisiae.

Science.gov (United States)

Robertson, Aaron; Schaltz, Kyle; Neimanis, Karina; Staples, James F; McDonald, Allison E

2016-10-01

Alternative oxidase (AOX) is a terminal oxidase within the inner mitochondrial membrane (IMM) present in many organisms where it functions in the electron transport system (ETS). AOX directly accepts electrons from ubiquinol and is therefore capable of bypassing ETS Complexes III and IV. The human genome does not contain a gene coding for AOX, so AOX expression has been suggested as a gene therapy for a range of human mitochondrial diseases caused by genetic mutations that render Complex III and/or IV dysfunctional. An effective means of screening mutations amenable to AOX treatment remains to be devised. We have generated such a tool by heterologously expressing AOX from the Pacific oyster (Crassostrea gigas) in the yeast Saccharomyces cerevisiae under the control of a galactose promoter. Our results show that this animal AOX is monomeric and is correctly targeted to yeast mitochondria. Moreover, when expressed in yeast, Pacific oyster AOX is a functional quinol oxidase, conferring cyanide-resistant growth and myxothiazol-resistant oxygen consumption to yeast cells and isolated mitochondria. This system represents a high-throughput screening tool for determining which Complex III and IV genetic mutations in yeast will be amenable to AOX gene therapy. As many human genes are orthologous to those found in yeast, our invention represents an efficient and cost-effective way to evaluate viable research avenues. In addition, this system provides the opportunity to learn more about the localization, structure, and regulation of AOXs from animals that are not easily reared or manipulated in the lab.

Effect of a heme oxygenase-1 inducer on NADPH oxidase ...

African Journals Online (AJOL)

Effect of a heme oxygenase-1 inducer on NADPH oxidase expression in ... and immunohistochemistry of hepatic NOX1 and NOX4 were investigated in week 4. ... (HO-1 inhibitor) administration caused upregulation of NOX gene expression ...

Biocatalytic properties and structural analysis of eugenol oxidase from Rhodococcus jostii RHA1 : a versatile oxidative biocatalyst

NARCIS (Netherlands)

Nguyen, Quoc-Thai; De Gonzalo, Gonzalo; Binda, Claudia; Rioz, Ana; Mattevi, Andrea; Fraaije, Marco

2016-01-01

Eugenol oxidase (EUGO) from Rhodococcus jostii RHA1 was previously shown to convert only a limited set of phenolic compounds. In this study, we have explored the biocatalytic potential of this flavoprotein oxidase resulting in a broadened substrate scope and a deeper insight into its structural

Process technology for the application of d-amino acid oxidases in pharmaceutical intermediate manufacturing

DEFF Research Database (Denmark)

Tindal, Stuart; Carr, Reuben; Archer, Ian V. J.

2011-01-01

Recent advances in biocatalysis have seen increased interest in the use of D-amino acid oxidase to synthesize optically pure amino acids. However, the creation of a genuine oxidase based platform technology will require suitable process technology as well as an understanding of the challenges...... and opportunities of a wider portfolio of synthetic targets. In this article we address some of the recent progress in process technology to enable the future development of a generic platform technology....

Extracellular oxidases and the transformation of solubilised low-rank coal by wood-rot fungi

Energy Technology Data Exchange (ETDEWEB)

Ralph, J.P. [Flinders Univ. of South Australia, Bedford Park (Australia). School of Biological Sciences; Graham, L.A. [Flinders Univ. of South Australia, Bedford Park (Australia). School of Biological Sciences; Catcheside, D.E.A. [Flinders Univ. of South Australia, Bedford Park (Australia). School of Biological Sciences

1996-12-31

The involvement of extracellular oxidases in biotransformation of low-rank coal was assessed by correlating the ability of nine white-rot and brown-rot fungi to alter macromolecular material in alkali-solubilised brown coal with the spectrum of oxidases they produce when grown on low-nitrogen medium. The coal fraction used was that soluble at 3.0{<=}pH{<=}6.0 (SWC6 coal). In 15-ml cultures, Gloeophyllum trabeum, Lentinus lepideus and Trametes versicolor produced little or no lignin peroxidase, manganese (Mn) peroxidase or laccase activity and caused no change to SWC6 coal. Ganoderma applanatum and Pycnoporus cinnabarinus also produced no detectable lignin or Mn peroxidases or laccase yet increased the absorbance at 400 nm of SWC6 coal. G. applanatum, which produced veratryl alcohol oxidase, also increased the modal apparent molecular mass. SWC6 coal exposed to Merulius tremellosus and Perenniporia tephropora, which secreted Mn peroxidases and laccase and Phanerochaete chrysosporium, which produced Mn and lignin peroxidases was polymerised but had unchanged or decreased absorbance. In the case of both P. chrysosporium and M. tremellosus, polymerisation of SWC6 coal was most extensive, leading to the formation of a complex insoluble in 100 mM NaOH. Rigidoporus ulmarius, which produced only laccase, both polymerised and reduced the A{sub 400} of SWC6 coal. P. chrysosporium, M. tremellosus and P. tephropora grown in 10-ml cultures produced a spectrum of oxidases similar to that in 15-ml cultures but, in each case, caused more extensive loss of A{sub 400}, and P. chrysosporium depolymerised SWC6 coal. It is concluded that the extracellular oxidases of white-rot fungi can transform low-rank coal macromolecules and that increased oxygen availability in the shallower 10-ml cultures favours catabolism over polymerisation. (orig.)

A multicopper oxidase-related protein is essential for insect viability, longevity and ovary development.

Science.gov (United States)

Peng, Zeyu; Green, Peter G; Arakane, Yasuyuki; Kanost, Michael R; Gorman, Maureen J

2014-01-01

Typical multicopper oxidases (MCOs) have ten conserved histidines and one conserved cysteine that coordinate four copper atoms. These copper ions are required for oxidase activity. During our studies of insect MCOs, we discovered a gene that we named multicopper oxidase-related protein (MCORP). MCORPs share sequence similarity with MCOs, but lack many of the copper-coordinating residues. We identified MCORP orthologs in many insect species, but not in other invertebrates or vertebrates. We predicted that MCORPs would lack oxidase activity due to the absence of copper-coordinating residues. To test this prediction, we purified recombinant Tribolium castaneum (red flour beetle) MCORP and analyzed its enzymatic activity using a variety of substrates. As expected, no oxidase activity was detected. To study MCORP function in vivo, we analyzed expression profiles of TcMCORP and Anopheles gambiae (African malaria mosquito) MCORP, and assessed RNAi-mediated knockdown phenotypes. We found that both MCORPs are constitutively expressed at a low level in all of the tissues we analyzed. Injection of TcMCORP dsRNA into larvae resulted in 100% mortality prior to adult eclosion, with death occurring mainly during the pharate pupal stage or late pharate adult stage. Injection of TcMCORP dsRNA into pharate pupae resulted in the death of approximately 20% of the treated insects during the pupal to adult transition and a greatly shortened life span for the remaining insects. In addition, knockdown of TcMCORP in females prevented oocyte maturation and, thus, greatly decreased the number of eggs laid. These results indicate that TcMCORP is an essential gene and that its function is required for reproduction. An understanding of the role MCORP plays in insect physiology may help to develop new strategies for controlling insect pests.

Pseudo-bi-enzyme glucose sensor: ZnS hollow spheres and glucose oxidase concerted catalysis glucose.

Science.gov (United States)

Shuai, Ying; Liu, Changhua; Wang, Jia; Cui, Xiaoyan; Nie, Ling

2013-06-07

This work creatively uses peroxidase-like ZnS hollow spheres (ZnS HSs) to cooperate with glucose oxidase (GOx) for glucose determinations. This approach is that the ZnS HSs electrocatalytically oxidate the enzymatically generated H2O2 to O2, and then the O2 circularly participates in the previous glucose oxidation by glucose oxidase. Au nanoparticles (AuNPs) and carbon nanotubes (CNTs) are used as electron transfer and enzyme immobilization matrices, respectively. The biosensor of glucose oxidase-carbon nanotubes-Au nanoparticles-ZnS hollow spheres-gold electrode (GOx-CNT-AuNPs-ZnS HSs-GE) exhibits a rapid response, a low detection limit (10 μM), a wide linear range (20 μM to 7 mM) as well as good anti-interference, long-term longevity and reproducibility.

Expression and Characterization of Glucose Oxidase from Aspergillus niger in Yarrowia lipolytica.

Science.gov (United States)

Khadivi Derakshan, Fatemeh; Darvishi, Farshad; Dezfulian, Mehrouz; Madzak, Catherine

2017-08-01

Glucose oxidase (GOX) is currently used in clinical, pharmaceutical, food and chemical industries. The aim of this study was expression and characterization of Aspergillus niger glucose oxidase gene in the yeast Yarrowia lipolytica. For the first time, the GOX gene of A. niger was successfully expressed in Y. lipolytica using a mono-integrative vector containing strong hybrid promoter and secretion signal. The highest total glucose oxidase activity was 370 U/L after 7 days of cultivation. An innovative method was used to cell wall disruption in current study, and it could be recommended to use for efficiently cell wall disruption of Y. lipolytica. Optimum pH and temperature for recombinant GOX activity were 5.5 and 37 °C, respectively. A single band with a molecular weight of 80 kDa similar to the native and pure form of A. niger GOX was observed for the recombinant GOX in SDS-PAGE analysis. Y. lipolytica is a suitable and efficient eukaryotic expression system to production of recombinant GOX in compered with other yeast expression systems and could be used to production of pure form of GOX for industrial applications.

Imaging Monoamine Oxidase in the Human Brain

Energy Technology Data Exchange (ETDEWEB)

Fowler, J. S.; Volkow, N. D.; Wang, G-J.; Logan, Jean

1999-11-10

Positron emission tomography (PET) studies mapping monoamine oxidase in the human brain have been used to measure the turnover rate for MAO B; to determine the minimum effective dose of a new MAO inhibitor drug lazabemide and to document MAO inhibition by cigarette smoke. These studies illustrate the power of PET and radiotracer chemistry to measure normal biochemical processes and to provide information on the effect of drug exposure on specific molecular targets.

Imaging Monoamine Oxidase in the Human Brain

International Nuclear Information System (INIS)

Fowler, J. S.; Volkow, N. D.; Wang, G-J.; Logan, Jean

1999-01-01

Positron emission tomography (PET) studies mapping monoamine oxidase in the human brain have been used to measure the turnover rate for MAO B; to determine the minimum effective dose of a new MAO inhibitor drug lazabemide and to document MAO inhibition by cigarette smoke. These studies illustrate the power of PET and radiotracer chemistry to measure normal biochemical processes and to provide information on the effect of drug exposure on specific molecular targets

A new methodology for the determination of enzyme activity based on carbon nanotubes and glucose oxidase.

Science.gov (United States)

Yeşiller, Gülden; Sezgintürk, Mustafa Kemal

2015-11-10

In this research, a novel enzyme activity analysis methodology is introduced as a new perspective for this area. The activity of elastase enzyme, which is a digestive enzyme mostly of found in the digestive system of vertebrates, was determined by an electrochemical device composed of carbon nanotubes and a second enzyme, glucose oxidase, which was used as a signal generator enzyme. In this novel methodology, a complex bioactive layer was constructed by using carbon nanotubes, glucose oxidase and a supporting protein, gelatin on a solid, conductive substrate. The activity of elastase was determined by monitoring the hydrolysis rate of elastase enzyme in the bioactive layer. As a result of this hydrolysis of elastase, glucose oxidase was dissociated from the bioactive layer, and following this the electrochemical signal due to glucose oxidase was decreased. The progressive elastase-catalyzed digestion of the bioactive layer containing glucose oxidase decreased the layer's enzymatic efficiency, resulting in a decrease of the glucose oxidation current as a function of the enzyme activity. The ratio of the decrease was correlated to elastase activity level. In this study, optimization experiments of bioactive components and characterization of the resulting new electrochemical device were carried out. A linear calibration range from 0.0303U/mL to 0.0729U/mL of elastase was reported. Real sample analyses were also carried out by the new electrochemical device. Copyright © 2015 Elsevier B.V. All rights reserved.

High-performance glucose biosensor based on chitosan-glucose oxidase immobilized polypyrrole/Nafion/functionalized multi-walled carbon nanotubes bio-nanohybrid film.

Science.gov (United States)

Shrestha, Bishnu Kumar; Ahmad, Rafiq; Mousa, Hamouda M; Kim, In-Gi; Kim, Jeong In; Neupane, Madhav Prasad; Park, Chan Hee; Kim, Cheol Sang

2016-11-15

A highly electroactive bio-nanohybrid film of polypyrrole (PPy)-Nafion (Nf)-functionalized multi-walled carbon nanotubes (fMWCNTs) nanocomposite was prepared on the glassy carbon electrode (GCE) by a facile one-step electrochemical polymerization technique followed by chitosan-glucose oxidase (CH-GOx) immobilization on its surface to achieve a high-performance glucose biosensor. The as-fabricated nanohybrid composite provides high surface area for GOx immobilization and thus enhances the enzyme-loading efficiency. The structural characterization revealed that the PPy-Nf-fMWCNTs nanocomposite films were uniformly formed on GCE and after GOx immobilization, the surface porosities of the film were decreased due to enzyme encapsulation inside the bio-nanohybrid composite materials. The electrochemical behavior of the fabricated biosensor was investigated by cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), and amperometry measurements. The results indicated an excellent catalytic property of bio-nanohybrid film for glucose detection with improved sensitivity of 2860.3μAmM(-1)cm(-2), the linear range up to 4.7mM (R(2)=0.9992), and a low detection limit of 5μM under a signal/noise (S/N) ratio of 3. Furthermore, the resulting biosensor presented reliable selectivity, better long-term stability, good repeatability, reproducibility, and acceptable measurement of glucose concentration in real serum samples. Thus, this fabricated biosensor provides an efficient and highly sensitive platform for glucose sensing and can open up new avenues for clinical applications. Copyright © 2016 Elsevier Inc. All rights reserved.

Nuclear expression of lysyl oxidase enzyme is an independent prognostic factor in rectal cancer patients

DEFF Research Database (Denmark)

Liu, Na; Cox, Thomas R; Cui, Weiyingqi

2017-01-01

Emerging evidence has implicated a pivotal role for lysyl oxidase (LOX) in cancer progression and metastasis. Whilst the majority of work has focused on the extracellular matrix cross-linking role of LOX, the exact function of intracellular LOX localisation remains unclear. In this study, we anal...... the nucleus of colon cancer cell lines by confocal microscopy and Western blot. Our results show a powerful link between nuclear LOX expression in tumours and patient survival, and offer a promising prognostic biomarker for rectal cancer patients....... analysed the LOX expression patterns in the nuclei of rectal cancer patient samples and determined the clinical significance of this expression. Nuclear LOX expression was significantly increased in patient lymph node metastases compared to their primary tumours. High nuclear LOX expression in tumours......Emerging evidence has implicated a pivotal role for lysyl oxidase (LOX) in cancer progression and metastasis. Whilst the majority of work has focused on the extracellular matrix cross-linking role of LOX, the exact function of intracellular LOX localisation remains unclear. In this study, we...

NADPH oxidase: an enzyme for multicellularity?

Science.gov (United States)

Lalucque, Hervé; Silar, Philippe

2003-01-01

Multicellularity has evolved several times during the evolution of eukaryotes. One evolutionary pressure that permits multicellularity relates to the division of work, where one group of cells functions as nutrient providers and the other in specialized roles such as defence or reproduction. This requires signalling systems to ensure harmonious development of multicellular structures. Here, we show that NADPH oxidases are specifically present in organisms that differentiate multicellular structures during their life cycle and are absent from unicellular life forms. The biochemical properties of these enzymes make them ideal candidates for a role in intercellular signalling.

Ag-doped CdO nanocatalysts: Preparation, characterization and catechol oxidase activity

Science.gov (United States)

El-Kemary, Maged; El-Mehasseb, Ibrahim; El-Shamy, Hany

2018-06-01

Silver doped cadmium oxide (Ag/CdO) nanoparticles with an average size of 41 nm have been successfully synthesized via thermal decomposition and liquid impregnation technique. The structural characterization has been performed by using several spectroscopic techniques, e.g., X-ray diffraction (XRD), scanning electron microscopy (SEM) and fourier-transform infrared (FT-IR). The catechol oxidase has been studied by UV-visible absorption spectroscopy and fourier-transform infrared as well as the mechanism has been assured by cyclic voltammetry and fluorescence spectroscopy. The results indicate that the oxidation does not occur in the presence of unsupported cadmium oxide particles by silver and in the same time, the catechol oxidase activity of silver doped CdO nanoparticles were improved by about three orders of magnitude than silver ions.

Brain monoamine oxidase B and A in human parkinsonian dopamine deficiency disorders.

Science.gov (United States)

Tong, Junchao; Rathitharan, Gausiha; Meyer, Jeffrey H; Furukawa, Yoshiaki; Ang, Lee-Cyn; Boileau, Isabelle; Guttman, Mark; Hornykiewicz, Oleh; Kish, Stephen J

2017-09-01

See Jellinger (doi:10.1093/awx190) for a scientific commentary on this article. The enzyme monoamine oxidases (B and A subtypes, encoded by MAOB and MAOA, respectively) are drug targets in the treatment of Parkinson's disease. Inhibitors of MAOB are used clinically in Parkinson's disease for symptomatic purposes whereas the potential disease-modifying effect of monoamine oxidase inhibitors is debated. As astroglial cells express high levels of MAOB, the enzyme has been proposed as a brain imaging marker of astrogliosis, a cellular process possibly involved in Parkinson's disease pathogenesis as elevation of MAOB in astrocytes might be harmful. Since brain monoamine oxidase status in Parkinson's disease is uncertain, our objective was to measure, by quantitative immunoblotting in autopsied brain homogenates, protein levels of both monoamine oxidases in three different degenerative parkinsonian disorders: Parkinson's disease (n = 11), multiple system atrophy (n = 11), and progressive supranuclear palsy (n = 16) and in matched controls (n = 16). We hypothesized that if MAOB is 'substantially' localized to astroglial cells, MAOB levels should be generally associated with standard astroglial protein measures (e.g. glial fibrillary acidic protein). MAOB levels were increased in degenerating putamen (+83%) and substantia nigra (+10%, non-significant) in multiple system atrophy; in caudate (+26%), putamen (+27%), frontal cortex (+31%) and substantia nigra (+23%) of progressive supranuclear palsy; and in frontal cortex (+33%), but not in substantia nigra of Parkinson's disease, a region we previously reported no increase in astrocyte protein markers. Although the magnitude of MAOB increase was less than those of standard astrocytic markers, significant positive correlations were observed amongst the astrocyte proteins and MAOB. Despite suggestions that MAOA (versus MAOB) is primarily responsible for metabolism of dopamine in dopamine neurons, there was no loss of the

ASIC-like currents in freshly isolated cerebral artery smooth muscle cells are inhibited by endogenous oxidase activity.

Science.gov (United States)

Chung, Wen-Shuo; Farley, Jerry M; Drummond, Heather A

2011-01-01

The aim of this study was to determine if VSMC ASIC-like currents are regulated by oxidative state. We used whole-cell patch clamp of isolated mouse cerebral VSMCs to determine if 1) reducing agents, such as DTT and GSH, and 2) inhibition of endogenous oxidase activity from NADPH and Xanthine oxidases potentiate active currents and activate electrically silent currents. Pretreatment with 2 mM DTT or GSH, increased the mean peak amplitude of ASIC-like currents evoked by pH 6.0 from 0.4 ± 0.1 to 14.9 ± 3.6 pA/pF, and from 0.9 ± 0.3 to 11.3 ± 2.4 pA/pF, respectively. Pretreatment with apocynin, a NADPH oxidase inhibitor, mimics the effect of the reducing agents, with the mean peak current amplitude increased from 0.9 ± 0.5 to 7.0 ± 2.6 pA/pF and from 0.5 ± 0.2 to 26.4 ± 6.8 pA/pF by 50 and 200 μM apocynin, respectively. Pretreatment with allopurinol, a xanthine oxidase inhibitor, also potentiates the VSMC ASIC-like activity. These findings suggest that VSMC ASIC-like channels are regulated by oxidative state and may be inhibited by basal endogenous oxidative sources such as NADPH and xanthine oxidase. Copyright © 2011 S. Karger AG, Basel.

Season-controlled changes in biochemical constituents and oxidase enzyme activities in tomato (Lycopersicon esculentum Mill.).

Science.gov (United States)

Sen, Supatra; Mukherji, S

2009-07-01

Season-controlled changes in biochemical constituents viz. carotenoids (carotene and xanthophyll) and pectic substances along with IAA-oxidase and polyphenol oxidase (PPO) enzyme activities were estimated/assayed in leaves of Lycopersicon esculentum Mill. (tomato) in two developmental stages--pre-flowering (35 days after sowing) and post-flowering (75 days after sowing) in three different seasons--summer rainy and winter Carotenoid content along with pectic substances were highest in winter and declined significantly in summer followed by rainy i.e. winter > summer > rainy. Carotenoid content was significantly higher in the pre-flowering as compared to post-flowering in all three seasons while pectic substances increased in the post-flowering as compared to pre-flowering throughout the annual cycle. IAA oxidase and PPO enzyme activities were enhanced in rainy and decreased sharply in summer and winter i.e. rainy > summer > winter. Both the enzymes exhibited higher activity in the post-flowering stage as compared to pre-flowering in all three seasons. These results indicate winter to be the most favourable season for tomato plants while rainy season environmental conditions prove to be unfavourable (stressful) with diminished content of carotenoid and pectic substances and low activities of IAA oxidase and PPO, ultimately leading to poor growth and productivity.

Electrochemical L-Lactic Acid Sensor Based on Immobilized ZnO Nanorods with Lactate Oxidase

Directory of Open Access Journals (Sweden)

Kimleang Khun

2012-02-01

Full Text Available In this work, fabrication of gold coated glass substrate, growth of ZnO nanorods and potentiometric response of lactic acid are explained. The biosensor was developed by immobilizing the lactate oxidase on the ZnO nanorods in combination with glutaraldehyde as a cross linker for lactate oxidase enzyme. The potentiometric technique was applied for the measuring the output (EMF response of L-lactic acid biosensor. We noticed that the present biosensor has wide linear detection range of concentration from 1 × 10−4–1 × 100 mM with acceptable sensitivity about 41.33 ± 1.58 mV/decade. In addition, the proposed biosensor showed fast response time less than 10 s, a good selectivity towards L-lactic acid in presence of common interfering substances such as ascorbic acid, urea, glucose, galactose, magnesium ions and calcium ions. The present biosensor based on immobilized ZnO nanorods with lactate oxidase sustained its stability for more than three weeks.

Electrochemical L-lactic acid sensor based on immobilized ZnO nanorods with lactate oxidase.

Science.gov (United States)

Ibupoto, Zafar Hussain; Shah, Syed Muhammad Usman Ali; Khun, Kimleang; Willander, Magnus

2012-01-01

In this work, fabrication of gold coated glass substrate, growth of ZnO nanorods and potentiometric response of lactic acid are explained. The biosensor was developed by immobilizing the lactate oxidase on the ZnO nanorods in combination with glutaraldehyde as a cross linker for lactate oxidase enzyme. The potentiometric technique was applied for the measuring the output (EMF) response of l-lactic acid biosensor. We noticed that the present biosensor has wide linear detection range of concentration from 1 × 10(-4)-1 × 10(0) mM with acceptable sensitivity about 41.33 ± 1.58 mV/decade. In addition, the proposed biosensor showed fast response time less than 10 s, a good selectivity towards l-lactic acid in presence of common interfering substances such as ascorbic acid, urea, glucose, galactose, magnesium ions and calcium ions. The present biosensor based on immobilized ZnO nanorods with lactate oxidase sustained its stability for more than three weeks.

Serum xanthine oxidase profile in stressed Marwari sheep from arid tracts in India

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Maan R.

2012-08-01

Full Text Available The present investigation was aimed to determine serum xanthine oxidase profile in stressed Marwari breed of sheep belonging to arid tracts in Rajasthan, India. Extreme hot and cold ambiences were considered as stress conditions to the animals. Blood samples were collected to obtain sera during moderate, extreme hot and cold ambiences. The mean value of serum xanthine oxidase during moderate ambience was 93.33±1.11 mU L-1.The mean value of serum xanthine oxidase was significantly (p≤0.05higher during hot and significantly (p≤0.05 lower during cold ambiences as compared to moderate mean value serving as control. The sex and age effects were significant (p≤0.05 in all ambiences. The mean values were significantly (p≤0.05 higher in males than females. In each ambience the age effect showed a significant (p≤0.05 increase in the mean values being highest in the animals of 2.5-4.5 years of age. The effects of extreme ambiences were observed on the male and female animals of all age groups as revealed by various interactions studied viz. ambience X age; ambience X sex and age X sex (p≤0.01. Further sex effect was present in the animals of each age group. It can be concluded that serum xanthine oxidase can be used as an effective marker to assess oxidative stress in these animals. Mean values obtained from large number of animals during moderate ambience will help in providing physiological reference values for future research and clinical interpretations.

Desbalanço redox: NADPH oxidase como um alvo terapêutico no manejo cardiovascular Desbalance redox: NADPH oxidasa como un objetivo terapéutico en el manejo cardiovascular Redox unbalance: NADPH oxidase as therapeutic target in blood pressure control

Directory of Open Access Journals (Sweden)

Luiza A. Rabêlo

2010-05-01

la mayor fuente de esas especies reactivas en la vasculatura. En condiciones fisiológicas, la formación y eliminación de esas sustancias aparecen balanceadas en la pared vascular. Durante el desbalance redox, sin embargo, hay un aumento en la actividad de la NADPH oxidasa y predominio de agentes prooxidantes, superando la capacidad de defensa orgánica antioxidante. Además de ello, tal hiperactividad enzimática reduce la biodisponibilidad del óxido nítrico, crucial para la vasodilatación y el mantenimiento de la función vascular normal. A pesar de que la NADPH oxidasa se relaciona directamente con la disfunción endotelial, fue primeramente descrita por su expresión en fagocitos, en los que su actividad enzimática determina la eficacia de los mecanismos de defensa orgánica contra patógenos. Las diferencias sutiles entre las unidades estructurales de las NADPH oxidasas, dependiendo del tipo celular que las expresa, pueden tener implicaciones terapéuticas, permitiendo la inhibición selectiva del desequilibrio redox inducido por la NADPH oxidasa, sin comprometer, con todo, su participación en las vías fisiológicas de señalización celular que garantizan la protección contra microorganismos.Several studies refer to reactive oxygen and nitrogen species (RONS as important agents in the pathogenesis of a number of heart diseases, including high blood pressure, arteriosclerosis and heart failure. Such species are highly bioactive molecules and a short life due chiefly to reduction of molecular oxygen. The enzyme complex of NADPH oxidase is the main source of these reactive species in vascular system. Under physiological conditions, formation and elimination of these substances seem balanced in vascular wall. During redox Unbalance, nonetheless, there is increase in NADPH oxidase activity and predominance of pro-oxidizing agents, surpassing the anti-oxidant capacity of the organism self-defense. Besides this, such enzyme hyperactivity reduces the

Isolation and purification of pyranose 2-oxidase from Phanerochaete chrysosporium and characterization of gene structure and regulation

Science.gov (United States)

Theodorus H. de Koker; Michael D. Mozuch; Daniel Cullen; Jill Gaskell; Philip J. Kersten

2004-01-01

Pyranose 2-oxidase (POX) was recovered from Phanerochaete chrysosporium BKM-F-1767 solid substrate culture using mild extraction conditions and was purified. 13C-nuclear magnetic resonance confirmed production of D- arabino -hexos-2-ulose (glucosone) from D-glucose with the oxidase. Peptide fingerprints generated by liquid chromatography-tandem mass spectrometry of...

Arthrobacter P 1, a Fast Growing Versatile Methylotroph with Amine Oxidase as a Key Enzyme in the Metabolism of Methylated Amines

NARCIS (Netherlands)

Dijken, J.P. van; Veenhuis, M.; Harder, W.

1981-01-01

A facultative methylotrophic bacterium was isolated from enrichment cultures containing methylamine as the sole carbon source. It was tentatively identified as an Arthrobacter species. Extracts of cells grown on methylamine or ethylamine contained high levels of amine oxidase (E.C. 1.4.3) activity.

Cytokinin oxidase or dehydrogenase? Mechanism of cytokinin degradation in cereals

DEFF Research Database (Denmark)

Galuszka, P.; Frebort, I.; Sebela, M.

2001-01-01

An enzyme degrading cytokinins with isoprenoid side chain, previously named cytokinin oxidase, was purified to near homogeneity from wheat and barley grains. New techniques were developed for the enzyme activity assay and staining on native electrophoretic gels to identify the protein. The purifi...

Lysyl oxidases regulate fibrillar collagen remodelling in idiopathic pulmonary fibrosis

NARCIS (Netherlands)

Tjin, Gavin; White, Eric S; Faiz, Alen; Sicard, Delphine; Tschumperlin, Daniel J; Mahar, Annabelle; Kable, Eleanor P W; Burgess, Janette K

2017-01-01

Idiopathic pulmonary fibrosis (IPF) is a progressive scarring disease of the lung with feweffective therapeutic options. Structural remodelling of the extracellular matrix [i.e. collagen cross-linkingmediated by the lysyl oxidase (LO) family of enzymes (LOX, LOXL1-4)] might contribute to disease

DNA damage and decrease of cellular oxidase activity in piglet ...

African Journals Online (AJOL)

DNA damage and decrease of cellular oxidase activity in piglet sertoli cells exposed to gossypol. Ming Zhang, Hui Yuan, Zuping He, Liyun Yuan, Jine Yi, Sijun Deng, Li Zhu, Chengzhi Guo, Yin Lu, Jing Wu, Lixin Wen, Qiang Wei, Liqun Xue ...

A novel L-amino acid oxidase from Trichoderma harzianum ETS 323 associated with antagonism of Rhizoctonia solani.

Science.gov (United States)

Yang, Chia-Ann; Cheng, Chi-Hua; Lo, Chaur-Tsuen; Liu, Shu-Ying; Lee, Jeng-Woei; Peng, Kou-Cheng

2011-05-11

Trichoderma spp. are used as biocontrol agents against phytopathogens such as Rhizoctonia solani, but their biocontrol mechanisms are poorly understood. A novel L-amino oxidase (Th-LAAO) was identified from the extracellular proteins of Trichoderma harzianum ETS 323. Here, we show a FAD-binding glycoprotein with the best substrate specificity constant for L-phenylalanine. Although the amino acid sequence of Th-LAAO revealed limited homology (16-24%) to other LAAO members, a highly conserved FAD-binding motif was identified in the N-terminus. Th-LAAO was shown to be a homodimeric protein, but the monomeric form was predominant when grown in the presence of deactivated Rhizoctonia solani. Furthermore, in vitro assays demonstrated that Th-LAAO had an antagonistic effect against Rhizoctonia solani and a stimulatory one on hyphal density and sporulation in T. harzianum ETS 323. These findings further our understanding of T. harzianum as a biocontrol agent and provide insight into the biological function of l-amino acid oxidase.

NADPH Oxidase Activity in Cerebral Arterioles Is a Key Mediator of Cerebral Small Vessel Disease—Implications for Prevention

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Mark F. McCarty

2015-04-01

Full Text Available Cerebral small vessel disease (SVD, a common feature of brain aging, is characterized by lacunar infarcts, microbleeds, leukoaraiosis, and a leaky blood-brain barrier. Functionally, it is associated with cognitive decline, dementia, depression, gait abnormalities, and increased risk for stroke. Cerebral arterioles in this syndrome tend to hypertrophy and lose their capacity for adaptive vasodilation. Rodent studies strongly suggest that activation of Nox2-dependent NADPH oxidase activity is a crucial driver of these structural and functional derangements of cerebral arterioles, in part owing to impairment of endothelial nitric oxide synthase (eNOS activity. This oxidative stress may also contribute to the breakdown of the blood-brain barrier seen in SVD. Hypertension, aging, metabolic syndrome, smoking, hyperglycemia, and elevated homocysteine may promote activation of NADPH oxidase in cerebral arterioles. Inhibition of NADPH oxidase with phycocyanobilin from spirulina, as well as high-dose statin therapy, may have potential for prevention and control of SVD, and high-potassium diets merit study in this regard. Measures which support effective eNOS activity in other ways—exercise training, supplemental citrulline, certain dietary flavonoids (as in cocoa and green tea, and capsaicin, may also improve the function of cerebral arterioles. Asian epidemiology suggests that increased protein intakes may decrease risk for SVD; conceivably, arginine and/or cysteine—which boosts tissue glutathione synthesis, and can be administered as N-acetylcysteine—mediate this benefit. Ameliorating the risk factors for SVD—including hypertension, metabolic syndrome, hyperglycemia, smoking, and elevated homocysteine—also may help to prevent and control this syndrome, although few clinical trials have addressed this issue to date.

Lactate Biosensor Based on Cellulose Acetate Membrane Bound Lactate Oxidase

Directory of Open Access Journals (Sweden)

Suman

2007-05-01

Full Text Available Lactate biosensor was fabricated by immobilizing lactate oxidase in cellulose acetate membrane and by mounting over the sensing part of Pt electrode (working and connected to Ag/AgCl electrode (reference along with auxillary electrode through potentiostat. The enzyme electrode was anodically polarized at +400 mV to generate electrons from H2O2, which was formed from oxidation of serum lactate by immobilized lactate oxidase. The minimum detection limit of the electrode was 0.1mmoles/L and sensitivity of the sensor was 0.008 mA/mM/L lactate. Assay coefficients of variation were < 2% .A good correlation (r=0.99 was found between lactate values obtained by colorimetric method and lactate biosensor. The self-life of the biosensor was 18 days at 4ºC and enzyme electrode can be re-used 150 times without any significant loss in enzyme activity.

Involvement of NADH Oxidase in Biofilm Formation in Streptococcus sanguinis.

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Xiuchun Ge

Full Text Available Biofilms play important roles in microbial communities and are related to infectious diseases. Here, we report direct evidence that a bacterial nox gene encoding NADH oxidase is involved in biofilm formation. A dramatic reduction in biofilm formation was observed in a Streptococcus sanguinis nox mutant under anaerobic conditions without any decrease in growth. The membrane fluidity of the mutant bacterial cells was found to be decreased and the fatty acid composition altered, with increased palmitic acid and decreased stearic acid and vaccenic acid. Extracellular DNA of the mutant was reduced in abundance and bacterial competence was suppressed. Gene expression analysis in the mutant identified two genes with altered expression, gtfP and Idh, which were found to be related to biofilm formation through examination of their deletion mutants. NADH oxidase-related metabolic pathways were analyzed, further clarifying the function of this enzyme in biofilm formation.

Boosting the oxidase mimicking activity of nanoceria by fluoride capping: rivaling protein enzymes and ultrasensitive F- detection

Science.gov (United States)

Liu, Biwu; Huang, Zhicheng; Liu, Juewen

2016-07-01

Nanomaterial-based enzyme mimics (nanozymes) are currently a new forefront of chemical research. However, the application of nanozymes is limited by their low catalytic activity and low turnover numbers. Cerium dioxide nanoparticles (nanoceria) are among the few with oxidase activity. Herein, we report an interesting finding addressing their limitations. The oxidase activity of nanoceria is improved by over 100-fold by fluoride capping, making it more close to real oxidases. The turnover number reached 700 in 15 min, drastically improved from ~15 turnovers for the naked particles. The mechanism is attributed to surface charge modulation and facilitated electron transfer by F- capping based on ζ-potential and free radical measurements. Ultrasensitive sensing of fluoride was achieved with a detection limit of 0.64 μM F- in water and in toothpastes, while no other tested anions can achieve the activity enhancement.Nanomaterial-based enzyme mimics (nanozymes) are currently a new forefront of chemical research. However, the application of nanozymes is limited by their low catalytic activity and low turnover numbers. Cerium dioxide nanoparticles (nanoceria) are among the few with oxidase activity. Herein, we report an interesting finding addressing their limitations. The oxidase activity of nanoceria is improved by over 100-fold by fluoride capping, making it more close to real oxidases. The turnover number reached 700 in 15 min, drastically improved from ~15 turnovers for the naked particles. The mechanism is attributed to surface charge modulation and facilitated electron transfer by F- capping based on ζ-potential and free radical measurements. Ultrasensitive sensing of fluoride was achieved with a detection limit of 0.64 μM F- in water and in toothpastes, while no other tested anions can achieve the activity enhancement. Electronic supplementary information (ESI) available: Methods, TMB oxidation kinetics and control experiments. See DOI: 10.1039/c6nr02730j

Renal denervation attenuates NADPH oxidase-mediated oxidative stress and hypertension in rats with hydronephrosis

DEFF Research Database (Denmark)

Peleli, Maria; Al-Mashhadi, Ammar; Yang, Ting

2016-01-01

Hydronephrosis is associated with development of salt-sensitive hypertension. Studies suggest that increased sympathetic nerve activity (SNA) and oxidative stress play important roles in renovascular hypertension. This study aimed to investigate the link between renal SNA and NADPH oxidase (NOX......) regulation in the development of hypertension in rats with hydronephrosis. Hydronephrosis was induced by partial unilateral ureteral obstruction (PUUO) in young rats. Sham surgery or renal denervation was performed at the same time. Blood pressure was measured during normal, high and low salt diets. Renal...

Electronegativity of aromatic amines as a basis for the development of ground state inhibitors of lysyl oxidase

International Nuclear Information System (INIS)

Williamson, P.R.; Kagan, H.M.

1987-01-01

Benzylamine derivatives containing para substituents of differing electronegativity as well as isomers of aminomethylpyridine have been assessed for their substrate and inhibitor potentials toward lysyl oxidase. Substituted benzylamines with increasingly electronegative para substituents had the lowest KI values and thus were the most effective inhibitors of the oxidation of elastin by lysyl oxidase. The kcat values for these compounds as substrates of lysyl oxidase were also reduced with increasingly electronegative para substituents. Both the Dkcat and D(kcat/Km) kinetic isotope effects decreased with increasingly electronegative p-substituents in [alpha, alpha'- 2 H]benzylamines. In contrast, there was no Dkcat solvent isotope effect with [ 2 H] H 2 O while the D(kcat/Km) solvent isotope effect tended to increase with increasingly electronegative p-substituents. These results are consistent with the stabilization of an enzyme-generated substrate carbanion and the retardation of substrate oxidation by electronegative substituents. Such ground state stabilization can result in compounds with increased potential for the inhibition of the oxidation of protein substrates of lysyl oxidase

NADPH oxidase-mediated generation of reactive oxygen species: A new mechanism for X-ray-induced HeLa cell death

International Nuclear Information System (INIS)

Liu Qing; He Xiaoqing; Liu Yongsheng; Du Bingbing; Wang Xiaoyan; Zhang Weisheng; Jia Pengfei; Dong Jingmei; Ma Jianxiu; Wang Xiaohu; Li Sha; Zhang Hong

2008-01-01

Oxidative damage is an important mechanism in X-ray-induced cell death. Radiolysis of water molecules is a source of reactive oxygen species (ROS) that contribute to X-ray-induced cell death. In this study, we showed by ROS detection and a cell survival assay that NADPH oxidase has a very important role in X-ray-induced cell death. Under X-ray irradiation, the upregulation of the expression of NADPH oxidase membrane subunit gp91 phox was dose-dependent. Meanwhile, the cytoplasmic subunit p47 phox was translocated to the cell membrane and localized with p22 phox and gp91 phox to form reactive NADPH oxidase. Our data suggest, for the first time, that NADPH oxidase-mediated generation of ROS is an important contributor to X-ray-induced cell death. This suggests a new target for combined gene transfer and radiotherapy.

H2O2 and NADPH oxidases involve in regulation of 2-(2-phenylethyl)chromones accumulation during salt stress in Aquilaria sinensis calli.

Science.gov (United States)

Wang, Xiaohui; Dong, Xianjuan; Feng, Yingying; Liu, Xiao; Wang, Jinling; Zhang, Zhongxiu; Li, Jun; Zhao, Yunfang; Shi, Shepo; Tu, Pengfei

2018-04-01

2-(2-Phenylethyl)chromones are the main compounds responsible for the quality of agarwood, which is widely used in traditional medicines, incenses and perfumes. H 2 O 2 and NADPH oxidases (also known as respiratory burst oxidase homologs, Rbohs) mediate diverse physiological and biochemical processes in environmental stress responses. However, little is known about the function of H 2 O 2 and NADPH oxidases in 2-(2-phenylethyl)chromones accumulation. In this study, we found that salt stress induced a transient increase in content of H 2 O 2 and 2-(2-phenylethyl)chromones accumulation in Aquilaria sinensis calli. Exogenous H 2 O 2 remarkably decreased the production of 2-(2-phenylethyl)chromones, while dimethylthiourea (DMTU), a scavenger of H 2 O 2 , significantly increased 2-(2-phenylethyl)chromones accumulation in salt treated calli. Three new H 2 O 2 -generating genes, named AsRbohA-C, were isolated and characterized from A. sinensis. Salt stress also induced a transient increase in AsRbohA-C expression and NADPH oxidase activity. Furthermore, exogenous H 2 O 2 increased AsRbohA-C expression and NADPH oxidase activity, while DMTU inhibited AsRbohA-C expression and NADPH oxidase activity under salt stress. Moreover, diphenylene iodonium (DPI), the inhibitor of NADPH oxidases, reduced AsRbohA-C expression and NADPH oxidase activity, but significantly induced 2-(2-phenylethyl)chromones accumulation during salt stress. These results clearly demonstrated the central role of H 2 O 2 and NADPH oxidases in regulation of salt-induced 2-(2-phenylethyl)chromones accumulation in A. sinensis calli. Copyright © 2018 Elsevier B.V. All rights reserved.

Effect of heat treatment on polyphenol oxidase and peroxidase ...

African Journals Online (AJOL)

Effect of heat treatment (55°C/20 min) on polyphenol oxidase (PPO) and peroxidase (POD) activities and total phenolic compounds was investigated in Algerian dates (Deglet Nour variety) at Tamar (fully ripe) stage and in dates stored for 5 months at ambient temperature and in cold storage (10°C). Results obtained ...

High-performance liquid chromatography coupled with post-column dual-bioactivity assay for simultaneous screening of xanthine oxidase inhibitors and free radical scavengers from complex mixture.

Science.gov (United States)

Li, D Q; Zhao, J; Li, S P

2014-06-06

Xanthine oxidase (XO) can catalyze hypoxanthine and xanthine to generate uric acid and reactive oxygen species (ROS), including superoxide anion radical (O₂(•-)) and hydrogen peroxide. XO inhibitors and free radical scavengers are beneficial to the treatment of gout and many related diseases. In the present study, an on-line high-performance liquid chromatography (HPLC) coupled with post-column dual-bioactivity assay was established and successfully applied to simultaneously screening of XO inhibitors and free radical scavengers from a complex mixture, Oroxylum indicum extract. The integrated system of HPLC separation, bioactivity screening and mass spectrometry identification was proved to be simple and effective for rapid and sensitive screening of individual bioactive compounds in complex mixtures. Copyright © 2014 Elsevier B.V. All rights reserved.

Structure of choline oxidase in complex with the reaction product glycine betaine.

Science.gov (United States)

Salvi, Francesca; Wang, Yuan-Fang; Weber, Irene T; Gadda, Giovanni

2014-02-01

Choline oxidase from Arthrobacter globiformis, which is involved in the biosynthesis of glycine betaine from choline, has been extensively characterized in its mechanistic and structural properties. Despite the knowledge gained on the enzyme, the details of substrate access to the active site are not fully understood. The `loop-and-lid' mechanism described for the glucose-methanol-choline enzyme superfamily has not been confirmed for choline oxidase. Instead, a hydrophobic cluster on the solvent-accessible surface of the enzyme has been proposed by molecular dynamics to control substrate access to the active site. Here, the crystal structure of the enzyme was solved in complex with glycine betaine at pH 6.0 at 1.95 Å resolution, allowing a structural description of the ligand-enzyme interactions in the active site. This structure is the first of choline oxidase in complex with a physiologically relevant ligand. The protein structures with and without ligand are virtually identical, with the exception of a loop at the dimer interface, which assumes two distinct conformations. The different conformations of loop 250-255 define different accessibilities of the proposed active-site entrance delimited by the hydrophobic cluster on the other subunit of the dimer, suggesting a role in regulating substrate access to the active site.

Liquid-Phase Packaging of a Glucose Oxidase Solution with Parylene Direct Encapsulation and an Ultraviolet Curing Adhesive Cover for Glucose Sensors

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Seiichi Takamatsu

2010-06-01

Full Text Available We have developed a package for disposable glucose sensor chips using Parylene encapsulation of a glucose oxidase solution in the liquid phase and a cover structure made of an ultraviolet (UV curable adhesive. Parylene was directly deposited onto a small volume (1 μL of glucose oxidase solution through chemical vapor deposition. The cover and reaction chamber were constructed on Parylene film using a UV-curable adhesive and photolithography. The package was processed at room temperature to avoid denaturation of the glucose oxidase. The glucose oxidase solution was encapsulated and unsealed. Glucose sensing was demonstrated using standard amperometric detection at glucose concentrations between 0.1 and 100 mM, which covers the glucose concentration range of diabetic patients. Our proposed Parylene encapsulation and UV-adhesive cover form a liquid phase glucose-oxidase package that has the advantages of room temperature processing and direct liquid encapsulation of a small volume solution without use of conventional solidifying chemicals.

Hofmeister effects on the glucose oxidase hydrogel-modified electrode

International Nuclear Information System (INIS)

Suzuki, Aimi; Tsujimura, Seiya

2016-01-01

We describe the consistent effect of salts in the electrolyte solution on glucose oxidation current production in the redox hydrogel-modified electrode containing glucose oxidase as an electrocatalyst and Os complex mediator. The ions affect not only on the electron transfer between the enzyme and the Os complex, but also on the hydrogel structure. This study found that the degree of the effect can be characterized by Hofmeister series. The relative decrease in oxidization current is the lowest in the middle of the Hofmeister series, and increases monotonically on either side. An increase of ionic strength inhibits the electron transfer from the active site of glucose oxidase to Os complex. In addition to this, the kosmotropic anions, which are strongly hydrated, caused hydrogel deswelling (shrinking). The more chaotropic an ion is, the more it adsorbs to uncharged parts of polymer/enzyme with dispersion force, and the swelling of the hydrogel decreases the catalytic current. This study impacts the design of hydrogel electrode and selection of electrolyte ions for bioelectronic applications.

Lysyl oxidase overexpression accelerates cardiac remodeling and aggravates angiotensin II-induced hypertrophy.

Science.gov (United States)

Galán, María; Varona, Saray; Guadall, Anna; Orriols, Mar; Navas, Miquel; Aguiló, Silvia; de Diego, Alicia; Navarro, María A; García-Dorado, David; Rodríguez-Sinovas, Antonio; Martínez-González, José; Rodriguez, Cristina

2017-09-01

Lysyl oxidase (LOX) controls matrix remodeling, a key process that underlies cardiovascular diseases and heart failure; however, a lack of suitable animal models has limited our knowledge with regard to the contribution of LOX to cardiac dysfunction. Here, we assessed the impact of LOX overexpression on ventricular function and cardiac hypertrophy in a transgenic LOX (TgLOX) mouse model with a strong cardiac expression of human LOX. TgLOX mice exhibited high expression of the transgene in cardiomyocytes and cardiofibroblasts, which are associated with enhanced LOX activity and H 2 O 2 production and with cardiofibroblast reprogramming. LOX overexpression promoted an age-associated concentric remodeling of the left ventricle and impaired diastolic function. Furthermore, LOX transgenesis aggravated angiotensin II (Ang II)-induced cardiac hypertrophy and dysfunction, which triggered a greater fibrotic response that was characterized by stronger collagen deposition and cross-linking and high expression of fibrotic markers. In addition, LOX transgenesis increased the Ang II-induced myocardial inflammatory infiltrate, exacerbated expression of proinflammatory markers, and decreased that of cardioprotective factors. Mechanistically, LOX overexpression enhanced oxidative stress and potentiated the Ang II-mediated cardiac activation of p38 MAPK while reducing AMPK activation. Our findings suggest that LOX induces an age-dependent disturbance of diastolic function and aggravates Ang II-induced hypertrophy, which provides novel insights into the role of LOX in cardiac performance.-Galán, M., Varona, S., Guadall, A., Orriols, M., Navas, M., Aguiló, S., de Diego, A., Navarro, M. A., García-Dorado, D., Rodríguez-Sinovas, A., Martínez-González, J., Rodriguez, C. Lysyl oxidase overexpression accelerates cardiac remodeling and aggravates angiotensin II-induced hypertrophy. © FASEB.

Suicide attempts, platelet monoamine oxidase and the average evoked response

International Nuclear Information System (INIS)

Buchsbaum, M.S.; Haier, R.J.; Murphy, D.L.

1977-01-01

The relationship between suicides and suicide attempts and two biological measures, platelet monoamine oxidase levels (MAO) and average evoked response (AER) augmenting was examined in 79 off-medication psychiatric patients and in 68 college student volunteers chosen from the upper and lower deciles of MAO activity levels. In the patient sample, male individuals with low MAO and AER augmenting, a pattern previously associated with bipolar affective disorders, showed a significantly increased incidence of suicide attempts in comparison with either non-augmenting low MAO or high MAO patients. Within the normal volunteer group, all male low MAO probands with a family history of suicide or suicide attempts were AER augmenters themselves. Four completed suicides were found among relatives of low MAO probands whereas no high MAO proband had a relative who committed suicide. These findings suggest that the combination of low platelet MAO activity and AER augmenting may be associated with a possible genetic vulnerability to psychiatric disorders. (author)

Substrate specificity of flavin-dependent vanillyl-alcohol oxidase from Penicillium simplicissimum.Evidence for the production of 4-hydroxycinnamyl alcohols from 4-allylphenols

OpenAIRE

Fraaije, Marco W.; Veeger, Cees; Berkel, Willem J.H. van

1995-01-01

The substrate specificity of the flavoprotein vanillyl-alcohol oxidase from Penicillium simplicissimum was investigated. Vanillyl-alcohol oxidase catalyzes besides the oxidation of 4-hydroxybenzyl alcohols, the oxidative deamination of 4-hydroxybenzylamines and the oxidative demethylation of 4-(methoxymethyl)phenols. During the conversion of vanillylamine to vanillin, a transient intermediate, most probably vanillylimine, is observed. Vanillyl-alcohol oxidase weakly interacts with 4-hydroxyph...

Effect of retinal impulse blockage on cytochrome oxidase-poor interpuffs in the macaque striate cortex: quantitative EM analysis of neurons.

Science.gov (United States)

Wong-Riley, M T; Trusk, T C; Kaboord, W; Huang, Z

1994-09-01

One of the hallmarks of the primate striate cortex is the presence of cytochrome oxidase-rich puffs in its supragranular layers. Neurons in puffs have been classified as type A, B, and C in ascending order of cytochrome oxidase content, with type C cells being the most vulnerable to retinal impulse blockade. The present study aimed at analysing cytochrome oxidase-poor interpuffs with reference to their metabolic cell types and the effect of intraretinal tetrodotoxin treatment. The same three metabolic types were found in interpuffs, except that type B and C neurons were smaller and less cytochrome oxidase-reactive in interpuffs than in puffs. Type A neurons had small perikarya, low levels of cytochrome oxidase, and received exclusively symmetric axosomatic synapses. The largest neurons were pyramidal, type B cells with moderate cytochrome oxidase activity and were also contacted exclusively by symmetric axosomatic synapses. Type C cells medium-sized with a rich supply of large, darkly reactive mitochondria and possessed all the characteristics of GABAergic neurons. They were the only cell type that received both symmetric and asymmetric axosomatic synapses. Two weeks of monocular tetrodotoxin blockade in adult monkeys caused all three major cell types in deprived interpuffs to suffer a significant downward shift in the size and cytochrome oxidase reactivity of their mitochondria, but the effects were more severe in type B and C neurons. In nondeprived interpuffs, all three cell types gained both in size and absolute number of mitochondria, and type A cells also had an elevated level of cytochrome oxidase, indicating that they might be functioning at a competitive advantage over cells in deprived columns. However, type B and C neurons showed a net loss of darkly reactive mitochondria, indicating that these cells became less active. Thus, mature interpuff neurons remained vulnerable to retinal impulse blockade and the metabolic capacity of these cells remains tightly

Characterization of polyphenol oxidase from blueberry (Vaccinium corymbosum L.).

Science.gov (United States)

Siddiq, M; Dolan, K D

2017-03-01

Polyphenol oxidase (PPO) was extracted and characterized from high-bush blueberries. PPO showed an optimum activity at pH 6.1-6.3 and 35°C, with the enzyme showing significant activity over a wide temperature range (25-60°C). Catechol was the most readily oxidized substrate followed by 4-methylcatechol, DL-DOPA, and dopamine. Blueberry PPO showed a K m of 15mM and V max of 2.57 ΔA 420 nm/min×10 -1 , determined with catechol. PPO was completely inactivated in 20min at 85°C, however, after 30minat 75°C it showed about 10% residual activity. Thermal treatment at 55 and 65°C for 30min resulted in the partial inactivation of PPO. Ascorbic acid, sodium diethyldithiocarbamic acid, L-cysteine, and sodium metabisulfite were effective inhibitors of PPO at 1.0mM. Benzoic acid and cinnamic acid series inhibitors showed relatively weak inhibition of PPO (21.8-27.6%), even at as high as 2.0mM concentration. Copyright © 2016 Elsevier Ltd. All rights reserved.

Nox1 oxidase suppresses influenza a virus-induced lung inflammation and oxidative stress.

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Stavros Selemidis

Full Text Available Influenza A virus infection is an ongoing clinical problem and thus, there is an urgent need to understand the mechanisms that regulate the lung inflammation in order to unravel novel generic pharmacological strategies. Evidence indicates that the Nox2-containing NADPH oxidase enzyme promotes influenza A virus-induced lung oxidative stress, inflammation and dysfunction via ROS generation. In addition, lung epithelial and endothelial cells express the Nox1 isoform of NADPH oxidase, placing this enzyme at key sites to regulate influenza A virus-induced lung inflammation. The aim of this study was to investigate whether Nox1 oxidase regulates the inflammatory response and the oxidative stress to influenza infection in vivo in mice. Male WT and Nox1-deficient (Nox1(-/y mice were infected with the moderately pathogenic HkX-31 (H3N2, 1×10(4 PFU influenza A virus for analysis of bodyweight, airways inflammation, oxidative stress, viral titre, lung histopathology, and cytokine/chemokine expression at 3 and 7 days post infection. HkX-31 virus infection of Nox1(-/y mice resulted in significantly greater: loss of bodyweight (Day 3; BALF neutrophilia, peri-bronchial, peri-vascular and alveolar inflammation; Nox2-dependent inflammatory cell ROS production and peri-bronchial, epithelial and endothelial oxidative stress. The expression of pro-inflammatory cytokines including CCL2, CCL3, CXCL2, IL-1β, IL-6, GM-CSF and TNF-α was higher in Nox1(-/y lungs compared to WT mice at Day 3, however, the expression of CCL2, CCL3, CXCL2, IFN-γ and the anti-inflammatory cytokine IL-10 were lower in lungs of Nox1(-/y mice vs. WT mice at Day 7. Lung viral titre, and airways infiltration of active CD8(+ and CD4(+ T lymphocytes, and of Tregs were similar between WT and Nox1(-/y mice. In conclusion, Nox1 oxidase suppresses influenza A virus induced lung inflammation and oxidative stress in mice particularly at the early phases of the infection. Nox1 and Nox2 oxidases appear

Associations Between Genetic Variants of NADPH Oxidase-Related Genes and Blood Pressure Responses to Dietary Sodium Intervention: The GenSalt Study.

Science.gov (United States)

Han, Xikun; Hu, Zunsong; Chen, Jing; Huang, Jianfeng; Huang, Chen; Liu, Fangchao; Gu, Charles; Yang, Xueli; Hixson, James E; Lu, Xiangfeng; Wang, Laiyuan; Liu, De-Pei; He, Jiang; Chen, Shufeng; Gu, Dongfeng

2017-04-01

The aim of this study was to comprehensively test the associations of genetic variants of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-related genes with blood pressure (BP) responses to dietary sodium intervention in a Chinese population. We conducted a 7-day low-sodium intervention followed by a 7-day high-sodium intervention among 1,906 participants in rural China. BP measurements were obtained at baseline and each dietary intervention using a random-zero sphygmomanometer. Linear mixed-effect models were used to assess the additive associations of 63 tag single-nucleotide polymorphisms in 11 NADPH oxidase-related genes with BP responses to dietary sodium intervention. Gene-based analyses were conducted using the truncated product method. The Bonferroni method was used to adjust for multiple testing in all analyses. Systolic BP (SBP) response to high-sodium intervention significantly decreased with the number of minor T allele of marker rs6967221 in RAC1 (P = 4.51 × 10-4). SBP responses (95% confidence interval) for genotypes CC, CT, and TT were 5.03 (4.71, 5.36), 4.20 (3.54, 4.85), and 0.56 (-1.08, 2.20) mm Hg, respectively, during the high-sodium intervention. Gene-based analyses revealed that RAC1 was significantly associated with SBP response to high-sodium intervention (P = 1.00 × 10-6) and diastolic BP response to low-sodium intervention (P = 9.80 × 10-4). These findings suggested that genetic variants of NADPH oxidase-related genes may contribute to the variation of BP responses to sodium intervention in Chinese population. Further replication of these findings is warranted. © American Journal of Hypertension, Ltd 2017. All rights reserved. For Permissions, please email: journals.permissions@oup.com

Evaluation of Fluorine-18-Labeled α1(I-N-Telopeptide Analogs as Substrate-Based Radiotracers for PET Imaging of Melanoma-Associated Lysyl Oxidase

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Manuela Kuchar

2018-04-01

Full Text Available Accumulating evidence suggests an unequivocal role of lysyl oxidases as key players of tumor progression and metastasis, which renders this enzyme family highly attractive for targeted non-invasive functional imaging of tumors. Considering their function in matrix remodeling, malignant melanoma appears as particularly interesting neoplasia in this respect. For the development of radiotracers that enable PET imaging of the melanoma-associated lysyl oxidase activity, substrates derived from the type I collagen α1 N-telopeptide were labeled with fluorine-18 using N-succinimidyl 4-[18F]fluorobenzoate ([18F]SFB as prosthetic reagent. With regards to potential crosslinking to tumor-associated collagen in vivo, their interaction with triple-helical type I collagen was studied by SPR. A mouse model of human melanoma was established on the basis of the A375 cell line, for which the expression of the oncologically relevant lysyl oxidase isoforms LOX and LOXL2 was demonstrated in Western blot and immunohistochemical experiments. The radiopharmacological profiles of the peptidic radiotracers were evaluated in normal rats and A375 melanoma-bearing mice by ex vivo metabolite analysis, whole-body biodistribution studies and dynamic PET imaging. Out of three 18F-labeled telopeptide analogs, the one with the most favorable substrate properties has shown favorable tumor uptake and tumor-to-muscle ratio. Lysyl oxidase-mediated tumor uptake was proven by pharmacological inhibition using β-aminopropionitrile and by employing negative-control analogs of impeded or abolished targeting capability. The latter were obtained by substituting the lysine residue by ornithine and norleucine, respectively. Comparing the tumor uptake of the lysine-containing peptide with that of the non-functional analogs indicate the feasibility of lysyl oxidase imaging in melanoma using substrate-based radiotracers.

Evaluation of Fluorine-18-Labeled α1(I)-N-Telopeptide Analogs as Substrate-Based Radiotracers for PET Imaging of Melanoma-Associated Lysyl Oxidase.

Science.gov (United States)

Kuchar, Manuela; Neuber, Christin; Belter, Birgit; Bergmann, Ralf; Lenk, Jens; Wodtke, Robert; Kniess, Torsten; Steinbach, Jörg; Pietzsch, Jens; Löser, Reik

2018-01-01

Accumulating evidence suggests an unequivocal role of lysyl oxidases as key players of tumor progression and metastasis, which renders this enzyme family highly attractive for targeted non-invasive functional imaging of tumors. Considering their function in matrix remodeling, malignant melanoma appears as particularly interesting neoplasia in this respect. For the development of radiotracers that enable PET imaging of the melanoma-associated lysyl oxidase activity, substrates derived from the type I collagen α1 N-telopeptide were labeled with fluorine-18 using N -succinimidyl 4-[ 18 F]fluorobenzoate ([ 18 F]SFB) as prosthetic reagent. With regards to potential crosslinking to tumor-associated collagen in vivo , their interaction with triple-helical type I collagen was studied by SPR. A mouse model of human melanoma was established on the basis of the A375 cell line, for which the expression of the oncologically relevant lysyl oxidase isoforms LOX and LOXL2 was demonstrated in Western blot and immunohistochemical experiments. The radiopharmacological profiles of the peptidic radiotracers were evaluated in normal rats and A375 melanoma-bearing mice by ex vivo metabolite analysis, whole-body biodistribution studies and dynamic PET imaging. Out of three 18 F-labeled telopeptide analogs, the one with the most favorable substrate properties has shown favorable tumor uptake and tumor-to-muscle ratio. Lysyl oxidase-mediated tumor uptake was proven by pharmacological inhibition using β-aminopropionitrile and by employing negative-control analogs of impeded or abolished targeting capability. The latter were obtained by substituting the lysine residue by ornithine and norleucine, respectively. Comparing the tumor uptake of the lysine-containing peptide with that of the non-functional analogs indicate the feasibility of lysyl oxidase imaging in melanoma using substrate-based radiotracers.

Thermal and pH stabilities of partially purified polyphenol oxidase ...

African Journals Online (AJOL)

Dr. Paul Chidoka Chikezie

2012-07-30

Jul 30, 2012 ... indices of the two PPO extracts is summarized in Table. 1. At the end of the .... activity near neutral pH values (Jime´nez-Atie´nzar et al.,. 2004; Dogan and Dogan, 2004). ..... oxidase from white cherry fruit (Starks gold). GIDA.

Layer by layer assembly of glucose oxidase and thiourea onto glassy carbon electrode: Fabrication of glucose biosensor

Energy Technology Data Exchange (ETDEWEB)

Salimi, Abdollah, E-mail: absalimi@yahoo.com [Department of Chemistry, University of Kurdistsn, P.O. Box 416, Sanandaj (Iran, Islamic Republic of); Research Center for Nanotechnology, University of Kurdistan, P.O. Box 416, Sanandaj (Iran, Islamic Republic of); Noorbakhsh, Abdollah [Department of Chemistry, University of Kurdistsn, P.O. Box 416, Sanandaj (Iran, Islamic Republic of); Department of Nanotechnology Engenering, Faculty of Advanced Science and Technology, University of Isfahan, 81746-73441 (Iran, Islamic Republic of)

2011-07-01

Highlights: > Although various enzymes immobilization have been approve for the construction of glucose biosensor, a layer by layer (LBL) technique has attracted more attention due to simplicity of the procedure, wide choice of materials that can be used, controllability of film thickness and unique mechanical properties. > In this paper, we described a novel and simple strategy for developing an amperometric glucose biosensor based on layer-by-layer self assembly of glucose oxidase on the glassy carbon electrode modified by thiourea. > Thiourea has two amino groups that the one can be immobilized on the activated glassy carbon electrode and the other can be used for the coupling of glucose oxidase enzyme. > The biosensor exhibited good performance for electrocatalytic oxidation of glucose, such as high sensitivity, low detection limit, short response time and wide concentration range. > Finally, the new method is strongly recommended for immobilization of many other enzymes or proteins containing carbaldehyde or carboxylic groups for fabricating third generation biosensors and bioelectronics devices. - Abstract: For the first time a novel, simple and facile approach is described to construct highly stable glucose oxidase (GOx) multilayer onto glassy carbon (GC) electrode using thiourea (TU) as a covalent attachment cross-linker. The layer by layer (LBL) attachment process was confirmed by cyclic voltammetry, electrochemical impedance spectroscopy and Fourier transform infrared reflection spectroscopy (FT-IR-RS) techniques. Immobilized GOx shows excellent electrocatalytic activity toward glucose oxidation using ferrocenemethanol as artificial electron transfer mediator and biosensor response was directly correlated to the number of bilayers. The surface coverage of active GOx per bilayer, heterogeneous electron transfer rate constant (k{sub s}) and Michaelis-Menten constant (K{sub M}), of immobilized GOx were 1.50 x 10{sup -12} mol cm{sup -2}, 9.2 {+-} 0.5 s{sup -1

Layer by layer assembly of glucose oxidase and thiourea onto glassy carbon electrode: Fabrication of glucose biosensor

International Nuclear Information System (INIS)

Salimi, Abdollah; Noorbakhsh, Abdollah

2011-01-01

Highlights: → Although various enzymes immobilization have been approve for the construction of glucose biosensor, a layer by layer (LBL) technique has attracted more attention due to simplicity of the procedure, wide choice of materials that can be used, controllability of film thickness and unique mechanical properties. → In this paper, we described a novel and simple strategy for developing an amperometric glucose biosensor based on layer-by-layer self assembly of glucose oxidase on the glassy carbon electrode modified by thiourea. → Thiourea has two amino groups that the one can be immobilized on the activated glassy carbon electrode and the other can be used for the coupling of glucose oxidase enzyme. → The biosensor exhibited good performance for electrocatalytic oxidation of glucose, such as high sensitivity, low detection limit, short response time and wide concentration range. → Finally, the new method is strongly recommended for immobilization of many other enzymes or proteins containing carbaldehyde or carboxylic groups for fabricating third generation biosensors and bioelectronics devices. - Abstract: For the first time a novel, simple and facile approach is described to construct highly stable glucose oxidase (GOx) multilayer onto glassy carbon (GC) electrode using thiourea (TU) as a covalent attachment cross-linker. The layer by layer (LBL) attachment process was confirmed by cyclic voltammetry, electrochemical impedance spectroscopy and Fourier transform infrared reflection spectroscopy (FT-IR-RS) techniques. Immobilized GOx shows excellent electrocatalytic activity toward glucose oxidation using ferrocenemethanol as artificial electron transfer mediator and biosensor response was directly correlated to the number of bilayers. The surface coverage of active GOx per bilayer, heterogeneous electron transfer rate constant (k s ) and Michaelis-Menten constant (K M ), of immobilized GOx were 1.50 x 10 -12 mol cm -2 , 9.2 ± 0.5 s -1 and 3.42(±0

On-chip highly sensitive saliva glucose sensing using multilayer films composed of single-walled carbon nanotubes, gold nanoparticles, and glucose oxidase

Directory of Open Access Journals (Sweden)

Wenjun Zhang

2015-06-01

Full Text Available It is very important for human health to rapidly and accurately detect glucose levels in biological environments, especially for diabetes mellitus. We proposed a simple, highly sensitive, accurate, convenient, low-cost, and disposable glucose biosensor on a single chip. A working (sensor electrode, a counter electrode, and a reference electrode are integrated on a single chip through micro-fabrication. The working electrode is functionalized through a layer-by-layer (LBL assembly of single-walled carbon nanotubes (SWNTs and multilayer films composed of chitosan (CS, gold nanoparticles (GNp, and glucose oxidase (GOx to obtain high sensitivity and accuracy. The glucose sensor has following features: (1 direct electron transfer between GOx and the electrode surface; (2 on-a-chip; (3 glucose detection down to 0.1 mg/dL (5.6 μM; (4 good sensing linearity over 0.017–0.81 mM; (5 high sensitivity (61.4 μA/mM-cm2 with a small reactive area (8 mm2; (6 fast response; (7 high reproducibility and repeatability; (8 reliable and accurate saliva glucose detection. Thus, this disposable biosensor will be an alternative for real time tracking of glucose levels from body fluids, e.g. saliva, in a noninvasive, pain-free, accurate, and continuous way. In addition to being used as a disposable glucose biosensor, it also provides a suitable platform for on-chip electrochemical sensing for other chemical agents and biomolecules.

Cytokinin oxidase from Phaseolus vulgaris callus tissues. Enhanced in vitro activity of the enzyme in the presence of copper-imidazole complexes

International Nuclear Information System (INIS)

Chatfield, J.M.; Armstrong, D.J.

1987-01-01

The effects of metal ions on cytokinin oxidase activity extracted from callus tissues of Phaseolus vulgaris L. cv Great Northern have been examined using an assay based on the oxidation of N 6 -(Δ 2 -isopentenyl)-adenine-2,8- 3 H (i 6 Ade) to adenine (Ade). The addition of cupric ions to reaction mixtures containing imidazole buffer markedly enhanced cytokinin oxidase activity. In the presence of optimal concentrations of copper and imidazole, cytokinin oxidase activity was stimulated more than 20-fold. The effect was enzyme dependent, specific for copper, and observed only in the presence of imidazole. The substrate specificity of the copper-imidazole enhanced reaction, as judged by substrate competition tests, was the same as that observed in the absence of copper and imidazole. Similarly, in tests involving DEAE-cellulose chromatography, elution profiles of cytokinin oxidase activity determined using a copper-imidazole enhanced assay were identical to those obtained using an assay without copper and imidazole. On the basis of these results, the addition of copper and imidazole to reaction mixtures used to assay for cytokinin oxidase activity is judged to provide a reliable and specific assay of greatly enhanced sensitivity for the enzyme. The mechanism by which copper and imidazole enhance cytokinin oxidase activity is not certain, but the reaction catalyzed by the enzyme was not inhibited by anaerobic conditions when these reagents were present. This observation suggests that copper-imidazole complexes are substituting for oxygen in the reaction mechanism by which cytokinin oxidase effects cleavage of the N 6 -side chain of i 6 Ade

Evaluation of different disinfectants on the performance of an on-meter dosed amperometric glucose-oxidase-based glucose meter.

Science.gov (United States)

Sarmaga, Don; Dubois, Jeffrey A; Lyon, Martha E

2011-11-01

Off-meter dosed photometric glucose-oxidase-based glucose meters have been reported to be susceptible to interference by hydrogen-peroxide-based disinfecting agents. The objective of this study was to determine if a single application of hydrogen-peroxide-containing Accel® wipe to disinfect an on-meter dosed amperometric glucose-oxidase-based glucose meter will influence its performance. The performance of five on-meter dosed amperometric glucose-oxidase-based glucose meters was determined before and after disinfecting the devices with a single application of either CaviWipes® (14.3% isopropanol and 0.23% diisobutyl-phenoxy-ethoxyethyl dimethyl benzyl ammonium chloride) or Accel (0.5% hydrogen peroxide) wipes. Replicate glucose measurements were conducted before disinfecting the devices, immediately after disinfecting, and then 1 and 2 min postdisinfecting, with measurements in triplicate. Analysis was sequentially completed for five different meters. Results were analyzed by a two-way analysis of variance (Analyze-it software). No clinical ( .05) in glucose concentration were detected when the on-meter dosed amperometric glucose-oxidase-based glucose meters were disinfected with either CaviWipes or Accel wipes and measured immediately or 1 or 2 min postdisinfecting. No clinically significant difference in glucose concentration was detected between meters (glucose oxidase amperometric-based glucose meters are not analytically susceptible to interference by a single application of hydrogen-peroxide-containing Accel disinfectant wipes. © 2011 Diabetes Technology Society.

Intermittent hypoxia-induced cognitive deficits are mediated by NADPH oxidase activity in a murine model of sleep apnea.

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Deepti Nair

Full Text Available In rodents, exposure to intermittent hypoxia (IH, a hallmark of obstructive sleep apnea (OSA, is associated with neurobehavioral impairments, increased apoptosis in the hippocampus and cortex, as well as increased oxidant stress and inflammation. Excessive NADPH oxidase activity may play a role in IH-induced CNS dysfunction.The effect of IH during light period on two forms of spatial learning in the water maze and well as markers of oxidative stress was assessed in mice lacking NADPH oxidase activity (gp91phox(_/Y and wild-type littermates. On a standard place training task, gp91phox(_/Y displayed normal learning, and were protected from the spatial learning deficits observed in wild-type littermates exposed to IH. Moreover, anxiety levels were increased in wild-type mice exposed to IH as compared to room air (RA controls, while no changes emerged in gp91phox(_/Y mice. Additionally, wild-type mice, but not gp91phox(_/Y mice had significantly elevated levels of NADPH oxidase expression and activity, as well as MDA and 8-OHDG in cortical and hippocampal lysates following IH exposures.The oxidative stress responses and neurobehavioral impairments induced by IH during sleep are mediated, at least in part, by excessive NADPH oxidase activity, and thus pharmacological agents targeting NADPH oxidase may provide a therapeutic strategy in sleep-disordered breathing.

Cloning and Functional Analysis of the Promoter of an Ascorbate Oxidase Gene from Gossypium hirsutum

OpenAIRE

Xin, Shan; Tao, Chengcheng; Li, Hongbin

2016-01-01

Apoplastic ascorbate oxidase (AO) plays significant roles in plant cell growth. However, the mechanism of underlying the transcriptional regulation of AO in Gossypium hirsutum remains unclear. Here, we obtained a 1,920-bp promoter sequence from the Gossypium hirsutum ascorbate oxidase (GhAO1) gene, and this GhAO1 promoter included a number of known cis-elements. Promoter activity analysis in overexpressing pGhAO1::GFP-GUS tobacco (Nicotiana benthamiana) showed that the GhAO1 promoter exhibite...

The effects of carbon nanotube addition and oxyfluorination on the glucose-sensing capabilities of glucose oxidase-coated carbon fiber electrodes

Energy Technology Data Exchange (ETDEWEB)

Im, Ji Sun; Yun, Jumi; Kim, Jong Gu [Department of Fine Chemical Engineering and Applied Chemistry, BK21-E2 M, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Bae, Tae-Sung [Department of Fine Chemical Engineering and Applied Chemistry, BK21-E2 M, Chungnam National University, Daejeon 305-764 (Korea, Republic of); Korea Basic Science Institute (KBSI), Jeonju 561-756 (Korea, Republic of); Lee, Young-Seak, E-mail: youngslee@cnu.ac.kr [Department of Fine Chemical Engineering and Applied Chemistry, BK21-E2 M, Chungnam National University, Daejeon 305-764 (Korea, Republic of)

2012-01-15

Glucose-sensing electrodes were constructed from carbon fibers by electrospinning and heat treatment. By controlling the pore size, the specific surface area and pore volume of the electrospun carbon fibers were increased for efficient immobilization of the glucose oxidase. Carbon nanotubes were embedded as an electrically conductive additive to improve the electrical property of the porous carbon fibers. In addition, the surface of the porous carbon fibers was modified with hydrophilic functional groups by direct oxyfluorination to increase the affinity between the hydrophobic carbon surface and the hydrophilic glucose oxidase molecules. The porosity of the carbon fibers was improved significantly with approximately 28- and 35-fold increases in the specific surface area and pore volume, respectively. The number of chemical bonds between carbon and oxygen were increased with higher oxygen content during oxyfluorination based on the X-ray photoelectron spectroscopy results. Glucose sensing was carried out by current voltagram and amperometric methods. A high-performance glucose sensor was obtained with high sensitivity and rapid response time as a result of carbon nanotube addition, physical activation and surface modification. The mechanism of the highly sensitive prepared glucose sensor was modeled by an enzyme kinetics study using the Michaelis-Menten equation.

Critical role of NADPH oxidase in neuronal oxidative damage and microglia activation following traumatic brain injury.

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Quan-Guang Zhang

Full Text Available BACKGROUND: Oxidative stress is known to play an important role in the pathology of traumatic brain injury. Mitochondria are thought to be the major source of the damaging reactive oxygen species (ROS following TBI. However, recent work has revealed that the membrane, via the enzyme NADPH oxidase can also generate the superoxide radical (O(2(-, and thereby potentially contribute to the oxidative stress following TBI. The current study thus addressed the potential role of NADPH oxidase in TBI. METHODOLOGY/PRINCIPAL FINDINGS: The results revealed that NADPH oxidase activity in the cerebral cortex and hippocampal CA1 region increases rapidly following controlled cortical impact in male mice, with an early peak at 1 h, followed by a secondary peak from 24-96 h after TBI. In situ localization using oxidized hydroethidine and the neuronal marker, NeuN, revealed that the O(2(- induction occurred in neurons at 1 h after TBI. Pre- or post-treatment with the NADPH oxidase inhibitor, apocynin markedly inhibited microglial activation and oxidative stress damage. Apocynin also attenuated TBI-induction of the Alzheimer's disease proteins β-amyloid and amyloid precursor protein. Finally, both pre- and post-treatment of apocynin was also shown to induce significant neuroprotection against TBI. In addition, a NOX2-specific inhibitor, gp91ds-tat was also shown to exert neuroprotection against TBI. CONCLUSIONS/SIGNIFICANCE: As a whole, the study demonstrates that NADPH oxidase activity and superoxide production exhibit a biphasic elevation in the hippocampus and cortex following TBI, which contributes significantly to the pathology of TBI via mediation of oxidative stress damage, microglial activation, and AD protein induction in the brain following TBI.

Ozone affects pollen viability and NAD(P)H oxidase release from Ambrosia artemisiifolia pollen.

Science.gov (United States)

Pasqualini, Stefania; Tedeschini, Emma; Frenguelli, Giuseppe; Wopfner, Nicole; Ferreira, Fatima; D'Amato, Gennaro; Ederli, Luisa

2011-10-01

Air pollution is frequently proposed as a cause of the increased incidence of allergy in industrialised countries. We investigated the impact of ozone (O(3)) on reactive oxygen species (ROS) and allergen content of ragweed pollen (Ambrosia artemisiifolia). Pollen was exposed to acute O(3) fumigation, with analysis of pollen viability, ROS and nitric oxide (NO) content, activity of nicotinamide adenine dinucleotide phosphate (NAD[P]H) oxidase, and expression of major allergens. There was decreased pollen viability after O(3) fumigation, which indicates damage to the pollen membrane system, although the ROS and NO contents were not changed or were only slightly induced, respectively. Ozone exposure induced a significant enhancement of the ROS-generating enzyme NAD(P)H oxidase. The expression of the allergen Amb a 1 was not affected by O(3), determined from the mRNA levels of the major allergens. We conclude that O(3) can increase ragweed pollen allergenicity through stimulation of ROS-generating NAD(P)H oxidase. Copyright © 2011 Elsevier Ltd. All rights reserved.

Steady-state oxidation of cholesterol catalyzed by cholesterol oxidase in lipid bilayer membranes on platinum electrodes

International Nuclear Information System (INIS)

Bokoch, Michael P.; Devadoss, Anando; Palencsar, Mariela S.; Burgess, James D.

2004-01-01

Cholesterol oxidase is immobilized in electrode-supported lipid bilayer membranes. Platinum electrodes are initially modified with a self-assembled monolayer of thiolipid. A vesicle fusion method is used to deposit an outer leaflet of phospholipids onto the thiolipid monolayer forming a thiolipid/lipid bilayer membrane on the electrode surface. Cholesterol oxidase spontaneously inserts into the electrode-supported lipid bilayer membrane from solution and is consequently immobilized to the electrode surface. Cholesterol partitions into the membrane from buffer solutions containing cyclodextrin. Cholesterol oxidase catalyzes the oxidation of cholesterol by molecular oxygen, forming hydrogen peroxide as a product. Amperometric detection of hydrogen peroxide for continuous solution flow experiments are presented, where flow was alternated between cholesterol solution and buffer containing no cholesterol. Steady-state anodic currents were observed during exposures of cholesterol solutions ranging in concentration from 10 to 1000 μM. These data are consistent with the Michaelis-Menten kinetic model for oxidation of cholesterol as catalyzed by cholesterol oxidase immobilized in the lipid bilayer membrane. The cholesterol detection limit is below 1 μM for cholesterol solution prepared in buffered cyclodextrin. The response of the electrodes to low density lipoprotein solutions is increased upon addition of cyclodextrin. Evidence for adsorption of low density lipoprotein to the electrode surface is presented

Pharmacological inhibition of NADPH oxidase protects against cisplatin induced nephrotoxicity in mice by two step mechanism.

Science.gov (United States)

Wang, Yimin; Luo, Xiao; Pan, Hao; Huang, Wei; Wang, Xueping; Wen, Huali; Shen, Kezhen; Jin, Baiye

2015-09-01

Cisplatin induced nephrotoxicity is primarily caused by ROS (Reactive Oxygen Species) induced proximal tubular cell death. NADPH oxidase is major source of ROS production by cisplatin. Here, we reported that pharmacological inhibition of NADPH oxidase by acetovanillone (obtained from medicinal herb Picrorhiza kurroa) led to reduced cisplatin nephrotoxicity in mice. In this study we used various molecular biology and biochemistry methods a clinically relevant model of nephropathy, induced by an important chemotherapeutic drug cisplatin. Cisplatin-induced nephrotoxicity was evident by histological damage from loss of the tubular structure. The damage was also marked by the increase in blood urea nitrogen, creatinine, protein nitration as well as cell death markers such as caspase 3/7 activity and DNA fragmentation. Tubular cell death by cisplatin led to pro-inflammatory response by production of TNFα and IL1β followed by leukocyte/neutrophil infiltration which resulted in new wave of ROS involving more NADPH oxidases. Cisplatin-induced markers of kidney damage such as oxidative stress, cell death, inflammatory cytokine production and nephrotoxicity were attenuated by acetovanillone. In addition to that, acetovanillone enhanced cancer cell killing efficacy of cisplatin. Thus, pharmacological inhibition of NADPH oxidase can be protective for cisplatin-induced nephrotoxicity in mice. Copyright © 2015. Published by Elsevier Ltd.

Diamine oxidase determination in serum - Low assay reproducibility and misclassification of healthy subjects

DEFF Research Database (Denmark)

Schnoor, Heidi Julius; Mosbech, Holger Fausbøll; Skov, Per Stahl

2013-01-01

Summary Background: Impaired histamine degradation based on reduced diamine oxidase (DAO) activity has been suggested to cause symptoms mimicking an allergic reaction. Aim: To test whether patients presenting with possible histamine-induced symptoms have a low serum activity of DAO compared...... in the test kit was 86 %. Only 11 out of the 31 subjects were uniformly classified in all three runs. Among the healthy subjects, 9–12 out of 18 showed reduced or highly reduced activities; in the patient group, 0–5 out of 11 showed reduced or highly reduced activities in the three measurements...... normal range caused misclassification in more than half of the cases. Using a commercial immunoassay, it was not possible to distinguish healthy subjects from patients showing potentially histamine-induced symptoms....

Evaluation of Antimicrobial Activity of Glucose Oxidase from Aspergillus niger EBL-A and Penicillium notatum

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Muhammad Anjum Zia

2013-12-01

Full Text Available This work aimed to study the production and purification of glucose oxidase by Aspergillus niger and Penicillium notatum using corn steep liquor as the substrate and evaluate its antimicrobial activity for use in pharmaceutical and food industries. The enzyme was purified by ammonium sulfate precipitation (60-85%, DEAE-cellulose ion exchange and Sephadex G-200 size exclusion chromatography. The crude enzyme extracts of A. niger and P. notatum showed 2.32 and 5.53 U mg-1 specific activities, respectively, which after desalting was 15.52 and 12.05 U mg-1, and after ion exchange and gel filtration chromatography was 29.09 - 62 and 25.72 - 59.37 U mg-1 for A. niger and P. notatum, respectively. The antimicrobial activity was determined by disc diffusion method against selected microbial strains where glucose oxidase from A. niger showed anti-bacterial activity, while no fungicidal effects were shown by both A. niger and P. notatum glucose oxidases.

Priming of the neutrophil NADPH oxidase activation: role of p47phox phosphorylation and NOX2 mobilization to the plasma membrane.

Science.gov (United States)

El-Benna, Jamel; Dang, Pham My-Chan; Gougerot-Pocidalo, Marie-Anne

2008-07-01

Neutrophils play an essential role in host defense against microbial pathogens and in the inflammatory reaction. Upon activation, neutrophils produce superoxide anion (O*2), which generates other reactive oxygen species (ROS) such as hydrogen peroxide (H2O2), hydroxyl radical (OH*) and hypochlorous acid (HOCl), together with microbicidal peptides and proteases. The enzyme responsible for O2* production is called the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase or respiratory burst oxidase. This multicomponent enzyme system is composed of two trans-membrane proteins (p22phox and gp91phox/NOX2, which form the cytochrome b558), three cytosolic proteins (p47phox, p67phox, p40phox) and a GTPase (Rac1 or Rac2), which assemble at membrane sites upon cell activation. NADPH oxidase activation in phagocytes can be induced by a large number of soluble and particulate factors. Three major events accompany NAPDH oxidase activation: (1) protein phosphorylation, (2) GTPase activation, and (3) translocation of cytosolic components to the plasma membrane to form the active enzyme. Actually, the neutrophil NADPH oxidase exists in different states: resting, primed, activated, or inactivated. The resting state is found in circulating blood neutrophils. The primed state can be induced by neutrophil adhesion, pro-inflammatory cytokines, lipopolysaccharide, and other agents and has been characterized as a "ready to go" state, which results in a faster and higher response upon exposure to a second stimulus. The active state is found at the inflammatory or infection site. Activation is induced by the pathogen itself or by pathogen-derived formylated peptides and other agents. Finally, inactivation of NADPH oxidase is induced by anti-inflammatory agents to limit inflammation. Priming is a "double-edged sword" process as it contributes to a rapid and efficient elimination of the pathogens but can also induce the generation of large quantities of toxic ROS by hyperactivation of

Cloning, expression, purification, crystallization and preliminary X-ray diffraction analysis of variants of monoamine oxidase from Aspergillus niger

Energy Technology Data Exchange (ETDEWEB)

Atkin, Kate E. [Structural Biology Laboratory, Department of Chemistry, University of York, York YO10 5YW (United Kingdom); Reiss, Renate; Turner, Nicholas J. [School of Chemistry, Manchester Interdisciplinary Biocentre, University of Manchester, 131 Princess Street, Manchester M1 7DN (United Kingdom); Brzozowski, Andrzej M.; Grogan, Gideon, E-mail: grogan@ysbl.york.ac.uk [Structural Biology Laboratory, Department of Chemistry, University of York, York YO10 5YW (United Kingdom)

2008-03-01

Crystals of A. niger monoamine oxidase variants display P2{sub 1} or P4{sub 1}2{sub 1}2/P4{sub 3}2{sub 1}2 symmetry, with eight or two molecules in the asymmetric unit, respectively. Monoamine oxidase from Aspergillus niger (MAO-N) is an FAD-dependent enzyme that catalyses the conversion of terminal amines to their corresponding aldehydes. Variants of MAO-N produced by directed evolution have been shown to possess altered substrate specificity. Crystals of two of these variants (MAO-N-3 and MAO-N-5) have been obtained; the former displays P2{sub 1} symmetry with eight molecules per asymmetric unit and the latter has P4{sub 1}2{sub 1}2 or P4{sub 3}2{sub 1}2 symmetry and two molecules per asymmetric unit. Solution of these structures will help shed light on the molecular determinants of improved activity and high enantioselectivity towards a broad range of substrates.

Structural characterization of acyl-CoA oxidases reveals a direct link between pheromone biosynthesis and metabolic state in Caenorhabditis elegans.

Science.gov (United States)

Zhang, Xinxing; Li, Kunhua; Jones, Rachel A; Bruner, Steven D; Butcher, Rebecca A

2016-09-06

Caenorhabditis elegans secretes ascarosides as pheromones to communicate with other worms and to coordinate the development and behavior of the population. Peroxisomal β-oxidation cycles shorten the side chains of ascaroside precursors to produce the short-chain ascaroside pheromones. Acyl-CoA oxidases, which catalyze the first step in these β-oxidation cycles, have different side chain-length specificities and enable C. elegans to regulate the production of specific ascaroside pheromones. Here, we determine the crystal structure of the acyl-CoA oxidase 1 (ACOX-1) homodimer and the ACOX-2 homodimer bound to its substrate. Our results provide a molecular basis for the substrate specificities of the acyl-CoA oxidases and reveal why some of these enzymes have a very broad substrate range, whereas others are quite specific. Our results also enable predictions to be made for the roles of uncharacterized acyl-CoA oxidases in C. elegans and in other nematode species. Remarkably, we show that most of the C. elegans acyl-CoA oxidases that participate in ascaroside biosynthesis contain a conserved ATP-binding pocket that lies at the dimer interface, and we identify key residues in this binding pocket. ATP binding induces a structural change that is associated with tighter binding of the FAD cofactor. Mutations that disrupt ATP binding reduce FAD binding and reduce enzyme activity. Thus, ATP may serve as a regulator of acyl-CoA oxidase activity, thereby directly linking ascaroside biosynthesis to ATP concentration and metabolic state.

Kinetic isotope effect studies on milk xanthine oxidase and on chicken liver xanthine dehydrogenase

International Nuclear Information System (INIS)

D'Ardenne, S.C.; Edmondson, D.E.

1990-01-01

The effect of isotopic substitution of the 8-H of xanthine (with 2 H and 3 H) on the rate of oxidation by bovine xanthine oxidase and by chicken xanthine dehydrogenase has been measured. V/K isotope effects were determined from competition experiments. No difference in H/T (V/K) values was observed between xanthine oxidase and xanthine dehydrogenase. Xanthine dehydrogenase exhibited a larger T/D (V/K) value than that observed for xanthine oxidase. Observed H/T (V/K) values for either enzyme are less than those H/T (V/K) values calculated with D/T (V/K) data. These discrepancies are suggested to arise from the presence of a rate-limiting step(s) prior to the irreversible C-H bond cleavage step in the mechanistic pathways of both enzymes. These kinetic complexities preclude examination of whether tunneling contributes to the reaction coordinate for the H-transfer step in each enzyme. No observable exchange of tritium with solvent is observed during the anaerobic incubation of [8- 3 H]xanthine with either enzyme, which suggests the reverse commitment to catalysis (C r ) is essentially zero. With the assumption of adherence to reduced mass relationships, the intrinsic deuterium isotope effect ( D k) for xanthine oxidation is calculated. By the use of these values and steady-state kinetic data, the minimal rate for the hydrogen-transfer step is calculated to be ∼75-fold faster than k cat for xanthine oxidase and ∼10-fold faster than k cat for xanthine dehydrogenase. Values calculated for each enzyme were found to be identical within experimental uncertainty

[THE INFLUENCE OF SEROTONIN TRANSPORTER AND MONOAMINE OXIDASE A GENES POLYMORPHISM ON PSYCHO-EMOTION AND KARYOLOGICAL STABILITY OF ATHLETES].

Science.gov (United States)

Kalaev, V N; Nechaeva, M S; Korneeva, O S; Cherenkov, D A

2015-11-01

The influence of polymorphism of the serotonin transporter and monoamine oxidase A genes, associated with man's aggressiveness on the psycho-emotional state and karyological status of single combat athletes. It was revealed that the carriers of less active ("short"), monoamine oxidase A gene variant have a high motivation to succeed and less rigidity and frustrated, compared to the carriers of more active ("long") version of the gene. Heterozygote carriers of less active ("short") variant of the serotonin transporter gene 5-HTTL had more physical aggression, guilt and were less frustrated compared with carriers of two long alleles. It has been revealed the association of studied genes with the karyological status of athletes. So fighters who are carriers of the short and long alleles of the serotonin transporter gene had more cells with nuclear abnormalities in the buccal epithelium than single combat athletes which both alleles were long.

Novel ferrocene-anchored ZnO nanoparticle/carbon nanotube assembly for glucose oxidase wiring: application to a glucose/air fuel cell.

Science.gov (United States)

Haddad, Raoudha; Mattei, Jean-Gabriel; Thery, Jessica; Auger, Aurélien

2015-06-28

Glucose oxidase (GOx) is immobilized on ZnO nanoparticle-modified electrodes. The immobilized glucose oxidase shows efficient mediated electron transfer with ZnO nanoparticles to which the ferrocenyl moiety is π-stacked into a supramolecular architecture. The constructed ZnO-Fc/CNT modified electrode exhibits high ferrocene surface coverage, preventing any leakage of the π-stacked ferrocene from the newly described ZnO hybrid nanoparticles. The use of the new architecture of ZnO supported electron mediators to shuttle electrons from the redox centre of the enzyme to the surface of the working electrode can effectively bring about successful glucose oxidation. These modified electrodes evaluated as a highly efficient architecture provide a catalytic current for glucose oxidation and are integrated in a specially designed glucose/air fuel cell prototype using a conventional platinum-carbon (Pt/C) cathode at physiological pH (7.0). The obtained architecture leads to a peak power density of 53 μW cm(-2) at 300 mV for the Nafion® based biofuel cell under "air breathing" conditions at room temperature.

Fatal Chromobacterium violaceum septicaemia in northern Laos, a modified oxidase test and post-mortem forensic family G6PD analysis.

Science.gov (United States)

Slesak, Günther; Douangdala, Phouvieng; Inthalad, Saythong; Silisouk, Joy; Vongsouvath, Manivanh; Sengduangphachanh, Amphonesavanh; Moore, Catrin E; Mayxay, Mayfong; Matsuoka, Hiroyuki; Newton, Paul N

2009-07-29

Chromobacterium violaceum is a Gram negative facultative anaerobic bacillus, found in soil and stagnant water, that usually has a violet pigmented appearance on agar culture. It is rarely described as a human pathogen, mostly from tropical and subtropical areas. A 53 year-old farmer died with Chromobacterium violaceum septicemia in Laos. A modified oxidase method was used to demonstrate that this violacious organism was oxidase positive. Forensic analysis of the glucose-6-phosphate dehydrogenase genotypes of his family suggest that the deceased patient did not have this possible predisposing condition. C. violaceum infection should be included in the differential diagnosis in patients presenting with community-acquired septicaemia in tropical and subtropical areas. The apparently neglected but simple modified oxidase test may be useful in the oxidase assessment of other violet-pigmented organisms or of those growing on violet coloured agar.

Direction of the reactivity of vanillyl-alcohol oxidase with 4-alkylphenols

NARCIS (Netherlands)

van den Heuvel, RHH; Fraaije, MW; van Berkel, WJH; Heuvel, Robert H.H. van den; Berkel, Willem J.H. van

2000-01-01

The covalent flavoprotein vanillyl-alcohol oxidase (VAO) predominantly converts short-chain 4-alkylphenols, like 4-ethylphenol, to (R)-1-(4'-hydroxyphenyl)alcohols and medium-chain 4-alkylphenols, like 4-butylphenol, to 1-(4'-hydroxyphenyl)alkenes. Crystallographic studies have indicated that the

New insights into the roles of NADPH oxidases in sexual development and ascospore germination in Sordaria macrospora.

Science.gov (United States)

Dirschnabel, Daniela Elisabeth; Nowrousian, Minou; Cano-Domínguez, Nallely; Aguirre, Jesus; Teichert, Ines; Kück, Ulrich

2014-03-01

NADPH oxidase (NOX)-derived reactive oxygen species (ROS) act as signaling determinants that induce different cellular processes. To characterize NOX function during fungal development, we utilized the genetically tractable ascomycete Sordaria macrospora. Genome sequencing of a sterile mutant led us to identify the NADPH oxidase encoding nox1 as a gene required for fruiting body formation, regular hyphal growth, and hyphal fusion. These phenotypes are shared by nor1, lacking the NOX regulator NOR1. Further phenotypic analyses revealed a high correlation between increased ROS production and hyphal fusion deficiencies in nox1 and other sterile mutants. A genome-wide transcriptional profiling analysis of mycelia and isolated protoperithecia from wild type and nox1 revealed that nox1 inactivation affects the expression of genes related to cytoskeleton remodeling, hyphal fusion, metabolism, and mitochondrial respiration. Genetic analysis of nox2, lacking the NADPH oxidase 2 gene, nor1, and transcription factor deletion mutant ste12, revealed a strict melanin-dependent ascospore germination defect, indicating a common genetic pathway for these three genes. We report that gsa3, encoding a G-protein α-subunit, and sac1, encoding cAMP-generating adenylate cyclase, act in a separate pathway during the germination process. The finding that cAMP inhibits ascospore germination in a melanin-dependent manner supports a model in which cAMP inhibits NOX2 activity, thus suggesting a link between both pathways. Our results expand the current knowledge on the role of NOX enzymes in fungal development and provide a frame to define upstream and downstream components of the NOX signaling pathways in fungi.

Effects of temperature and glycerol and methanol-feeding profiles on the production of recombinant galactose oxidase in Pichia pastoris

Science.gov (United States)

Anasontzis, George E; Salazar Penã, Margarita; Spadiut, Oliver; Brumer, Harry; Olsson, Lisbeth

2014-01-01

Optimization of protein production from methanol-induced Pichia pastoris cultures is necessary to ensure high productivity rates and high yields of recombinant proteins. We investigated the effects of temperature and different linear or exponential methanol-feeding rates on the production of recombinant Fusarium graminearum galactose oxidase (EC 1.1.3.9) in a P. pastoris Mut+ strain, under regulation of the AOX1 promoter. We found that low exponential methanol feeding led to 1.5-fold higher volumetric productivity compared to high exponential feeding rates. The duration of glycerol feeding did not affect the subsequent product yield, but longer glycerol feeding led to higher initial biomass concentration, which would reduce the oxygen demand and generate less heat during induction. A linear and a low exponential feeding profile led to productivities in the same range, but the latter was characterized by intense fluctuations in the titers of galactose oxidase and total protein. An exponential feeding profile that has been adapted to the apparent biomass concentration results in more stable cultures, but the concentration of recombinant protein is in the same range as when constant methanol feeding is employed. © 2014 The Authors Biotechnology Progress published by Wiley Periodicals, Inc. on behalf of American Institute of Chemical Engineers Biotechnol. Prog., 30:728–735, 2014 PMID:24493559

Shared features of S100B immunohistochemistry and cytochrome oxidase histochemistry in the ventroposterior thalamus and lateral habenula in neonatal rats.

Science.gov (United States)

Muneoka, Katsumasa; Funahashi, Hisayuki; Ogawa, Tetsuo; Whitaker-Azmitia, Patricia M; Shioda, Seiji

2012-10-01

The ventroposterior thalamus and the habenular nuclei of the epithalamus are relevant to the monoaminergic system functionally and anatomically. The glia-derived S100B protein plays a critical role in the development of the nervous system including the monoaminergic systems. In this study, we performed an immunohistochemical study of glia-related proteins including S100B, serotonin transporter, and microtubule-associated protein 2, as well as cytochrome oxidase histochemistry in neonatal rats. Results showed the same findings for S100B immunohistochemistry between the ventroposterior thalamus and the lateral habenula at postnatal day 7: intense staining in cell bodies of astrocytes, diffusely spread immunoproduct in the intercellular space, and S100B-free areas as well as a strong reaction to cytochrome oxidase histochemistry. Further common features were the scarcity of glial fibrillary acidic protein-positive astrocytes and the few apoptotic cells observed. The results of the cytochrome oxidase reaction suggested that S100B is released actively into intercellular areas in restricted brain regions showing high neuronal activity at postnatal day 7. Pathology of the ventroposterior thalamus and the habenula is suggested in mental disorders, and S100B might be a key factor for investigations in these areas. Copyright © 2012 ISDN. Published by Elsevier Ltd. All rights reserved.

Reconstitution of apoglucose oxidase with FAD conjugates for biosensoring of progesterone

NARCIS (Netherlands)

Posthuma-Trumpie, G.A.; Berg, van den W.A.M.; Wiel, van de D.F.M.; Schaaper, W.M.M.; Korf, J.; Berkel, van W.J.H.

2007-01-01

The reconstitution of Aspergillus niger apoglucose oxidase (apoGOx) with FAD conjugates for biosensoring of progesterone was investigated. ApoGOx prepared by partial unfolding of the protein under acidic conditions consisted of reconstitutable monomers (50 ± 10%), reconstitutable dimers (20 ± 10%)

Export of functional Streptomyces coelicolor alditol oxidase to the periplasm or cell surface of Escherichia coli and its application in whole-cell biocatalysis

NARCIS (Netherlands)

van Bloois, Edwin; Winter, Remko T.; Janssen, Dick B.; Fraaije, Marco W.

Streptomyces coelicolor A3(2) alditol oxidase (AldO) is a soluble monomeric flavoprotein in which the flavin cofactor is covalently linked to the polypeptide chain. AldO displays high reactivity towards different polyols such as xylitol and sorbitol. These characteristics make AldO industrially

Localization of Glucose Oxidase and Catalase Activities in Aspergillus niger

NARCIS (Netherlands)

Witteveen, Cor F.B.; Veenhuis, Marten; Visser, Jaap

The subcellular localization of glucose oxidase (EC 1.1.3.4) in Aspergillus niger N400 (CBS 120.49) was investigated by (immuno)cytochemical methods. By these methods, the bulk of the enzyme was found to be localized in the cell wall. In addition, four different catalases (EC 1.11.1.6) were

Mechanism-Based Inhibitors of Cytokinin Oxidase/Dehydrogenase Attack FAD Cofactor

Czech Academy of Sciences Publication Activity Database

Kopečný, D.; Šebela, M.; Briozzo, P.; Spíchal, Lukáš; Houba-Hérin, N.; Mašek, V.; Joly, N.; Madzak, C.; Anzenbacher, P.; Laloue, M.

2008-01-01

Roč. 380, č. 5 (2008), s. 886-899 ISSN 0022-2836 R&D Projects: GA ČR(CZ) GP522/08/P113 Institutional research plan: CEZ:AV0Z50380511 Keywords : cytokinin oxidase/dehydrogenase * cytokinin signaling * protein structure Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.146, year: 2008

Cytochrome oxidase subunit II gene in mitochondria of Oenothera has no intron

Science.gov (United States)

Hiesel, Rudolf; Brennicke, Axel

1983-01-01

The cytochrome oxidase subunit II gene has been localized in the mitochondrial genome of Oenothera berteriana and the nucleotide sequence has been determined. The coding sequence contains 777 bp and, unlike the corresponding gene in Zea mays, is not interrupted by an intron. No TGA codon is found within the open reading frame. The codon CGG, as in the maize gene, is used in place of tryptophan codons of corresponding genes in other organisms. At position 742 in the Oenothera sequence the TGG of maize is changed into a CGG codon, where Trp is conserved as the amino acid in other organisms. Homologous sequences occur more than once in the mitochondrial genome as several mitochondrial DNA species hybridize with DNA probes of the cytochrome oxidase subunit II gene. ImagesFig. 5. PMID:16453484

Fatal Chromobacterium violaceum septicaemia in northern Laos, a modified oxidase test and post-mortem forensic family G6PD analysis

Directory of Open Access Journals (Sweden)

Mayxay Mayfong

2009-07-01

Full Text Available Abstract Background Chromobacterium violaceum is a Gram negative facultative anaerobic bacillus, found in soil and stagnant water, that usually has a violet pigmented appearance on agar culture. It is rarely described as a human pathogen, mostly from tropical and subtropical areas. Case presentation A 53 year-old farmer died with Chromobacterium violaceum septicemia in Laos. A modified oxidase method was used to demonstrate that this violacious organism was oxidase positive. Forensic analysis of the glucose-6-phosphate dehydrogenase genotypes of his family suggest that the deceased patient did not have this possible predisposing condition. Conclusion C. violaceum infection should be included in the differential diagnosis in patients presenting with community-acquired septicaemia in tropical and subtropical areas. The apparently neglected but simple modified oxidase test may be useful in the oxidase assessment of other violet-pigmented organisms or of those growing on violet coloured agar.

Structural characterization of acyl-CoA oxidases reveals a direct link between pheromone biosynthesis and metabolic state in Caenorhabditis elegans

Science.gov (United States)

Zhang, Xinxing; Jones, Rachel A.; Bruner, Steven D.; Butcher, Rebecca A.

2016-01-01

Caenorhabditis elegans secretes ascarosides as pheromones to communicate with other worms and to coordinate the development and behavior of the population. Peroxisomal β-oxidation cycles shorten the side chains of ascaroside precursors to produce the short-chain ascaroside pheromones. Acyl-CoA oxidases, which catalyze the first step in these β-oxidation cycles, have different side chain-length specificities and enable C. elegans to regulate the production of specific ascaroside pheromones. Here, we determine the crystal structure of the acyl-CoA oxidase 1 (ACOX-1) homodimer and the ACOX-2 homodimer bound to its substrate. Our results provide a molecular basis for the substrate specificities of the acyl-CoA oxidases and reveal why some of these enzymes have a very broad substrate range, whereas others are quite specific. Our results also enable predictions to be made for the roles of uncharacterized acyl-CoA oxidases in C. elegans and in other nematode species. Remarkably, we show that most of the C. elegans acyl-CoA oxidases that participate in ascaroside biosynthesis contain a conserved ATP-binding pocket that lies at the dimer interface, and we identify key residues in this binding pocket. ATP binding induces a structural change that is associated with tighter binding of the FAD cofactor. Mutations that disrupt ATP binding reduce FAD binding and reduce enzyme activity. Thus, ATP may serve as a regulator of acyl-CoA oxidase activity, thereby directly linking ascaroside biosynthesis to ATP concentration and metabolic state. PMID:27551084

Graphene-glucose oxidase bioanodes for enzymatic biofuel cells

DEFF Research Database (Denmark)

Tang, Jing; Werchmeister, Rebecka Maria Larsen; Engelbrekt, Christian

2017-01-01

as supporting material, polyethyleneimine (PEI) as linker and glucose oxidase (GOD) as the chosen enzyme. GOD can catalyze oxidation of glucose to gluconolactone, but needs a mediator to assist electron transfer between the enzyme and electrodes. The redox molecule ferrocene carboxylic acid (Fc...... and systematically investigated. The assembled EBFCs show good reproducibility. EBFCs provide maximum output power density 2.47 μW cm-2 at 35 ℃, indicating the optimized activity of EBFCs fed with glucose....

Auxin-activated NADH oxidase activity of soybean plasma membranes is distinct from the constitutive plasma membrane NADH oxidase and exhibits prion-like properties

Science.gov (United States)

Morre, D. James; Morre, Dorothy M.; Ternes, Philipp

2003-01-01

The hormone-stimulated and growth-related cell surface hydroquinone (NADH) oxidase activity of etiolated hypocotyls of soybeans oscillates with a period of about 24 min or 60 times per 24-h day. Plasma membranes of soybean hypocotyls contain two such NADH oxidase activities that have been resolved by purification on concanavalin A columns. One in the apparent molecular weight range of 14-17 kDa is stimulated by the auxin herbicide 2,4-dichlorophenoxyacetic acid (2,4-D). The other is larger and unaffected by 2,4-D. The 2,4-D-stimulated activity absolutely requires 2,4-D for activity and exhibits a period length of about 24 min. Also exhibiting 24-min oscillations is the rate of cell enlargement induced by the addition of 2,4-D or the natural auxin indole-3-acetic acid (IAA). Immediately following 2,4-D or IAA addition, a very complex pattern of oscillations is frequently observed. However, after several hours a dominant 24-min period emerges at the expense of the constitutive activity. A recruitment process analogous to that exhibited by prions is postulated to explain this behavior.

Enzymatic halogenation and oxidation using an alcohol oxidase-vanadium chloroperoxidase cascade

NARCIS (Netherlands)

But, Andrada; Noord, Van Aster; Poletto, Francesca; Sanders, Johan P.M.; Franssen, Maurice C.R.; Scott, Elinor L.

2017-01-01

The chemo-enzymatic cascade which combines alcohol oxidase from Hansenula polymorpha (AOXHp) with vanadium chloroperoxidase (VCPO), for the production of biobased nitriles from amino acids was investigated. In the first reaction H2O2 (and acetaldehyde) are generated from ethanol and oxygen by AOXHp.

Depletion of rat cortical norepinephrine and the inhibition of [3H]norepinephrine uptake by xylamine does not require monoamine oxidase activity

International Nuclear Information System (INIS)

Dudley, M.W.

1988-01-01

Inhibition of monoamine oxidase A through pretreatment of rats with clorgyline or the pro-drug MDL 72,394 did not block the amine-depleting action of xylamine. Xylamine treatment resulted in a loss of approximately 60% of the control level of norepinephrine in the cerebral cortex. A 1-hr pretreatment, but not a 24-hr pretreatment, with the monoamine oxidase B inhibitor, L-deprenyl, prevented the depletion of norepinephrine by xylamine. In addition, pretreatment with MDL 72,974, a monoamine oxidase B inhibitor without amine-releasing or uptake - inhibiting effects, did not prevent cortical norepinephrine levels. Inhibition of monoamine oxidase by either MDL 72,974 or MDL 72,394 did not prevent the inhibition of [ 3 H]norepinephrine uptake into rat cortical synaptosomes by xylamine. These data indicate that monoamine oxidase does not mediate the amine-releasing or uptake inhibiting properties of xylamine. The protection afforded by L-deprenyl following a 1-hr pretreatment most probably was due to accumulation of its metabolite, L-amphetamine, which would inhibit the uptake carrier. A functional carrier is required for depletion since desipramine administered 1 hr prior to xylamine, was also able to prevent depletion of norepinephrine

COI (cytochrome oxidase-I) sequence based studies of Carangid fishes from Kakinada coast, India.

Science.gov (United States)

Persis, M; Chandra Sekhar Reddy, A; Rao, L M; Khedkar, G D; Ravinder, K; Nasruddin, K

2009-09-01

Mitochondrial DNA, cytochrome oxidase-1 gene sequences were analyzed for species identification and phylogenetic relationship among the very high food value and commercially important Indian carangid fish species. Sequence analysis of COI gene very clearly indicated that all the 28 fish species fell into five distinct groups, which are genetically distant from each other and exhibited identical phylogenetic reservation. All the COI gene sequences from 28 fishes provide sufficient phylogenetic information and evolutionary relationship to distinguish the carangid species unambiguously. This study proves the utility of mtDNA COI gene sequence based approach in identifying fish species at a faster pace.

Iododerivative of pargyline: A potential tracer for the exploration of monoamine oxidase sites by SPECT

International Nuclear Information System (INIS)

Lena, Isabelle; Ombetta, Jean-Edouard; Chalon, Sylvie; Dognon, Anne-Marie; Baulieu, Jean-Louis; Frangin, Yves; Garreau, Lucette; Besnard, Jean-Claude; Guilloteau, Denis

1995-01-01

Monoamine oxidases are important in the regulation of monoaminergic neurotransmission. An increase in monoamine oxidase B (MAO B) has been observed in some neurodegenerative diseases, and therefore quantification of cerebral MAO B activity by SPECT would be useful for the diagnosis and therapeutic follow-up of these disorders. We have developed an iodinated derivative of pargyline, a selective inhibitor of MAO B, in order to explore this enzyme by SPECT. Stable bromo and iodo derivatives of pargyline were synthesized and chemically characterized. The radioiodinated ligand [ 125 I]-2-iodopargyline was obtained with high specific activity from the bromo precursor by nucleophilic exchange. Affinity and selectivity of 2-iodopargyline were tested in vitro. Biodistribution study of [ 125 I]-2-iodopargyline was performed in rats. Radioiodinated ligand were obtained in a no-carrier-added form. 2-iodopargyline has a higher in vitro affinity for MAO B than pargyline. However, the in vitro selectivity for MAO B was better for pargyline than for 2-iodopargyline. Ex vivo autoradiographic studies and in vivo saturation studies with selective inhibitors of MAO showed that the cerebral biodistribution of [ 125 I]-2-iodopargyline in the rat is consistent with high level binding to MAO B sites in the pineal gland and in the thalamus. In conclusion, 2-iodopargyline preferentially binds in vivo to MAO B sites with high affinity. However, its selectivity for MAO B in rats is not very high, whereas this ligand binds to a lesser extent to MAO A. It will be then of great value to evaluate the specificity of 2-iodopargyline in humans. This new ligand labeled with 123 I should therefore be a suitable tool for SPECT exploration of MAO B in the human brain

Biocatalytic Properties and Structural Analysis of Eugenol Oxidase from Rhodococcus jostii RHA1: A Versatile Oxidative Biocatalyst

OpenAIRE

Nguyen, Quoc-Thai; de Gonzalo, Gonzalo; Binda, Claudia; Rioz-Martínez, Ana; Mattevi, Andrea; Fraaije, Marco W.

2016-01-01

Abstract Eugenol oxidase (EUGO) from Rhodococcus jostii RHA1 had previously been shown to convert only a limited set of phenolic compounds. In this study, we have explored the biocatalytic potential of this flavoprotein oxidase, resulting in a broadened substrate scope and a deeper insight into its structural properties. In addition to the oxidation of vanillyl alcohol and the hydroxylation of eugenol, EUGO can efficiently catalyze the dehydrogenation of various phenolic ketones and the selec...

Low-Dose Dextromethorphan, a NADPH Oxidase Inhibitor, Reduces Blood Pressure and Enhances Vascular Protection in Experimental Hypertension

Science.gov (United States)

Wu, Tao-Cheng; Chao, Chih-Yu; Lin, Shing-Jong; Chen, Jaw-Wen

2012-01-01

Background Vascular oxidative stress may be increased with age and aggravate endothelial dysfunction and vascular injury in hypertension. This study aimed to investigate the effects of dextromethorphan (DM), a NADPH oxidase inhibitor, either alone or in combination treatment, on blood pressure (BP) and vascular protection in aged spontaneous hypertensive rats (SHRs). Methodology/Principal Findings Eighteen-week-old WKY rats and SHRs were housed for 2 weeks. SHRs were randomly assigned to one of the 12 groups: untreated; DM monotherapy with 1, 5 or 25 mg/kg/day; amlodipine (AM, a calcium channel blocker) monotherapy with 1 or 5 mg/kg/day; and combination therapy of DM 1, 5 or 25 mg/kg/day with AM 1 or 5 mg/kg/day individually for 4 weeks. The in vitro effects of DM were also examined. In SHRs, AM monotherapy dose-dependently reduced arterial systolic BP. DM in various doses significantly and similarly reduced arterial systolic BP. Combination of DM with AM gave additive effects on BP reduction. DM, either alone or in combination with AM, improved aortic endothelial function indicated by ex vivo acetylcholine-induced relaxation. The combination of low-dose DM with AM gave most significant inhibition on aortic wall thickness in SHRs. Plasma total antioxidant status was significantly increased by all the therapies except for the combination of high-dose DM with high-dose AM. Serum nitrite and nitrate level was significantly reduced by AM but not by DM or the combination of DM with AM. Furthermore, in vitro treatment with DM reduced angiotensin II-induced reactive oxygen species and NADPH oxidase activation in human aortic endothelial cells. Conclusions/Significance Treatment of DM reduced BP and enhanced vascular protection probably by inhibiting vascular NADPH oxidase in aged hypertensive animals with or without AM treatment. It provides the potential rationale to a novel combination treatment with low-dose DM and AM in clinical hypertension. PMID:23049937

Low-dose dextromethorphan, a NADPH oxidase inhibitor, reduces blood pressure and enhances vascular protection in experimental hypertension.

Directory of Open Access Journals (Sweden)

Tao-Cheng Wu

Full Text Available BACKGROUND: Vascular oxidative stress may be increased with age and aggravate endothelial dysfunction and vascular injury in hypertension. This study aimed to investigate the effects of dextromethorphan (DM, a NADPH oxidase inhibitor, either alone or in combination treatment, on blood pressure (BP and vascular protection in aged spontaneous hypertensive rats (SHRs. METHODOLOGY/PRINCIPAL FINDINGS: Eighteen-week-old WKY rats and SHRs were housed for 2 weeks. SHRs were randomly assigned to one of the 12 groups: untreated; DM monotherapy with 1, 5 or 25 mg/kg/day; amlodipine (AM, a calcium channel blocker monotherapy with 1 or 5 mg/kg/day; and combination therapy of DM 1, 5 or 25 mg/kg/day with AM 1 or 5 mg/kg/day individually for 4 weeks. The in vitro effects of DM were also examined. In SHRs, AM monotherapy dose-dependently reduced arterial systolic BP. DM in various doses significantly and similarly reduced arterial systolic BP. Combination of DM with AM gave additive effects on BP reduction. DM, either alone or in combination with AM, improved aortic endothelial function indicated by ex vivo acetylcholine-induced relaxation. The combination of low-dose DM with AM gave most significant inhibition on aortic wall thickness in SHRs. Plasma total antioxidant status was significantly increased by all the therapies except for the combination of high-dose DM with high-dose AM. Serum nitrite and nitrate level was significantly reduced by AM but not by DM or the combination of DM with AM. Furthermore, in vitro treatment with DM reduced angiotensin II-induced reactive oxygen species and NADPH oxidase activation in human aortic endothelial cells. CONCLUSIONS/SIGNIFICANCE: Treatment of DM reduced BP and enhanced vascular protection probably by inhibiting vascular NADPH oxidase in aged hypertensive animals with or without AM treatment. It provides the potential rationale to a novel combination treatment with low-dose DM and AM in clinical hypertension.

Inhibition of xanthine oxidase reduces oxidative stress and improves skeletal muscle function in response to electrically stimulated isometric contractions in aged mice

Science.gov (United States)

Ryan, Michael J.; Jackson, Janna R.; Hao, Yanlei; Leonard, Stephen S.; Alway, Stephen E.

2012-01-01

Oxidative stress is a putative factor responsible for reducing function and increasing apoptotic signaling in skeletal muscle with aging. This study examined the contribution and functional significance of the xanthine oxidase enzyme as a potential source of oxidant production in aged skeletal muscle during repetitive in situ electrically stimulated isometric contractions. Xanthine oxidase activity was inhibited in young adult and aged mice via a subcutaneously placed time release (2.5 mg/day) allopurinol pellet, 7 days prior to the start of in situ electrically stimulated isometric contractions. Gastrocnemius muscles were electrically activated with 20 maximal contractions for three consecutive days. Xanthine oxidase activity was 65% greater in the gastrocnemius muscle of aged mice compared to young mice. Xanthine oxidase activity also increased after in situ electrically stimulated isometric contractions in muscles from both young (33%) and aged (28%) mice, relative to contralateral non-contracted muscles. Allopurinol attenuated the exercise-induced increase in oxidative stress, but it did not affect the elevated basal levels of oxidative stress that was associated with aging. In addition, inhibition of xanthine oxidase activity decreased caspase 3 activity, but it had no effect on other markers of mitochondrial associated apoptosis. Our results show that compared to control conditions, suppression of xanthine oxidase activity by allopurinol reduced xanthine oxidase activity, H2O2 levels, lipid peroxidation and caspase-3 activity, prevented the in situ electrically stimulated isometric contraction-induced loss of glutathione, prevented the increase of catalase and copper-zinc superoxide dismutase activities, and increased maximal isometric force in the plantar flexor muscles of aged mice after repetitive electrically evoked contractions. PMID:21530649

Protoporphyrinogen oxidase: high affinity tetrahydrophthalimide radioligand for the inhibitor/herbicide-binding site in mouse liver mitochondria.

Science.gov (United States)

Birchfield, N B; Casida, J E

1996-01-01

Protoporphyrinogen oxidase (protox), the last common enzyme in heme and chlorophyll biosynthesis, is the target of several classes of herbicides acting as inhibitors in both plants and mammals. N-(4-Chloro-2-fluoro-5-(propargyloxy)phenyl)-3,4,5,6-tetrahydro phthalimide (a potent protox inhibitor referred to as THP) was synthesized as a candidate radioligand ([3H]-THP) by selective catalytic reduction of 3,6-dihydrophthalic anhydride (DHPA) with tritium gas followed by condensation in 45% yield with 4-chloro-2-fluoro-5-(propargyloxy)aniline. Insertion of tritium at the 3 and 6 carbons of DHPA as well as the expected 4 and 5 carbons resulted in high specific activity [3H]THP (92 Ci/mmol). This radioligand undergoes rapid, specific, saturable, and reversible binding to the inhibitor/herbicide binding site of the protox component of cholate-solubilized mouse liver mitochondria with an apparent Kd of 0.41 nM and Bmax of 0.40 pmol/mg of protein. In the standard assay, mouse preparation (150 micrograms of protein) and [3H]THP (0.5 nM) are incubated in 500 microL of phosphate buffer at pH 7.2 for 15 min at 25 degrees C followed by addition of ammonium sulfate and filtration with glass fiber filters. The potencies of five nitrodiphenyl ethers and two other herbicides as inhibitors of [3H]THP binding correlate well with those for inhibition of protox activity (r2 = 0.97, n = 7), thus validating the binding assay as relevant to enzyme inhibition. It is also suitable to determine in vivo block as illustrated by an approximately 50% decrease in [3H]THP binding in liver mitochondria from mice treated ip with oxyfluorfen at 4 mg/kg. This is the first report of a binding assay for protox in mammals. The high affinity and specific activity of [3H]THP facilitate quantitation of protox and therefore research on a sensitive inhibition site for porphyrin biosynthesis.

$^{111m}$Cd- and $^{199m}$Hg-derivatives of blue oxidases

CERN Multimedia

2002-01-01

The rack-induced bonding concept (H.B.Gray & B.G.~Malmstroem, Comments Inorg. Chem, 2, 203, 1983) postulates that the bound metal ion in metalloproteins is forced to adopt a coordination geometry determined by the rigid peptide conformation of the protein. Alternatively, the metal ion could create its own favoured coordination geometry in a soft peptide conformation. In order to decide who is slave or master the changes of coordination and rigidity of metal sites in blue copper proteins due to metal and ligand exchange were studied by $^{111m}$Cd and $^{199m}$Hg $\\gamma$-$\\gamma$-perturbed angular correlation (PAC). To get a better understanding of the so called " Type 1 Copper Site " of the blue oxidases laccase (LAC) and ascorbate oxidase (AO) we concentrated our investigations on the small blue copper proteins azurin and plastocyanin. \\\\ \\\\In azurin~(Az), the metal ligand methionine 121~(M121) was replaced by several amino acids, e.g. asparagine~(N), glutamic acid~(E), via site directed mutagenesis. Di...

Cytokinin oxidase/dehydrogenase genes in barley and wheat. Cloning and heterologous expression

Czech Academy of Sciences Publication Activity Database

Galuszka, P.; Frébortová, Jitka; Werner, T.; Yamada, M.; Strnad, Miroslav; Schmülling, T.; Frébort, I.

2004-01-01

Roč. 271, č. 20 (2004), s. 3990-4002 ISSN 0014-2956 Institutional research plan: CEZ:AV0Z5038910 Keywords : cereals * cloning * cytokinin oxidase/dehydrogenase Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.260, year: 2004

Effects of phenylated compounds of methylglyoxal bis(guanylhydrazone) on diamine oxidase activity from rat small intestine.

Science.gov (United States)

Balaña-Fouce, R; Pulido, T G; Escudero, D O; Sanz-Sanchez, F

1986-01-01

Two phenylated compounds of methylglyoxal bis(guanylhydrazone), potentially inhibitors of diamine oxidase activity, have been synthesized: phenylglyoxal bis(guanylhydrazone) and diphenylglyoxal bis(guanylhydrazone). Their inhibitory capacity was tested: while PGBG was able to reduce the enzyme activity by 50% at 1.3 microM, DPGBG was only able to reduce diamine oxidase activity by less than 2% at a concentration 1000-fold higher. The inhibition of PGBG was non-competitive and the Ki calculated by a Dixon plot was estimated as 1.7 microM.

In Vivo Metabolic Trapping Radiotracers for Imaging Monoamine Oxidase-A and –B Enzymatic Activity

Science.gov (United States)

Brooks, Allen F.; Shao, Xia; Quesada, Carole A.; Sherman, Phillip; Scott, Peter J.H.; Kilbourn, Michael R.

2017-01-01

The isozymes of monoamine oxidase (MAO-A and MAO-B) are important enzymes involved in the metabolism of numerous biogenic amines, including the neurotransmitters serotonin, dopamine and norepinephrine. Recently, changes in concentrations of MAO-B have been proposed as an in vivo marker of neuroinflammation associated with Alzheimer’s disease. Previous developments of in vivo radiotracers for imaging changes in MAO enzyme expression or activity have utilized the irreversible propargylamine-based suicide inhibitors, or high-affinity reversibly-binding inhibitors. As an alternative approach, we have investigated 1-[11C]methyl-4-aryloxy-1,2,3,6-tetrahydropyridines as metabolic trapping agents for the monoamine oxidases. MAO-mediated oxidation and spontaneous hydrolysis yields 1-[11C]methyl-2,3-dihydro-4-pyridinone as a hydrophilic metabolite that is trapped within brain tissues. Radiotracers with phenyl, biphenyl and 7-coumarinyl ethers were evaluated using microPET imaging in rat and primate brain. No isozyme selectivity for radiotracer trapping was observed in the rat brain for any compound, but in the monkey brain the phenyl ether demonstrated MAO-A selectivity, and the coumarinyl ether showed MAO-B selectivity. These are lead compounds for further development of 1-[11C]methyl-4-aryloxy-1,2,3,6-tetrahydropyridines with optimized brain pharmacokinetics and isozyme selectivity. PMID:26393369

Pretreatment with mixed-function oxidase inducers increases the sensitivity of the hepatocyte/DNA repair assay

International Nuclear Information System (INIS)

Shaddock, J.G.; Heflich, R.H.; McMillan, D.C.; Hinson, J.A.; Casciano, D.A.

1989-01-01

A recent National Toxicology Program evaluation indicates that the rat hepatocyte/DNA repair assay has a high false-negative rate and that it is insensitive to some genotoxic hepatocarcinogens as well as other species and organ-specific carcinogens. In this study, the authors examined whether the sensitivity of the hepatocyte/DNA repair assay might be increased through animal pretreatment with various hepatic mixed-function oxidase inducers, i.e., Aroclor 1254, phenobarbital, and 3,3',4,4'-tetrachloroazobenzene (TCAB). The effects on unscheduled DNA synthesis (UDS), a measured of DNA damage and repair, were studied in cultures exposed to known and/or potential carcinogens that had been evaluated as negative or questionable or that produced conflicting results with hepatocytes isolated from uninduced animals. 4,4'-Oxydianiline, 1-nitropy-rene, and TCAB produced concentration-dependent increases in UDS in hepatocytes from rats pretreated with Aroclor 1254. 4,4'-Oxydianiline and TCAB also induced a dose-dependent increase in DNA repair in hepatocytes from rats pretreated with phenobarbital, whereas 1-nitropyrene was negative. These data indicate that the limited sensitivity to chemical carcinogens displayed by the hepatocyte/DNA repair assay may be increased by using hepatocytes isolated from animals exposed to hepatic mixed-function oxidase inducers

Pretreatment with mixed-function oxidase inducers increases the sensitivity of the hepatocyte/DNA repair assay

Energy Technology Data Exchange (ETDEWEB)

Shaddock, J.G.; Heflich, R.H.; McMillan, D.C.; Hinson, J.A.; Casciano, D.A. (National Center for Toxicological Research, Jefferson, AK (USA) Univ. of Arkansas for Medical Sciences, Little Rock (USA))

1989-01-01

A recent National Toxicology Program evaluation indicates that the rat hepatocyte/DNA repair assay has a high false-negative rate and that it is insensitive to some genotoxic hepatocarcinogens as well as other species and organ-specific carcinogens. In this study, the authors examined whether the sensitivity of the hepatocyte/DNA repair assay might be increased through animal pretreatment with various hepatic mixed-function oxidase inducers, i.e., Aroclor 1254, phenobarbital, and 3,3{prime},4,4{prime}-tetrachloroazobenzene (TCAB). The effects on unscheduled DNA synthesis (UDS), a measured of DNA damage and repair, were studied in cultures exposed to known and/or potential carcinogens that had been evaluated as negative or questionable or that produced conflicting results with hepatocytes isolated from uninduced animals. 4,4{prime}-Oxydianiline, 1-nitropy-rene, and TCAB produced concentration-dependent increases in UDS in hepatocytes from rats pretreated with Aroclor 1254. 4,4{prime}-Oxydianiline and TCAB also induced a dose-dependent increase in DNA repair in hepatocytes from rats pretreated with phenobarbital, whereas 1-nitropyrene was negative. These data indicate that the limited sensitivity to chemical carcinogens displayed by the hepatocyte/DNA repair assay may be increased by using hepatocytes isolated from animals exposed to hepatic mixed-function oxidase inducers.

Azure B, a metabolite of methylene blue, is a high-potency, reversible inhibitor of monoamine oxidase

Energy Technology Data Exchange (ETDEWEB)

Petzer, Anél, E-mail: 12264954@nwu.ac.za [Unit for Drug Research and Development, School of Pharmacy, North-West University, Private Bag X6001, Potchefstroom, 2520 (South Africa); Harvey, Brian H. [Division of Pharmacology, School of Pharmacy, North-West University, Private Bag X6001, Potchefstroom, 2520 (South Africa); Wegener, Gregers [Centre for Psychiatric Research, Aarhus University Hospital-Risskov, Skovagervej 2, 8240 Risskov (Denmark); Petzer, Jacobus P. [Division of Pharmaceutical Chemistry, School of Pharmacy, North-West University, Private Bag X6001, Potchefstroom, 2520 (South Africa)

2012-02-01

Methylene blue (MB) has been shown to act at multiple cellular and molecular targets and as a result possesses diverse medical applications. Among these is a high potency reversible inhibition of monoamine oxidase A (MAO-A) that may, at least in part, underlie its adverse effects but also its psycho- and neuromodulatory actions. MB is metabolized to yield N-demethylated products of which azure B, the monodemethyl species, is the major metabolite. Similar to MB, azure B also displays a variety of biological activities and may therefore contribute to the pharmacological profile of MB. Based on these observations, the present study examines the interactions of azure B with recombinant human MAO-A and -B. The results show that azure B is a potent MAO-A inhibitor (IC{sub 50} = 11 nM), approximately 6-fold more potent than is MB (IC{sub 50} = 70 nM) under identical conditions. Measurements of the time-dependency of inhibition suggest that the interaction of azure B with MAO-A is reversible. Azure B also reversibly inhibits the MAO-B isozyme with an IC{sub 50} value of 968 nM. These results suggest that azure B may be a hitherto under recognized contributor to the pharmacology and toxicology of MB by blocking central and peripheral MAO-A activity and as such needs to be considered during its use in humans and animals. Highlights: ► Methylene blue (MB) is a known potent MAO-A inhibitor. ► Azure B, the major metabolite of MB, is more potent as a MAO-A inhibitor. ► Azure B may be a contributor to the CNS pharmacology and toxicology of MB.

Substrate specificity of flavin-dependent vanillyl-alcohol oxidase from Penicillium simplicissimum.Evidence for the production of 4-hydroxycinnamyl alcohols from 4-allylphenols

NARCIS (Netherlands)

Fraaije, Marco W.; Veeger, Cees; Berkel, Willem J.H. van

1995-01-01

The substrate specificity of the flavoprotein vanillyl-alcohol oxidase from Penicillium simplicissimum was investigated. Vanillyl-alcohol oxidase catalyzes besides the oxidation of 4-hydroxybenzyl alcohols, the oxidative deamination of 4-hydroxybenzylamines and the oxidative demethylation of

Structure, organization, and transcriptional regulation of a family of copper radical oxidase genes in the lignin-degrading basidiomycete Phanerochaete chrysosporium

Science.gov (United States)

Amber Vanden Wymelenberg; Grzegorz Sabat; Michael Mozuch; Philip J. Kersten; Dan Cullen; Robert A. Blanchette

2006-01-01

The white rot basidiomycete Phanerochaete chrysosporium produces an array of nonspecific extracellular enzymes thought to be involved in lignin degradation, including lignin peroxidases, manganese peroxidases, and the H2O2-generating copper radical oxidase, glyoxal oxidase (GLX). Preliminary analysis of the P. chrysosporium draft genome had identified six sequences...

Addition of glucose oxidase for the improvement of refrigerated dough quality

Science.gov (United States)

Refrigerated dough encompasses a wide range of products and is a very popular choice for consumers. Two of the largest problems that occur during refrigerated dough storage are dough syruping and loss of dough strength. The goal of this study was to evaluate glucose oxidase as an additive to refri...

Copper radical oxidases and related extracellular oxidoreductases of wood-decay Agaricomycetes

Science.gov (United States)

Phil Kersten; Dan Cullen

2014-01-01

Extracellular peroxide generation, a key component of oxidative lignocellulose degradation, has been attributed to various enzymes including the copper radical oxidases. Encoded by a family of structurally related sequences, the genes are widely distributed among wood decay fungi including three recently completed polypore genomes. In all cases, core catalytic residues...

Monoamine oxidase inhibitory activity in tobacco particulate matter: Are harman and norharman the only physiologically relevant inhibitors?

Science.gov (United States)

Truman, Penelope; Grounds, Peter; Brennan, Katharine A

2017-03-01

Monoamine oxidase inhibition is significant in smokers, but it is still unclear how the inhibition that is seen in the brains and bodies of smokers is brought about. Our aim was to test the contribution of the harman and norharman in tobacco smoke to MAO-A inhibition from tobacco smoke preparations, as part of a re-examination of harman and norharman as the cause of the inhibition of MAO-A inhibition in the brain. Tobacco smoke particulate matter and cigarette smoke particulate matter were prepared and the amounts of harman and norharman measured. The results were compared with the total monoamine oxidase-A inhibitory activity. At a nicotine concentration of 0.6μM (a "physiological" concentration in blood) the total monoamine oxidase-A inhibitory activity measured in these samples was sufficient to inhibit the enzyme by approximately 10%. Of this inhibitory activity, only a small proportion of the total was found to be due to harman and norharman. These results show that harman and norharman provide only a moderate contribution to the total monoamine oxidase-A inhibitory activity of tobacco smoke, perhaps under 10%. This suggests that other inhibitors (either known or unknown) may be more significant contributors to total inhibitory activity than has yet been established, and deserve closer examination. Copyright © 2017 Elsevier B.V. All rights reserved.

Differences in activity of cytochrome C oxidase in brain between sleep and wakefulness.

Science.gov (United States)

Nikonova, Elena V; Vijayasarathy, Camasamudram; Zhang, Lin; Cater, Jacqueline R; Galante, Raymond J; Ward, Stephen E; Avadhani, Narayan G; Pack, Allan I

2005-01-01

Increased mRNA level of subunit 1 cytochrome c oxidase (COXI) during wakefulness and after short-term sleep deprivation has been described in brain. We hypothesized that this might contribute to increased activity of cytochrome oxidase (COX) enzyme during wakefulness, as part of the mechanisms to provide sufficient amounts of adenosine triphosphate to meet increased neuronal energy demands. COX activity was measured in isolated mitochondria from different brain regions in groups of rats with 3 hours of spontaneous sleep, 3 hours of spontaneous wake, and 3 hours of sleep deprivation. The group with 3 hours of spontaneous wake was added to delineate the circadian component of changes in the enzyme activity. Northern blot analysis was performed to examine the mRNA levels of 2 subunits of the enzyme COXI and COXIV, encoded by mitochondrial and nuclear DNA, respectively. Laboratory of Biochemistry, Department of Animal Biology, and Center for Sleep and Respiratory Neurobiology, University of Pennsylvania. 2-month-old male Fischer rats (N = 21) implanted for polygraphic recording. For COX activity, there was a main effect by analysis of variance of experimental group (P sleep-deprived groups as compared to the sleep group. A main effect of brain region was also significant (P sleep. There is an increase in COX activity after both 3 hours of spontaneous wake and 3 hours of sleep deprivation as compared with 3 hours of spontaneous sleep in diverse brain regions, which could be, in part, explained by the increased levels of bigenomic transcripts of the enzyme. This likely contributes to increased adenosine triphosphate production during wakefulness. ADP, adenosine diphosphate; ATP, adenosine triphosphate; COXI, cytochrome c oxidase subunit 1 mRNA; COX, cytochrome c oxidase (protein); CREB, cyclic AMP response element binding protein; DNA, deoxyribonucleic acid; EDTA, ethylenediaminetetraacetic acid; EEG, electroencephalography; EMG, electromyography; GABP, GA binding

Cyanide inhibition and pyruvate-induced recovery of cytochrome c oxidase

Czech Academy of Sciences Publication Activity Database

Nůsková, Hana; Vrbacký, Marek; Drahota, Zdeněk; Houštěk, Josef

2010-01-01

Roč. 42, č. 5 (2010), s. 395-403 ISSN 0145-479X R&D Projects: GA ČR(CZ) GA303/07/0781; GA MŠk(CZ) 1M0520; GA MŠk OC08017 Institutional research plan: CEZ:AV0Z50110509 Keywords : cytochrom c oxidase * cyanide * oxygen affinity Subject RIV: CE - Biochemistry Impact factor: 3.637, year: 2010

Redox-flexible NADH oxidase biosensor: A platform for various dehydrogenase bioassays and biosensors

International Nuclear Information System (INIS)

Serban, Simona; El Murr, Nabil

2006-01-01

A generic amperometric bioassay based on the enzymatic oxidation catalysed by the stable NADH oxidase (NAox) from Thermus thermophilus has been developed for NADH measurements. The NAox uses O 2 as its natural electron acceptor and produces H 2 O 2 in a two-electron process. Electrochemical and spectrophotometric experiments showed that the NAox used in this work, presents a very good activity towards its substrate and, in contrary to previously mentioned NADH oxidases, does not require the addition of any exogenous flavin cofactor neither to promote nor to maintain its activity. In addition, the NAox used also works with artificial electron acceptors like ferrocene derivatives. O 2 was successfully replaced by redox mediators such as hydroxymethyl ferrocene (FcCH 2 OH) for the regeneration of the active enzyme. Combining the NAox with the mediator and the horseradish peroxidase we developed an original, high sensitive 'redox-flexible' NADH amperometric bioassay working in a large window of applied potentials in both oxidation and reduction modes. The biosensor has a continuous and complementary linearity range permitting to measure NADH concentrations starting from 5 x 10 -6 M in reduction until 2 x 10 3 M in oxidation. This redox-flexibility allows choosing the applied potential in order to avoid electrochemical interferences. The association of the 'redox-flexible' concept with NADH dependent enzymes opens a novel strategy for dehydrogenases based bioassays and biosensors. The great number of dehydrogenases available makes the concept applicable for numerous substrates to analyse. Moreover it allows the development of a wide range of biosensors on the basis of a generic platform. This gives several advantages over the previous manufacturing techniques and offers a general and flexible scheme for the fabrication of biosensors presenting high sensitivities, wide calibration ranges and less affected by electrochemical interferences

Molecular characterisation and expression analysis of acc oxidase gene from guzmania ruiz and pav

International Nuclear Information System (INIS)

Jianxin, L.; Huaqiao, D.; Weiyong, W.; Danqing, T.

2017-01-01

ACC oxidase is the last key enzyme of ethylene synthesis pathway, while ethylene is a key factor affecting flowering in ornamental bromeliad. To understand ACC oxidase gene's characteristics and its effect on ornamental bromeliad flowering, we cloned 1504bp full-length cDNA sequence (GenBank: JX972145) and 2546bp corresponding genomic sequence (GenBank: JX972146)of GoACO1 (ACC oxidase gene) from Guzmania variety: Ostara. Prokaryotic expression study showed that expression of GoACO1 can produced a 41 KD protein precipitation in Escherichia coli DE3(BL-21); Real-time quantitative analysis showed that GoACO1 can express in all tested tissues including floral organ, bract, leaf and scape, and expression quantity in bract was the highest. Through constructing plant overexpression vector, transforming into Arabidopsis thaliana, and investigating blossom character of T2 generation seeds, we found that first flowering time of the goal Arabidopsis thaliana was 1.5 days earlier, and their peak flowering time(the number of flowering more than 50%) was 1.8 days earlier, compared with wild type one. Taken together, our results suggested that GoACO1can express in all kinds of tissues and seems to promote Arabidopsis thaliana flowering earlier. (author)

Asp170 is crucial for the redox properties of vanillyl-alcohol oxidase

NARCIS (Netherlands)

Heuvel, van den R.H.H.; Fraaije, M.W.; Mattevi, A.; Berkel, van W.J.H.

2000-01-01

Vanillyl-alcohol oxidase is a flavoprotein containing a covalent flavin that catalyzes the oxidation of 4-(methoxymethyl)phenol to 4-hydroxybenzaldehyde. The reaction proceeds through the formation of a p-quinone methide intermediate, after which, water addition takes place. Asp-170, located near

Asp-170 Is Crucial for the Redox Properties of Vanillyl-alcohol Oxidase

NARCIS (Netherlands)

Heuvel, Robert H.H. van den; Fraaije, Marco W.; Mattevi, Andrea; Berkel, Willem J.H. van

2000-01-01

Vanillyl-alcohol oxidase is a flavoprotein containing a covalent flavin that catalyzes the oxidation of 4-(methoxymethyl)phenol to 4-hydroxybenzaldehyde. The reaction proceeds through the formation of a p-quinone methide intermediate, after which, water addition takes place. Asp-170, located near

Glucose Oxidase Catalyzed Self-Assembly of Bioelectroactive Gold Nanostructures

Science.gov (United States)

2010-01-01

polymer matrix), however, electrons generated at the FAD/FADH2 active site of glucose oxidase (GOx) must tunnel ca. 15 through the protein shell...described as a surface bound thiolate [33]. Recently, the presence of free thiol groups has been proposed as a mechanism for gold reduction in pure enzymes...simultaneously [38]. The oxidative polymerization of the amines proceeds simulta- neously with the formation of gold nanoparticles such that the polymerized amine

[Substrate-inhibitory analysis of monoamine oxidase from hepatopancreas of the octopus Bathypolypus arcticus].

Science.gov (United States)

Basova, I N; Iagodina, O V

2012-01-01

Study of the substrate-inhibitory specificity of mitochondrial monoamine oxidase (MAO) of hepatopancreas of the octopus Bathypolypus arcticus revealed distinctive peculiarities of catalytic properties of this enzyme. The studied enzyme, on one hand, like the classic MAO of homoiothermal animals, is able to deaminate tyramine, serotonin, benzylamine, tryptamine, beta-phenylethylamine, while, on the other hand, deaminates histamine and does not deaminate putrescine--classic substrates of diamine oxidase (DAO). Results of the substrate-inhibitory analysis with use of chlorgiline and deprenyl are indirect proofs of the existence in the octopus hepatopancreas of one molecular MAO form. Semicarbazide and pyronine G turned out to be weak irreversible inhibitors, four derivatives of acridine--irreversible inhibitors of the intermediate effectiveness with respect to the octopus hepatopancreas MAO; specificity of action of inhibitors at deamination of different substrates was equal.

Nitric oxide partitioning into mitochondrial membranes and the control of respiration at cytochrome c oxidase

Science.gov (United States)

Shiva, Sruti; Brookes, Paul S.; Patel, Rakesh P.; Anderson, Peter G.; Darley-Usmar, Victor M.

2001-06-01

An emerging and important site of action for nitric oxide (NO) within cells is the mitochondrial inner membrane, where NO binds to and inhibits members of the electron transport chain, complex III and cytochrome c oxidase. Although it is known that inhibition of cytochrome c oxidase by NO is competitive with O2, the mechanisms that underlie this phenomenon remain unclear, and the impact of both NO and O2 partitioning into biological membranes has not been considered. These properties are particularly interesting because physiological O2 tensions can vary widely, with NO having a greater inhibitory effect at low O2 tensions (mitochondrial membranes in the absence of substrate, in a nonsaturable process that is O2 dependent. This consumption modulates inhibition of cytochrome c oxidase by NO and is enhanced by the addition of exogenous membranes. From these data, it is evident that the partition of NO into mitochondrial membranes has a major impact on the ability of NO to control mitochondrial respiration. The implications of this conclusion are discussed in the context of mitochondrial lipid:protein ratios and the importance of NO as a regulator of respiration in pathophysiology.

Cloning and sequencing of phenol oxidase 1 (pox1) gene from ...

African Journals Online (AJOL)

The gene (pox1) encoding a phenol oxidase 1 from Pleurotus ostreatus was sequenced and the corresponding pox1-cDNA was also synthesized, cloned and sequenced. The isolated gene is flanked by an upstream region called the promoter (399 bp) prior to the start codon (ATG). The putative metalresponsive elements ...

Gastric mucosal injury in the rat. Role of iron and xanthine oxidase

International Nuclear Information System (INIS)

Smith, S.M.; Grisham, M.B.; Manci, E.A.; Granger, D.N.; Kvietys, P.R.

1987-01-01

Recent studies have implicated oxygen free radicals in ischemia-reperfusion injury to the gastric mucosa. The aims of the present study were to test the hypothesis that the enzyme xanthine oxidase is the source of the oxygen radicals in the ischemic stomach and determine the importance of the iron-catalyzed Haber-Weiss reaction in generating the cytotoxic oxygen radicals. Gastric mucosal clearance of 51 Cr-labeled red blood cells was measured during a 30-min control period, a 30-min ischemic period (hemorrhage to 25 mmHg arterial pressure), and a 60-80-min reperfusion period (reinfusion of shed blood). In untreated (control) rats, a dramatic rise (100-fold) in the leakage of 51 Cr-labeled red blood cells into the gastric lumen was observed only during the reperfusion period. After the reperfusion period, gastric mucosal damage was further assessed using gross lesion area and histology. Rats were placed on a sodium tungstate diet (to inactivate xanthine oxidase), or treated with either deferoxamine (an iron chelating agent) or superoxide dismutase (a superoxide scavenger). All three interventions substantially reduced 51 Cr-labeled red blood cell clearance and gross lesion area relative to untreated rats. However, tissue injury assessed histologically was similar in both treated and untreated animals. The results of this study support the hypothesis that oxygen free radicals mediate the hemorrhagic shock-induced extravasation of red blood cells. The data also indicate that xanthine oxidase is the source of the oxy-radicals and that the iron-catalyzed Haber-Weiss reaction is largely responsible for hydroxyl radical generation in this model

Characterization of three bioenergetically active respiratory terminal oxidases in the cyanobacterium Synechocystis sp. strain PCC 6803.

Science.gov (United States)

Pils, D; Schmetterer, G

2001-09-25

Synechocystis sp. PCC 6803 contains three respiratory terminal oxidases (RTOs): cytochrome c oxidase (Cox), quinol oxidase (Cyd), and alternate RTO (ARTO). Mutants lacking combinations of the RTOs were used to characterize these key enzymes of respiration. Pentachlorophenol and 2-heptyl-4-hydroxy-quinoline-N-oxide inhibited Cyd completely, but had little effect on electron transport to the other RTOs. KCN inhibited all three RTOs but the in vivo K(I) for Cox and Cyd was quite different (7 vs. 27 microM), as was their affinity for oxygen (K(M) 1.0 vs. 0.35 microM). ARTO has a very low respiratory activity. However, when uptake of 3-O-methylglucose, an active H+ co-transport, was used to monitor energization of the cytoplasmic membrane, ARTO was similarly effective as the other RTOs. As removal of the gene for cytochrome c(553) had the same effects as removal of ARTO genes, we propose that the ARTO might be a second Cox. The possible functions, localization and regulation of the RTOs are discussed.

Comparison of the vanadate oxidase method with the diazo method for serum bilirubin determination in dog, monkey, and rat.

Science.gov (United States)

Ameri, Mehrdad; Schnaars, Henry; Sibley, John; Honor, David

2011-01-01

The most widely used method for bilirubin concentration determination is the diazo method, which measures the color of azobilirubin. The vanadate oxidase method is based on oxidation of bilirubin to biliverdin by vanadate. The objective of this study was to compare total and direct bilirubin concentration ([Bt] and [Bd], respectively) determined by the diazo and vanadate oxidase methods in pooled serum samples from dogs, monkeys, and rats spiked with panels of different concentrations of bilirubin standards. Pooled serum samples from 40 dogs, 40 monkeys, and 60 rats were spiked with either ditaurine conjugates of bilirubin or a standard reference material. The results obtained from both assays were compared using Deming regression analysis. The intra- and interassay precision, expressed as a percentage of the coefficient of variation (%CV), was determined for [Bt] and [Bd], and the mean percentage of recovery was calculated. The vanadate oxidase method displayed an excellent correlation (r  =  0.99-1.00) with the diazo method. Using Deming regression, there were minimal negative or positive constant and proportional biases for [Bt] and [Bd]. The precision studies revealed that the vanadate oxidase method has comparable between-run and within-run CVs to those of the diazo method. The recovery study demonstrated that the diazo method more closely approximates the expected values of [Bt]. In conclusion, the vanadate oxidase method is a simple and rapid method that can be employed as an alternative to the diazo method when interfering substances are present in the serum samples of dog, monkey, and rat.

Free radicals in hypoxic rat diaphragm contractility: no role for xanthine oxidase.

NARCIS (Netherlands)

Heunks, L.M.A.; Machiels, H.A.; Abreu, R.A. de; Zhu, X.; Heijden, E. van der; Dekhuijzen, P.N.R.

2001-01-01

Recent evidence indicates that hypoxia enhances the generation of oxidants. Little is known about the role of free radicals in contractility of the rat diaphragm during hypoxia. We hypothesized that antioxidants improve contractility of the hypoxic rat diaphragm and that xanthine oxidase (XO) is an

A kinetic study of soluble glucose oxidase using a rotating-disc electrode

NARCIS (Netherlands)

Stroe-Biezen, van S.A.M.; Janssen, A.P.M.; Janssen, L.J.J.

1994-01-01

In order to determine the kinetic parameters of glucose oxidation catalysed by the enzyme glucose oxidase (GO) the initial velocity of hydrogen peroxide formation was measured using a rotating disc electrode (RDE). The major advantage of this method is the possibility of continuous measurement of

Covalent immobilization of glucose oxidase on amino MOFs via post-synthetic modification

NARCIS (Netherlands)

Tudisco, C.G.; Zolubas, G.; Seoane de la Cuesta, B.; Zafarani, H.; Kazemzad Asiabi, M.; Gascon Sabate, J.; Hagedoorn, P.L.; Rassaei, L.

2016-01-01

The post-synthetic modification (PSM) of two amino-MOFs with glucose oxidase is reported in this study. The multi-step approach preserved the MOFs' structure and allowed the production of enzyme-functionalized MOFs (MOFs@GOx), which retained the enzymatic activity and showed selective properties

Electron transfer reactivity of the Arabidopsis thaliana sulfhydryl oxidase AtErv1

DEFF Research Database (Denmark)

Farver, Ole; Vitu, Elvira; Wherland, Scot

2009-01-01

The redox reactivity of the three disulfide bridges and the flavin present in each protomer of the wild-type Arabidopsis thaliana mitochondrial sulfhydryl oxidase (AtErv1) homodimer has been investigated. Pulse radiolytically produced CO2- radical ions were found to reduce the disulfide bridges...

Antiglycation, radical scavenging, and semicarbazide-sensitive amine oxidase inhibitory activities of acetohydroxamic acid in vitro

Directory of Open Access Journals (Sweden)

Liu YH

2017-07-01

Full Text Available Yuh-Hwa Liu,1,2,* Yeh-Lin Lu,3,* Der-Zen Liu,4 Wen-Chi Hou5 1Division of Gastroenterology, Shin Kong Wu Ho-Su Memorial Hospital, Taipei, Taiwan; 2Department of General Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan; 3Department of Pharmacy, Taipei Medical University, Taipei, Taiwan; 4Graduate Institute of Biomedical Materials and Tissue Engineering, Taipei Medical University, Taipei, Taiwan; 5Graduate Institute of Pharmacognosy, Taipei Medical University, Taipei, Taiwan *These authors contributed equally to this work Abstract: Advanced glycation endproducts (AGEs can promote intracellular reactive oxygen species production, and the levels of AGEs are highly correlated with cardiovascular disease and diabetes complications. Acetohydroxamic acid (acetH is a bacterial urease inhibitor drug used to treat kidney stones and infections in the urinary tract, and hydroxyurea (HU is a drug used for antineoplasm and sickle cell diseases. Both acetH and HU are hydroxamic acid derivatives. It was found that acetH and HU at 2.5 or 5 mM showed anti-AGE formation by lowering the AGEs’ fluorescent intensities and Nε-(carboxymethyllysine formation in bovine serum albumin/galactose models, and both showed better and significant differences (P<0.05 compared to the positive control of aminoguanidine. Regarding radical scavenging activities, the half-inhibition concentrations (IC50 of acetH against α,α-diphenyl-β-picrylhydrazyl radical and hydroxyl radical were 34.86 and 104.42 µM, respectively. The IC50 of acetH against semicarbazide-sensitive amine oxidase was 10.56 µM, and acetH showed noncompetitive inhibition respective to the substrates (benzylamine. The antiglycation, antioxidant, and semicarbazide-sensitive amine oxidase inhibitory activities of acetH prove that it has the potential for treating cardiovascular disease and diabetes complications and it needs further investigation in animal models. Keywords: acetH, AGEs, Nε

New biotechnological perspectives of a NADH oxidase variant from Thermus thermophilus HB27 as NAD+-recycling enzyme

Directory of Open Access Journals (Sweden)

Rocha-Martín Javier

2011-11-01

Full Text Available Abstract Background The number of biotransformations that use nicotinamide recycling systems is exponentially growing. For this reason one of the current challenges in biocatalysis is to develop and optimize more simple and efficient cofactor recycling systems. One promising approach to regenerate NAD+ pools is the use of NADH-oxidases that reduce oxygen to hydrogen peroxide while oxidizing NADH to NAD+. This class of enzymes may be applied to asymmetric reduction of prochiral substrates in order to obtain enantiopure compounds. Results The NADH-oxidase (NOX presented here is a flavoenzyme which needs exogenous FAD or FMN to reach its maximum velocity. Interestingly, this enzyme is 6-fold hyperactivated by incubation at high temperatures (80°C under limiting concentrations of flavin cofactor, a change that remains stable even at low temperatures (37°C. The hyperactivated form presented a high specific activity (37.5 U/mg at low temperatures despite isolation from a thermophile source. Immobilization of NOX onto agarose activated with glyoxyl groups yielded the most stable enzyme preparation (6-fold more stable than the hyperactivated soluble enzyme. The immobilized derivative was able to be reactivated under physiological conditions after inactivation by high solvent concentrations. The inactivation/reactivation cycle could be repeated at least three times, recovering full NOX activity in all cases after the reactivation step. This immobilized catalyst is presented as a recycling partner for a thermophile alcohol dehydrogenase in order to perform the kinetic resolution secondary alcohols. Conclusion We have designed, developed and characterized a heterogeneous and robust biocatalyst which has been used as recycling partner in the kinetic resolution of rac-1-phenylethanol. The high stability along with its capability to be reactivated makes this biocatalyst highly re-useable for cofactor recycling in redox biotransformations.

In vivo oxalate degradation by liposome encapsulated oxalate oxidase in rat model of hyperoxaluria

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Tulika Dahiya

2013-01-01

Interpretation & conclusions: EMA-oxalate oxidase encapsulated liposome caused oxalate degradation in experimental hyperoxaluria indicating that the enzyme could be used as a therapeutic agent in hyperoxaluria leading to urinary stones.

Galactose oxidase labeling of membrane proteins from human brain white matter

International Nuclear Information System (INIS)

Hukkanen, V.; Frey, H.; Salmi, A.

1981-01-01

Membrane proteins of human autopsy brain white matter were subjected to a galactose oxidase/NaB 3 H 4 labeling procedure and the membranes labeled by this method or by [ 3 H]acetic anhydride techniques were studied by lectin affinity chromatography using Lens culinaris phytohemagglutinin (lentil lectin) attached to Sepharose 4B beads. (Auth.)

Design of carbon nanotube-based gas-diffusion cathode for O{sub 2} reduction by multicopper oxidases

Energy Technology Data Exchange (ETDEWEB)

Lau, Carolin; Adkins, Emily R.; Atanassov, Plamen [University of New Mexico, Center for Emerging Energy Technologies, Albuquerque, NM (United States); Ramasamy, Ramaraja P. [Microbiology and Applied Biochemistry, Airbase Sciences, Air Force Research Laboratory, Tyndall Air Force Base, FL (United States); Nano-Electrochemistry Laboratory, Faculty of Engineering, University of Georgia, Athens, GA (United States); Luckarift, Heather R.; Johnson, Glenn R. [Microbiology and Applied Biochemistry, Airbase Sciences, Air Force Research Laboratory, Tyndall Air Force Base, FL (United States)

2012-01-15

Multicopper oxidases, such as laccase or bilirubin oxidase, are known to reduce molecular oxygen at very high redox potentials, which makes them attractive biocatalysts for enzymatic cathodes in biological fuel cells. By designing an enzymatic gas-diffusion electrode, molecular oxygen can be supplied through the gaseous phase, avoiding solubility and diffusion limitations typically associated with liquid electrolytes. In doing so, the current density of enzymatic cathodes can theoretically be enhanced. This publication presents a material study of carbon/Teflon composites that aim to optimize the functionality of the gas-diffusion and catalytic layers for application in enzymatic systems. The modification of the catalytic layer with multiwalled carbon nanotubes, for example, creates the basis for stronger {pi}-{pi} stacking interactions through tethered enzymatic linkers, such as pyrenes or perylene derivates. Cyclic voltammograms show the effective direct electron contact of laccase with carbon nanotube-modified electrodes via tethered crosslinking molecules as a model system. The polarization behavior of laccase-modified gas-diffusion electrodes reveals open-circuit potentials of +550 mV (versus Ag/AgCl) and current densities approaching 0.5 mA cm{sup 2} (at zero potential) in air-breathing mode. (Copyright copyright 2012 WILEY-VCH Verlag GmbH and Co. KGaA, Weinheim)

Pyranose 2-oxidase from Phanerochaete chrysosporium--Expression in E.coli and Biochemical Characterization

Czech Academy of Sciences Publication Activity Database

Pisanelli, I.; Kujawa, M.; Spadiut, O.; Kittl, R.; Halada, Petr; Volc, Jindřich; Mozuch, M. D.; Kersten, P.; Haltrich, D.; Peterbauer, C.

2009-01-01

Roč. 142, č. 2 (2009), s. 97-106 ISSN 0168-1656 Institutional research plan: CEZ:AV0Z50200510 Keywords : Pyranose 2-oxidase * Phanerochaete chrysosporium * Lignocellulose degradation Subject RIV: CE - Biochemistry Impact factor: 2.881, year: 2009

Involvement of NADH Oxidase in Competition and Endocarditis Virulence in Streptococcus sanguinis.

Science.gov (United States)

Ge, Xiuchun; Yu, Yang; Zhang, Min; Chen, Lei; Chen, Weihua; Elrami, Fadi; Kong, Fanxiang; Kitten, Todd; Xu, Ping

2016-05-01

Here, we report for the first time that the Streptococcus sanguinis nox gene encoding NADH oxidase is involved in both competition with Streptococcus mutans and virulence for infective endocarditis. An S. sanguinis nox mutant was found to fail to inhibit the growth of Streptococcus mutans under microaerobic conditions. In the presence of oxygen, the recombinant Nox protein of S. sanguinis could reduce oxygen to water and oxidize NADH to NAD(+) The oxidation of NADH to NAD(+) was diminished in the nox mutant. The nox mutant exhibited decreased levels of extracellular H2O2; however, the intracellular level of H2O2 in the mutant was increased. Furthermore, the virulence of the nox mutant was attenuated in a rabbit endocarditis model. The nox mutant also was shown to be more sensitive to blood killing, oxidative and acid stresses, and reduced growth in serum. Thus, NADH oxidase contributes to multiple phenotypes related to competitiveness in the oral cavity and systemic virulence. Copyright © 2016 Ge et al.

Preparation of biosensors by immobilization of polyphenol oxidase in conducting copolymers and their use in determination of phenolic compounds in red wine.

Science.gov (United States)

Böyükbayram, A Elif; Kiralp, Senem; Toppare, Levent; Yağci, Yusuf

2006-10-01

Electrochemically produced graft copolymers of thiophene capped polytetrahydofuran (TPTHF1 and TPTHF2) and pyrrole were achieved by constant potential electrolysis using sodium dodecylsulfate (SDS) as the supporting electrolyte. Characterizations were based on Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). Electrical conductivities were measured by the four-probe technique. Novel biosensors for phenolic compounds were constructed by immobilizing polyphenol oxidase (PPO) into conducting copolymers prepared by electropolymerization of pyrrole with thiophene capped polytetrahydrofuran. Kinetic parameters, maximum reaction rate (V(max)) and Michaelis-Menten constant (K(m)) and optimum conditions regarding temperature and pH were determined for the immobilized enzyme. Operational stability and shelf-life of the enzyme electrodes were investigated. Enzyme electrodes of polyphenol oxidase were used to determine the amount of phenolic compounds in two brands of Turkish red wines and found very useful owing to their high kinetic parameters and wide pH working range.

Effects of high pressure homogenization on the activity, stability, kinetics and three-dimensional conformation of a glucose oxidase produced by Aspergillus niger.

Directory of Open Access Journals (Sweden)

Alline Artigiani Lima Tribst

Full Text Available High pressure homogenization (HPH is a non-thermal method, which has been employed to change the activity and stability of biotechnologically relevant enzymes. This work investigated how HPH affects the structural and functional characteristics of a glucose oxidase (GO from Aspergillus niger. The enzyme was homogenized at 75 and 150 MPa and the effects were evaluated with respect to the enzyme activity, stability, kinetic parameters and molecular structure. The enzyme showed a pH-dependent response to the HPH treatment, with reduction or maintenance of activity at pH 4.5-6.0 and a remarkable activity increase (30-300% at pH 6.5 in all tested temperatures (15, 50 and 75°C. The enzyme thermal tolerance was reduced due to HPH treatment and the storage for 24 h at high temperatures (50 and 75°C also caused a reduction of activity. Interestingly, at lower temperatures (15°C the activity levels were slightly higher than that observed for native enzyme or at least maintained. These effects of HPH treatment on function and stability of GO were further investigated by spectroscopic methods. Both fluorescence and circular dichroism revealed conformational changes in the molecular structure of the enzyme that might be associated with the distinct functional and stability behavior of GO.

Isolation and Characterization of a Catabolite Repression-Insensitive Mutant of a Methanol Yeast, Candida boidinii A5, Producing Alcohol Oxidase in Glucose-Containing Medium

OpenAIRE

Sakai, Yasuyoshi; Sawai, Tohru; Tani, Yoshiki

1987-01-01

Mutants exhibiting alcohol oxidase (EC 1.1.3.13) activity when grown on glucose in the presence of methanol were found among 2-deoxyglucose-resistant mutants derived from a methanol yeast, Candida boidinii A5. One of these mutants, strain ADU-15, showed the highest alcohol oxidase activity in glucose-containing medium. The growth characteristics and also the induction and degradation of alcohol oxidase were compared with the parent strain and mutant strain ADU-15. In the parent strain, initia...

2-acetylphenol analogs as potent reversible monoamine oxidase inhibitors

Directory of Open Access Journals (Sweden)

Legoabe LJ

2015-07-01

Full Text Available Lesetja J Legoabe,1 Anél Petzer,1 Jacobus P Petzer1,21Centre of Excellence for Pharmaceutical Sciences, 2Department of Pharmaceutical Chemistry, School of Pharmacy, North-West University, Potchefstroom, South AfricaAbstract: Based on a previous report that substituted 2-acetylphenols may be promising leads for the design of novel monoamine oxidase (MAO inhibitors, a series of C5-substituted 2-acetylphenol analogs (15 and related compounds (two were synthesized and evaluated as inhibitors of human MAO-A and MAO-B. Generally, the study compounds exhibited inhibitory activities against both MAO-A and MAO-B, with selectivity for the B isoform. Among the compounds evaluated, seven compounds exhibited IC50 values <0.01 µM for MAO-B inhibition, with the most selective compound being 17,000-fold selective for MAO-B over the MAO-A isoform. Analyses of the structure–activity relationships for MAO inhibition show that substitution on the C5 position of the 2-acetylphenol moiety is a requirement for MAO-B inhibition, and the benzyloxy substituent is particularly favorable in this regard. This study concludes that C5-substituted 2-acetylphenol analogs are potent and selective MAO-B inhibitors, appropriate for the design of therapies for neurodegenerative disorders such as Parkinson’s disease.Keywords: monoamine oxidase, MAO, inhibition, 2-acetylphenol, structure–activity relationship