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Sample records for bilirubin oxidase bound

  1. Bilirubin oxidases in bioelectrochemistry: features and recent findings.

    PubMed

    Mano, Nicolas; Edembe, Lise

    2013-12-15

    Bilirubin oxidases, a sub class of the Multicopper oxidases family, were discovered in 1981 by Tanaka and Murao (Murao and Tanaka, 1981) and first used for the detection of bilirubin. Since 2001 and the pioneering work of Tsujimura, these BODs have attracted a lot of attention for the reduction of O2. Unlike laccases, these BODs are stable in physiological conditions (20mM phosphate buffer, pH 7.4, 0.14 M NaCl, 37 °C) and more than 120 papers have been published in the last 7 years. Here, we will first briefly describe some general features of BODs and then review the use of BODs for bilirubin biosensors and the recent achievements and progress toward the elaboration of efficient O2 reducing cathodes. PMID:23911663

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

    Technology Transfer Automated Retrieval System (TEKTRAN)

    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...

  3. Oxygen biosensor based on bilirubin oxidase immobilized on a nanostructured gold electrode.

    PubMed

    Pita, Marcos; Gutierrez-Sanchez, Cristina; Toscano, Miguel D; Shleev, Sergey; De Lacey, Antonio L

    2013-12-01

    Gold disk electrodes modified with gold nanoparticles have been used as a scaffold for the covalent immobilization of bilirubin oxidase. The nanostructured bioelectrodes were tested as mediator-less biosensors for oxygen in a buffer that mimics the content and the composition of human physiological fluids. Chronoamperometry measurements showed a detection limit towards oxygen of 6 ± 1 μM with a linear range of 6-300 μM, i.e. exceeding usual physiological ranges of oxygen in human tissues and fluids. The biosensor presented is the first ever-reported oxygen amperometric biosensor based on direct electron transfer of bilirubin oxidase. PMID:23973738

  4. Protein-Support Interactions for Rationally Designed Bilirubin Oxidase Based Cathode: A Computational Study.

    PubMed

    Matanovic, Ivana; Babanova, Sofia; Chavez, Madelaine Seow; Atanassov, Plamen

    2016-04-21

    An example of biocathode based on bilirubin oxidase (BOx) was used to demonstrate how density functional theory can be combined with docking simulations in order to study the interface interactions between the enzyme and specifically designed electrode surface. The electrode surface was modified through the adsorption of bilirubin, the natural substrate for BOx, and the prepared electrode was electrochemically characterized using potentiostatic measurements. The experimentally determined current densities showed that the presence of bilirubin led to significant improvement of the cathode operation. On the basis of the computationally calculated binding energies of bilirubin to the graphene support and BOx and the analysis of the positioning of bilirubin relative to the support and T1 Cu atom of the enzyme, we hypothesize that the bilirubin serves as a geometric and electronic extension of the support. The computational results further confirm that the modification of the electrode surface with bilirubin provides an optimal orientation of BOx toward the support but also show that bilirubin facilitates the interfacial electron transfer by decreasing the distance between the electrode surface and the T1 Cu atom. PMID:27015361

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

    SciTech Connect

    Abu-Bakar, A'edah; Arthur, Dionne Maioha; Aganovic, Simona; Ng, Jack C.; Lang, Matti A.

    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 change the hepatic CYP2A5

  6. Fully Oriented Bilirubin Oxidase on Porphyrin-Functionalized Carbon Nanotube Electrodes for Electrocatalytic Oxygen Reduction.

    PubMed

    Lalaoui, Noémie; Le Goff, Alan; Holzinger, Michael; Cosnier, Serge

    2015-11-16

    The efficient immobilization and orientation of bilirubin oxidase from Myrothecium verrucaria on multi-walled carbon nanotube electrodes by using π-stacked porphyrins as a direct electron-transfer promoter is reported. By comparing the use of different types of porphyrin, the rational effect of the porphyrin structure on both the immobilization and orientation of the enzyme is demonstrated. The best performances were obtained for protoporphyrin IX, which is the natural precursor of bilirubin. These electrodes exhibit full orientation of the enzyme, as confirmed by the observable non-catalytic redox system corresponding to the T1 copper center associated with pure Nernstian electrocatalytic behavior with high catalytic currents of almost 5 mA cm(-2) at neutral pH. PMID:26449635

  7. Circular dichroism study of the interaction between mutagens and bilirubin bound to different binding sites of serum albumins.

    PubMed

    Orlov, Sergey; Goncharova, Iryna; Urbanová, Marie

    2014-05-21

    Although recent investigations have shown that bilirubin not only has a negative role in the organism but also exhibits significant antimutagenic properties, the mechanisms of interactions between bilirubin and mutagens are not clear. In this study, interaction between bilirubin bound to different binding sites of mammalian serum albumins with structural analogues of the mutagens 2-aminofluorene, 2,7-diaminofluorene and mutagen 2,4,7-trinitrofluorenone were investigated by circular dichroism and absorption spectroscopy. Homological human and bovine serum albumins were used as chiral matrices, which preferentially bind different conformers of bilirubin in the primary binding sites and make it observable by circular dichroism. These molecular systems approximated a real system for the study of mutagens in blood serum. Differences between the interaction of bilirubin bound to primary and to secondary binding sites of serum albumins with mutagens were shown. For bilirubin bound to secondary binding sites with low affinity, partial displacement and the formation of self-associates were observed in all studied mutagens. The associates of bilirubin bound to primary binding sites of serum albumins are formed with 2-aminofluorene and 2,4,7-trinitrofluorenone. It was proposed that 2,7-diaminofluorene does not interact with bilirubin bound to primary sites of human and bovine serum albumins due to the spatial hindrance of the albumins binding domains. The spatial arrangement of the bilirubin bound to serum albumin along with the studied mutagens was modelled using ligand docking, which revealed a possibility of an arrangement of the both bilirubin and 2-aminofluorene and 2,4,7-trinitrofluorenone in the primary binding site of human serum albumin. PMID:24589992

  8. Bilirubin oxidase based enzymatic air-breathing cathode: Operation under pristine and contaminated conditions.

    PubMed

    Santoro, Carlo; Babanova, Sofia; Erable, Benjamin; Schuler, Andrew; Atanassov, Plamen

    2016-04-01

    The performance of bilirubin oxidase (BOx) based air breathing cathode was constantly monitored over 45 days. The effect of electrolyte composition on the cathode oxygen reduction reaction (ORR) output was investigated. Particularly, deactivation of the electrocatalytic activity of the enzyme in phosphate buffer saline (PBS) solution and in activated sludge (AS) was evaluated. The greatest drop in current density was observed during the first 3 days of constant operation with a decrease of ~60 μA cm(-2) day(-1). The rate of decrease slowed to ~10 μA cm(-2) day(-1) (day 3 to 9) and then to ~1.5 μA cm(-2)day(-1) thereafter (day 9 to 45). Despite the constant decrease in output, the BOx cathode generated residual current after 45 days operations with an open circuit potential (OCP) of 475 mV vs. Ag/AgCl. Enzyme deactivation was also studied in AS to simulate an environment close to the real waste operation with pollutants, solid particles and bacteria. The presence of low-molecular weight soluble contaminants was identified as the main reason for an immediate enzymatic deactivation within few hours of cathode operation. The presence of solid particles and bacteria does not affect the natural degradation of the enzyme. PMID:26544631

  9. Increasing the catalytic activity of Bilirubin oxidase from Bacillus pumilus: Importance of host strain and chaperones proteins.

    PubMed

    Gounel, Sébastien; Rouhana, Jad; Stines-Chaumeil, Claire; Cadet, Marine; Mano, Nicolas

    2016-07-20

    Aggregation of recombinant proteins into inclusion bodies (IBs) is the main problem of the expression of multicopper oxidase in Escherichia coli. It is usually attributed to inefficient folding of proteins due to the lack of copper and/or unavailability of chaperone proteins. The general strategies reported to overcome this issue have been focused on increasing the intracellular copper concentration. Here we report a complementary method to optimize the expression in E. coli of a promising Bilirubin oxidase (BOD) isolated from Bacillus pumilus. First, as this BOD has a disulfide bridge, we switched E.coli strain from BL21 (DE3) to Origami B (DE3), known to promote the formation of disulfide bridges in the bacterial cytoplasm. In a second step, we investigate the effect of co-expression of chaperone proteins on the protein production and specific activity. Our strategy allowed increasing the final amount of enzyme by 858% and its catalytic rate constant by 83%. PMID:27165502

  10. Bilirubin Oxidase from Myrothecium verrucaria Physically Absorbed on Graphite Electrodes. Insights into the Alternative Resting Form and the Sources of Activity Loss

    PubMed Central

    Tasca, Federico; Farias, Diego; Castro, Carmen; Acuna-Rougier, Cristina; Antiochia, Riccarda

    2015-01-01

    The oxygen reduction reaction is one of the most important chemical processes in energy converting systems and living organisms. Mediator-less, direct electro-catalytic reduction of oxygen to water was achieved on spectrographite electrodes modified by physical adsorption of bilirubin oxidases from Myrothecium verrucaria. The existence of an alternative resting form of the enzyme is validated. The effect on the catalytic cycle of temperature, pH and the presence of halogens in the buffer was investigated. Previous results on the electrochemistry of bilirubin oxidase and on the impact of the presence of halogens are reviewed and reinterpreted. PMID:26196288

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

    SciTech Connect

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

    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.

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

    PubMed Central

    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

    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 P63. 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. PMID:20606269

  13. Substrate recycling scheme for tetrachloro-p-benzoquinone using bilirubin oxidase and NADH: Application for pentachlorophenol assay

    SciTech Connect

    Cybulski, D.; Luong, J.H.T.; Male, K.B.; Scharer, J.M.; Moo-Young, M.

    1999-03-01

    A novel assay for tetrachloro-p-benzoquinone (TCBQ), the main oxidation product of pentachlorophenol (PCP), was developed using bilirubin oxidase (BOX) in the presence of excess NADH. TCBQ was easily and rapidly reduced by NADH to 1,4-tetrachlorohydroquinone (TCHQ), which was then recycled back to TCBQ by the enzyme. BOX exhibited no reactivity toward NADH while its catalytic activity for the oxidation of TCHQ was very high. Under an optimized condition, the rate of NADH consumption determined by measuring the absorbance decrease at 340 nm yielded a detection limit for TCBQ of 110 nM. Fluorescence detection of the NADH using a lower enzyme concentration with excitation and emission wavelengths of 345 and 450 nm, respectively, allowed for a TCBQ detection limit of 30 nM. PCP was oxidized to TCBQ with high yield using bis(trifluoroacetoxy)iodobenzene in 0.05 M trichloroacetic acid. Coupling this oxidation reaction to the BOX/NADH assay attained PCT detection limits of 170 and 50 nM using absorbance and fluorescence measurements, respectively. When tested on PCP-contaminated soil samples, the BOX assay compared very well with HPLC measurements.

  14. How the Intricate Interactions between Carbon Nanotubes and Two Bilirubin Oxidases Control Direct and Mediated O2 Reduction.

    PubMed

    Mazurenko, Ievgen; Monsalve, Karen; Rouhana, Jad; Parent, Philippe; Laffon, Carine; Goff, Alan Le; Szunerits, Sabine; Boukherroub, Rabah; Giudici-Orticoni, Marie-Thérèse; Mano, Nicolas; Lojou, Elisabeth

    2016-09-01

    Due to the lack of a valid approach in the design of electrochemical interfaces modified with enzymes for efficient catalysis, many oxidoreductases are still not addressed by electrochemistry. We report in this work an in-depth study of the interactions between two different bilirubin oxidases, (from the fungus Myrothecium verrucaria and from the bacterium Bacillus pumilus), catalysts of oxygen reduction, and carbon nanotubes bearing various surface charges (pristine, carboxylic-, and pyrene-methylamine-functionalized). The surface charges and dipole moment of the enzymes as well as the surface state of the nanomaterials are characterized as a function of pH. An original electrochemical approach allows determination of the best interface for direct or mediated electron transfer processes as a function of enzyme, nanomaterial type, and adsorption conditions. We correlate these experimental results to theoric voltammetric curves. Such an integrative study suggests strategies for designing efficient bioelectrochemical interfaces toward the elaboration of biodevices such as enzymatic fuel cells for sustainable electricity production. PMID:27533778

  15. Bilirubin - urine

    MedlinePlus

    ... or gallbladder Considerations Bilirubin can break down in light. That is why babies with jaundice are sometimes placed under blue fluorescent lamps. Alternative Names Conjugated bilirubin - urine; Direct bilirubin - ...

  16. Bilirubin Test

    MedlinePlus

    ... test in conjunction with other laboratory tests ( alkaline phosphatase , aspartate aminotransferase , alanine aminotransferase ) when someone shows signs ... Gilbert syndrome, due to low levels of the enzyme that produces conjugated bilirubin If conjugated (direct) bilirubin ...

  17. Biofuel cells based on direct enzyme-electrode contacts using PQQ-dependent glucose dehydrogenase/bilirubin oxidase and modified carbon nanotube materials.

    PubMed

    Scherbahn, V; Putze, M T; Dietzel, B; Heinlein, T; Schneider, J J; Lisdat, F

    2014-11-15

    Two types of carbon nanotube electrodes (1) buckypaper (BP) and (2) vertically aligned carbon nanotubes (vaCNT) have been used for elaboration of glucose/O2 enzymatic fuel cells exploiting direct electron transfer. For the anode pyrroloquinoline quinone dependent glucose dehydrogenase ((PQQ)GDH) has been immobilized on [poly(3-aminobenzoic acid-co-2-methoxyaniline-5-sulfonic acid), PABMSA]-modified electrodes. For the cathode bilirubin oxidase (BOD) has been immobilized on PQQ-modified electrodes. PABMSA and PQQ act as promoter for enzyme bioelectrocatalysis. The voltammetric characterization of each electrode shows current densities in the range of 0.7-1.3 mA/cm(2). The BP-based fuel cell exhibits maximal power density of about 107 µW/cm(2) (at 490 mV). The vaCNT-based fuel cell achieves a maximal power density of 122 µW/cm(2) (at 540 mV). Even after three days and several runs of load a power density over 110 µW/cm(2) is retained with the second system (10mM glucose). Due to a better power exhibition and an enhanced stability of the vaCNT-based fuel cells they have been studied in human serum samples and a maximal power density of 41 µW/cm(2) (390 mV) can be achieved. PMID:24967753

  18. Membraneless enzymatic ethanol/O2 fuel cell: Transitioning from an air-breathing Pt-based cathode to a bilirubin oxidase-based biocathode

    NASA Astrophysics Data System (ADS)

    Aquino Neto, Sidney; Milton, Ross D.; Hickey, David P.; De Andrade, Adalgisa R.; Minteer, Shelley D.

    2016-08-01

    The bioelectrooxidation of ethanol was investigated in a fully enzymatic membraneless ethanol/O2 biofuel cell assembly using hybrid bioanodes containing multi-walled carbon nanotube (MWCNT)-decorated gold metallic nanoparticles with either a pyrroloquinoline quinone (PQQ)-dependent alcohol dehydrogenase (ADH) enzyme or a nicotinamide adenine dinucleotide (NAD+)-dependent ADH enzyme. The biofuel cell anode was prepared with the PQQ-dependent enzyme and designed using either a direct electron transfer (DET) architecture or via a mediated electron transfer (MET) configuration through a redox polymer, 1,1‧-dimethylferrocene-modified linear polyethyleneimine (FcMe2-C3-LPEI). In the case of the bioanode containing the NAD+-dependent enzyme, only the mediated electron transfer mechanism was employed using an electropolymerized methylene green film to regenerate the NAD+ cofactor. Regardless of the enzyme being employed at the anode, a bilirubin oxidase-based biocathode prepared within a DET architecture afforded efficient electrocatalytic oxygen reduction in an ethanol/O2 biofuel cell. The power curves showed that DET-based bioanodes via the PQQ-dependent ADH still lack high current densities, whereas the MET architecture furnished maximum power density values as high as 226 ± 21 μW cm-2. Considering the complete membraneless enzymatic biofuel cell with the NAD+-dependent ADH-based bioanode, power densities as high as 111 ± 14 μW cm-2 were obtained. This shows the advantage of PQQ-dependent ADH for membraneless ethanol/O2 biofuel cell applications.

  19. Membraneless enzymatic ethanol/O2 fuel cell: Transitioning from an air-breathing Pt-based cathode to a bilirubin oxidase-based biocathode

    NASA Astrophysics Data System (ADS)

    Aquino Neto, Sidney; Milton, Ross D.; Hickey, David P.; De Andrade, Adalgisa R.; Minteer, Shelley D.

    2016-08-01

    The bioelectrooxidation of ethanol was investigated in a fully enzymatic membraneless ethanol/O2 biofuel cell assembly using hybrid bioanodes containing multi-walled carbon nanotube (MWCNT)-decorated gold metallic nanoparticles with either a pyrroloquinoline quinone (PQQ)-dependent alcohol dehydrogenase (ADH) enzyme or a nicotinamide adenine dinucleotide (NAD+)-dependent ADH enzyme. The biofuel cell anode was prepared with the PQQ-dependent enzyme and designed using either a direct electron transfer (DET) architecture or via a mediated electron transfer (MET) configuration through a redox polymer, 1,1‧-dimethylferrocene-modified linear polyethyleneimine (FcMe2-C3-LPEI). In the case of the bioanode containing the NAD+-dependent enzyme, only the mediated electron transfer mechanism was employed using an electropolymerized methylene green film to regenerate the NAD+ cofactor. Regardless of the enzyme being employed at the anode, a bilirubin oxidase-based biocathode prepared within a DET architecture afforded efficient electrocatalytic oxygen reduction in an ethanol/O2 biofuel cell. The power curves showed that DET-based bioanodes via the PQQ-dependent ADH still lack high current densities, whereas the MET architecture furnished maximum power density values as high as 226 ± 21 μW cm-2. Considering the complete membraneless enzymatic biofuel cell with the NAD+-dependent ADH-based bioanode, power densities as high as 111 ± 14 μW cm-2 were obtained. This shows the advantage of PQQ-dependent ADH for membraneless ethanol/O2 biofuel cell applications.

  20. Bilirubin - blood

    MedlinePlus

    ... bilirubin is not processed normally by the liver ( Gilbert disease ) The following problems with gallbladder or bile ... Choledocholithiasis Cirrhosis Crigler-Najjar syndrome Dubin-Johnson syndrome Gilbert disease Glucose-6-phosphate dehydrogenase deficiency Hemoglobin Hemolytic ...

  1. Two-dimensional crystallization of monomeric bovine cytochrome c oxidase with bound cytochrome c in reconstituted lipid membranes.

    PubMed

    Osuda, Yukiho; Shinzawa-Itoh, Kyoko; Tani, Kazutoshi; Maeda, Shintaro; Yoshikawa, Shinya; Tsukihara, Tomitake; Gerle, Christoph

    2016-06-01

    Mitochondrial cytochrome c oxidase utilizes electrons provided by cytochrome c for the active vectorial transport of protons across the inner mitochondrial membrane through the reduction of molecular oxygen to water. Direct structural evidence on the transient cytochrome c oxidase-cytochrome c complex thus far, however, remains elusive and its physiological relevant oligomeric form is unclear. Here, we report on the 2D crystallization of monomeric bovine cytochrome c oxidase with tightly bound cytochrome c at a molar ratio of 1:1 in reconstituted lipid membranes at the basic pH of 8.5 and low ionic strength. PMID:26754561

  2. Purification and structural analysis of membrane-bound polyphenol oxidase from Fuji apple.

    PubMed

    Liu, Fang; Zhao, Jin-Hong; Wen, Xin; Ni, Yuan-Ying

    2015-09-15

    Membrane-bound polyphenol oxidase (mPPO) in Fuji apple (Malus domestica Borkh. cv. Red Fuji) was purified and analyzed with a nanoelectrospray ionization mass spectrometer. The three-dimensional model and binding site of mPPO to 4-methyl catechol were also studied using molecular docking. mPPO was purified 54.41-fold using temperature-induced phase partitioning technique and ion exchange chromatography. mPPO had a molecular weight of 67.3kDa. Even though a significant level of homology was observed between mPPO and the soluble polyphenol oxidase in the copper binding sequence, there was another region, rich in histidine residues, which differed in 13 amino acids. The three-dimensional structure of mPPO consisted of six α-helices, two short β-strands, and ten random coils. The putative substrate-binding pocket contained six polar or charged amino acids, His191, His221, Trp224, Trp228, Phe227, and Val190. Trp224 and Trp228 formed hydrogen bonds with 4-methyl-catechol. PMID:25863612

  3. The reaction pathway of membrane-bound rat liver mitochondrial monoamine oxidase

    PubMed Central

    Houslay, Miles D.; Tipton, Keith F.

    1973-01-01

    1. A preparation of a partly purified mitochondrial outer-membrane fraction suitable for kinetic investigations of monoamine oxidase is described. 2. An apparatus suitable for varying the O2 concentration in a spectrophotometer cuvette is described. 3. The reaction catalysed by the membrane-bound enzyme is shown to proceed by a double-displacement (Ping Pong) mechanism, and a formal mechanism is proposed. 4. KCN, NaN3, benzyl cyanide and 4-cyanophenol are shown to be reversible inhibitors of the enzyme. 5. The non-linear reciprocal plot obtained with impure preparations of benzylamine, which is typical of high substrate inhibition, is shown to be due to aldehyde contamination of the substrate. PMID:4778271

  4. Bilirubin present in diverse angiosperms

    PubMed Central

    Pirone, Cary; Johnson, Jodie V.; Quirke, J. Martin E.; Priestap, Horacio A.; Lee, David

    2010-01-01

    Background and aims Bilirubin is an orange-yellow tetrapyrrole produced from the breakdown of heme by mammals and some other vertebrates. Plants, algae and cyanobacteria synthesize molecules similar to bilirubin, including the protein-bound bilins and phytochromobilin which harvest or sense light. Recently, we discovered bilirubin in the arils of Strelitzia nicolai, the White Bird of Paradise Tree, which was the first example of this molecule in a higher plant. Subsequently, we identified bilirubin in both the arils and the flowers of Strelitzia reginae, the Bird of Paradise Flower. In the arils of both species, bilirubin is present as the primary pigment, and thus functions to produce colour. Previously, no tetrapyrroles were known to generate display colour in plants. We were therefore interested in determining whether bilirubin is broadly distributed in the plant kingdom and whether it contributes to colour in other species. Methodology In this paper, we use HPLC/UV and HPLC/UV/electrospray ionization-tandem mass spectrometry (HPLC/UV/ESI-MS/MS) to search for bilirubin in 10 species across diverse angiosperm lineages. Principal results Bilirubin was present in eight species from the orders Zingiberales, Arecales and Myrtales, but only contributed to colour in species within the Strelitziaceae. Conclusions The wide distribution of bilirubin in angiosperms indicates the need to re-assess some metabolic details of an important and universal biosynthetic pathway in plants, and further explore its evolutionary history and function. Although colour production was limited to the Strelitziaceae in this study, further sampling may indicate otherwise. PMID:22476078

  5. Two-dimensional crystallization of monomeric bovine cytochrome c oxidase with bound cytochrome c in reconstituted lipid membranes

    PubMed Central

    Osuda, Yukiho; Shinzawa-Itoh, Kyoko; Tani, Kazutoshi; Maeda, Shintaro; Yoshikawa, Shinya; Tsukihara, Tomitake; Gerle, Christoph

    2016-01-01

    Mitochondrial cytochrome c oxidase utilizes electrons provided by cytochrome c for the active vectorial transport of protons across the inner mitochondrial membrane through the reduction of molecular oxygen to water. Direct structural evidence on the transient cytochrome c oxidase–cytochrome c complex thus far, however, remains elusive and its physiological relevant oligomeric form is unclear. Here, we report on the 2D crystallization of monomeric bovine cytochrome c oxidase with tightly bound cytochrome c at a molar ratio of 1:1 in reconstituted lipid membranes at the basic pH of 8.5 and low ionic strength. PMID:26754561

  6. Purification and characterization of membrane-bound semicarbazide-sensitive amine oxidase (SSAO) from bovine lung.

    PubMed Central

    Lizcano, J M; Tipton, K F; Unzeta, M

    1998-01-01

    Semicarbazide-sensitive amine oxidase (SSAO) has been purified from bovine lung microsomes in a form which is catalytically active and stable to storage. The enzyme, an integral membrane protein, was solubilized with Triton X-100 and purification was achieved, in the presence of detergent, by chromatography with Cibacron Blue 3GA-agarose, hydroxylapatite, Lens culinaris-agarose, Resource Q-FPLC and gel filtration on Superdex 200 HR-FPLC. This is the first reported procedure for the extensive purification of a membrane-bound SSAO. The purified enzyme had an apparent Mr of 400000 but exhibited microheterogeneity with SDS/PAGE and isoelectric focusing, probably as a result of its glycoprotein nature. It behaved as a tetramer with subunits with apparent Mr values of 100. Antibodies raised towards the purified enzyme cross-reacted with the enzymes from human lung and bovine plasma. Redox-cycling staining and reaction with carbonyl reagents were consistent with the presence of a quinone cofactor, possibly topa quinone. The enzyme was also shown to contain two mol of Cu/mol of enzyme and removal of half of this bound copper resulted essentially in complete inhibition of enzyme activity. In contrast to the reported behaviour of the SSAO enzymes from plasma, the bovine lung enzyme was relatively insensitive to inhibition by cyanide, copper-chelating agents and amiloride. The specificity of the bovine lung enzyme was also narrower than reported for soluble SSAO. It catalysed the oxidative deamination of benzylamine, methylamine, 2-phenylethylamine and histamine but had no significant activity towards dopamine, 5-hydroxytryptamine, tryptamine or tyramine. PMID:9512463

  7. Fluorescent protein-based detection of unconjugated bilirubin in newborn serum

    PubMed Central

    Iwatani, Sota; Nakamura, Hajime; Kurokawa, Daisuke; Yamana, Keiji; Nishida, Kosuke; Fukushima, Sachiyo; Koda, Tsubasa; Nishimura, Noriyuki; Nishio, Hisahide; Iijima, Kazumoto; Miyawaki, Atsushi; Morioka, Ichiro

    2016-01-01

    Increased serum levels of unconjugated bilirubin are associated with the development of brain damage in newborns. In current clinical settings, there are no methods for directly determining serum levels of unconjugated bilirubin. UnaG, a fluorescent protein from Japanese eel muscle that specifically binds to unconjugated bilirubin was used in this study. Linear regression analysis was carried out to compare unconjugated bilirubin levels measured by UnaG and conventional bilirubin oxidase methods. Unconjugated bilirubin levels in the serum of newborns who were untreated or treated with phototherapy were compared. Effects of interfering factors in the serum (conjugated bilirubin, hemoglobin, and lipid) on unconjugated bilirubin concentration measured by the UnaG method were also evaluated. Unconjugated bilirubin levels measured by the UnaG method were highly correlated with those determined by the bilirubin oxidase assay. Unconjugated bilirubin levels determined by bilirubin oxidase and UnaG assays were similar in serum samples containing conjugated bilirubin. The performance of the UnaG assay was unaffected by phototherapy and the presence of serum hemoglobin and lipid emulsion. These results demonstrate the clinical applicability of the UnaG method for direct measurement of unconjugated bilirubin levels in newborn serum. PMID:27324682

  8. Fluorescent protein-based detection of unconjugated bilirubin in newborn serum.

    PubMed

    Iwatani, Sota; Nakamura, Hajime; Kurokawa, Daisuke; Yamana, Keiji; Nishida, Kosuke; Fukushima, Sachiyo; Koda, Tsubasa; Nishimura, Noriyuki; Nishio, Hisahide; Iijima, Kazumoto; Miyawaki, Atsushi; Morioka, Ichiro

    2016-01-01

    Increased serum levels of unconjugated bilirubin are associated with the development of brain damage in newborns. In current clinical settings, there are no methods for directly determining serum levels of unconjugated bilirubin. UnaG, a fluorescent protein from Japanese eel muscle that specifically binds to unconjugated bilirubin was used in this study. Linear regression analysis was carried out to compare unconjugated bilirubin levels measured by UnaG and conventional bilirubin oxidase methods. Unconjugated bilirubin levels in the serum of newborns who were untreated or treated with phototherapy were compared. Effects of interfering factors in the serum (conjugated bilirubin, hemoglobin, and lipid) on unconjugated bilirubin concentration measured by the UnaG method were also evaluated. Unconjugated bilirubin levels measured by the UnaG method were highly correlated with those determined by the bilirubin oxidase assay. Unconjugated bilirubin levels determined by bilirubin oxidase and UnaG assays were similar in serum samples containing conjugated bilirubin. The performance of the UnaG assay was unaffected by phototherapy and the presence of serum hemoglobin and lipid emulsion. These results demonstrate the clinical applicability of the UnaG method for direct measurement of unconjugated bilirubin levels in newborn serum. PMID:27324682

  9. Blood Test: Bilirubin

    MedlinePlus

    ... two forms in the body: indirect (unconjugated) and direct (conjugated). Indirect bilirubin, which doesn't dissolve in ... liver to be changed into the soluble form, direct bilirubin. Why It's Done Healthy newborns — especially those ...

  10. Enzymic oxidation of unconjugated bilirubin by rat liver.

    PubMed Central

    Cardenas-Vazquez, R; Yokosuka, O; Billing, B H

    1986-01-01

    The presence of the enzyme bilirubin oxidase, which degrades bilirubin in vitro, was demonstrated in the liver. Subcellular-fractionation experiments indicate that bilirubin oxidase is located in both the inner and outer membranes of the mitochondria. The mean rate of the reaction is 1.57 +/- 0.38 (S.D.) nmol of bilirubin degraded/min per mg of mitochondrial protein (munits/mg of protein). With respect to the overall breakdown of bilirubin, the enzyme has a Km' of 136 microM-bilirubin and a Vmax.' of 9.13 munits/mg of protein. Its activity is influenced by the ionic strength of the media and is inhibited by KCN, thiol reagents, NADH and albumin. The enzyme is aerobic, and between 1 and 1.5 mol of O2 are consumed per mol of bilirubin degraded. The products of the reaction include propentdyopents. The hepatic bilirubin oxidase activity of the jaundiced Gunn-rat liver is not significantly different from that of the Sprague-Dawley rat, and it is not induced by beta-naphthoflavone. PMID:3790083

  11. [Cytoprotective effects of bilirubin].

    PubMed

    Vítek, L

    2005-01-01

    Bilirubin, a major product of heme catabolism, belongs to compounds with pleiotropic biologic effects. For a long time bilirubin was considered as a metabolite dangerous for human health, neonatologists know well serious clinical complication of neonatal jaundice called bilirubin encephalopathy. Nevertheless, recent data has demonstrated that bilirubin exhibits potent antioxidant and even anti-inflammatory effects with substantial clinical impacts. The aim of the present study was to summarize present knowledge in this rapidly evolving field and suggest further possible clinical consequences. PMID:15981989

  12. Catalytically active bovine serum amine oxidase bound to fluorescent and magnetically drivable nanoparticles

    PubMed Central

    Sinigaglia, Giulietta; Magro, Massimiliano; Miotto, Giovanni; Cardillo, Sara; Agostinelli, Enzo; Zboril, Radek; Bidollari, Eris; Vianello, Fabio

    2012-01-01

    Novel superparamagnetic surface-active maghemite nanoparticles (SAMNs) characterized by a diameter of 10 ± 2 nm were modified with bovine serum amine oxidase, which used rhodamine B isothiocyanate (RITC) adduct as a fluorescent spacer-arm. A fluorescent and magnetically drivable adduct comprised of bovine serum copper-containing amine oxidase (SAMN–RITC–BSAO) that immobilized on the surface of specifically functionalized magnetic nanoparticles was developed. The multifunctional nanomaterial was characterized using transmission electron microscopy, infrared spectroscopy, mass spectrometry, and activity measurements. The results of this study demonstrated that bare magnetic nanoparticles form stable colloidal suspensions in aqueous solutions. The maximum binding capacity of bovine serum amine oxidase was approximately 6.4 mg g−1 nanoparticles. The immobilization procedure reduced the catalytic activity of the native enzyme to 30% ± 10% and the Michaelis constant was increased by a factor of 2. We suggest that the SAMN–RITC–BSAO complex, characterized by a specific activity of 0.81 IU g−1, could be used in the presence of polyamines to create a fluorescent magnetically drivable H2O2 and aldehydes-producing system. Selective tumor cell destruction is suggested as a potential future application of this system. PMID:22619559

  13. An amperometric cholesterol biosensor based on epoxy resin membrane bound cholesterol oxidase

    PubMed Central

    Pundir, C.S.; Narang, Jagriti; Chauhan, Nidhi; Sharma, Preety; Sharma, Renu

    2012-01-01

    Background & objectives: The use of epoxy resin membrane as a support for immobilization of enzyme has resulted into improved sensitivity and stability of biosensors for uric acid, ascorbic acid and polyphenols. The present work was aimed to prepare an improved amperometric biosensor for determination of serum cholesterol required in the diagnostics and management of certain pathological conditions. Methods: Epoxy resin membrane with immobilized cholesterol oxidase was mounted on the cleaned platinum (Pt) electrode with a parafilm to construct a working electrode. This working electrode along with Ag/AgCl as reference and Ag wire as an auxiliary electrode were connected through a three terminal electrometer to construct a cholesterol biosensor. Results: The sensor showed optimum response within 25 sec at pH 7.0 and 45°C. The linear working range of biosensor was 1.0 to 8.0 mM cholesterol. Km and Imax for cholesterol were 5.0 mM and 9.09 μA, respectively. The biosensor measured serum cholesterol. The minimum detection limit of the sensor was 1.0 mM. The mean analytical recoveries of added cholesterol in serum (2.84 and 4.13 mM) were 91.4±2.8 and 92.3±3.1 per cent (n=6), respectively. Within and between assay coefficient of variation (CV) were <2 and <4 per cent, respectively. Biosensor had a storage life of 6 months at 4°C. Interpretation & conclusions: The use of epoxy resin membrane as a support for immobilization of cholesterol oxidase has resulted into an improved amperometric cholesterol biosensor. The present biosensor had an advantage over the existing biosensors as it worked at comparatively lower potential. PMID:23168704

  14. An antibody that confers plant disease resistance targets a membrane-bound glyoxal oxidase in Fusarium.

    PubMed

    Song, Xiu-Shi; Xing, Shu; Li, He-Ping; Zhang, Jing-Bo; Qu, Bo; Jiang, Jin-He; Fan, Chao; Yang, Peng; Liu, Jin-Long; Hu, Zu-Quan; Xue, Sheng; Liao, Yu-Cai

    2016-05-01

    Plant germplasm resources with natural resistance against globally important toxigenic Fusarium are inadequate. CWP2, a Fusarium genus-specific antibody, confers durable resistance to different Fusarium pathogens that infect cereals and other crops, producing mycotoxins. However, the nature of the CWP2 target is not known. Thus, investigation of the gene coding for the CWP2 antibody target will likely provide critical insights into the mechanism underlying the resistance mediated by this disease-resistance antibody. Immunoblots and mass spectrometry analysis of two-dimensional electrophoresis gels containing cell wall proteins from Fusarium graminearum (Fg) revealed that a glyoxal oxidase (GLX) is the CWP2 antigen. Cellular localization studies showed that GLX is localized to the plasma membrane. This GLX efficiently catalyzes hydrogen peroxide production; this enzymatic activity was specifically inhibited by the CWP2 antibody. GLX-deletion strains of Fg, F. verticillioides (Fv) and F. oxysporum had significantly reduced virulence on plants. The GLX-deletion Fg and Fv strains had markedly reduced mycotoxin accumulation, and the expression of key genes in mycotoxin metabolism was downregulated. This study reveals a single gene-encoded and highly conserved cellular surface antigen that is specifically recognized by the disease-resistance antibody CWP2 and regulates both virulence and mycotoxin biosynthesis in Fusarium species. PMID:26720747

  15. Inherited Disorders of Bilirubin Clearance

    PubMed Central

    Memon, Naureen; Weinberger, Barry I; Hegyi, Thomas; Aleksunes, Lauren M

    2016-01-01

    Inherited disorders of hyperbilirubinemia may be caused by increased bilirubin production or decreased bilirubin clearance. Reduced hepatic bilirubin clearance can be due to defective 1) unconjugated bilirubin uptake and intrahepatic storage, 2) conjugation of glucuronic acid to bilirubin (e.g. Gilbert syndrome, Crigler-Najjar syndrome, Lucey-Driscoll syndrome, breast milk jaundice), 3) bilirubin excretion into bile (Dubin-Johnson syndrome), or 4) conjugated bilirubin re-uptake (Rotor syndrome). In this review, the molecular mechanisms and clinical manifestations of these conditions are described, as well as current approaches to diagnosis and therapy. PMID:26595536

  16. Optical transcutaneous bilirubin detector

    DOEpatents

    Kronberg, James W.

    1993-01-01

    A transcutaneous bilirubin detector comprising a source of light having spectral components absorbable and not absorbable by bilirubin, a handle assembly, electronic circuitry and a fiber optic bundle connecting the assembly to the light source and circuitry. Inside the assembly is a prism that receives the light from one end of the fiber optic bundle and directs it onto the skin and directs the reflected light back into the bundle. The other end of the bundle is trifucated, with one end going to the light source and the other two ends going to circuitry that determines how much light of each kind has been reflected. A relatively greater amount absorbed by the skin from the portion of the spectrum absorbable by bilirubin may indicate the presence of the illness. Preferably, two measurements are made, one on the kneecap and one on the forehead, and compared to determine the presence of bilirubin. To reduce the impact of light absorption by hemoglobin in the blood carried by the skin, pressure is applied with a plunger and spring in the handle assembly, the pressure limited by points of a button slidably carried in the assembly that are perceived by touch when the pressure applied is sufficient.

  17. Optical transcutaneous bilirubin detector

    DOEpatents

    Kronberg, J.W.

    1991-03-04

    This invention consists of a transcutaneous bilirubin detector comprising a source of light having spectral components absorbable and not absorbable by bilirubin, a handle assembly, electronic circuitry and a fiber optic bundle connecting the assembly to the light source and circuitry. Inside the assembly is a prism that receives the light from one end of the fiber optic bundle and directs it onto the skin and directs the reflected light back into the bundle. The other end of the bundle is trifucated, with one end going to the light source and the other two ends going to circuitry that determines how much light of each kind has been reflected. A relatively greater amount absorbed by the skin from the portion of the spectrum absorbable by bilirubin may indicate the presence of the illness. Preferably, two measurements are made, one on the kneecap and one on the forehead, and compared to determine the presence of bilirubin. To reduce the impact of light absorption by hemoglobin in the blood carried by the skin, pressure is applied with a plunger and spring in the handle assembly, the pressure limited by points of a button slidably carried in the assembly that are perceived by touch when the pressure applied is sufficient.

  18. Optical transcutaneous bilirubin detector

    DOEpatents

    Kronberg, J.W.

    1993-11-09

    A transcutaneous bilirubin detector is designed comprising a source of light having spectral components absorbable and not absorbable by bilirubin, a handle assembly, electronic circuitry and a fiber optic bundle connecting the assembly to the light source and circuitry. Inside the assembly is a prism that receives the light from one end of the fiber optic bundle and directs it onto the skin and directs the reflected light back into the bundle. The other end of the bundle is trifucated, with one end going to the light source and the other two ends going to circuitry that determines how much light of each kind has been reflected. A relatively greater amount absorbed by the skin from the portion of the spectrum absorbable by bilirubin may indicate the presence of the illness. Preferably, two measurements are made, one on the kneecap and one on the forehead, and compared to determine the presence of bilirubin. To reduce the impact of light absorption by hemoglobin in the blood carried by the skin, pressure is applied with a plunger and spring in the handle assembly, the pressure limited by points of a button slidably carried in the assembly that are perceived by touch when the pressure applied is sufficient. 6 figures.

  19. Bilirubin measurements in neonates

    NASA Astrophysics Data System (ADS)

    Newman, Gregory J.

    2000-04-01

    Infant Jaundice is a physiologic condition of elevated bilirubin in the tissue that affects nearly 60 percent of all term newborns and virtually 100 percent of premature infants. The high production of bilirubin in the newborn circulatory system and the inability of the immature liver to process and eliminate it case the condition. When the bilirubin levels rise, it starts to deposit in the baby's skin and in the brain. The deposits in the brain can cause neurologic impairment and death. The BiliCheck is a handheld, battery-powered device that measures the level of jaundice non-invasively using BioPhotonics at the point of care. The result is displayed on an LCD screen immediately, so physicians can now make treatment decision without waiting for results to return from the lab. The BiliCheck System has been marketed worldwide since April of 1998 and has received FDA clearance for use in the USA on pre-photo therapy infants in March of 1999.

  20. Destabilization of the Quaternary Structure of Bovine Heart Cytochrome c Oxidase upon Removal of Tightly Bound Cardiolipin.

    PubMed

    Sedlák, Erik; Robinson, Neal C

    2015-09-15

    The quaternary structural stability of cardiolipin-containing (CcO(CL+)) versus CL-free cytochrome c oxidase (CcO(CL-)) was compared using structural perturbants as probes. Exposure to increasing concentrations of urea or guanidinium chloride causes sequential dissociation of five subunits from CcO(CL+) in the order VIa and VIb, followed by III and VIIa, and ultimately Vb. Removal of CL from CcO destabilizes the association of each of these five subunits with the core of CcO. Subunits VIa and VIb spontaneously dissociate from CcO(CL-) even in the absence of denaturant and are no longer present after purification of the CL-free 11-subunit complex by ion exchange chromatography. The other 11 subunits remain associated in a partially active complex, but the association of subunits III, VIIa, and Vb is weakened; i.e., the midpoints for the subunit dissociation curves are each shifted to a lower perturbant concentration (lower by 1.1-1.7 M urea; lower by 0.3-0.4 M GdmCl). This corresponds to a decrease of ∼9 kJ in the Gibbs free association energy for each of these subunits when CL is removed from CcO. With either CcO(CL+) or CcO(CL-), loss of enzymatic activity occurs coincident with dissociation of subunits III and VIIa. The loss of activity is irreversible, and reactivation of CcO(CL-) by exogenous CL occurs only if both subunits remain associated with the core of CcO. Inclusion of sulfate anions stabilizes the association of VIIa more than III, resulting in a slight separation of the urea-induced dissociation curves. In this case, activity loss correlates much better with dissociation of subunit VIIa than III. We conclude that (1) bound cardiolipin is an important stabilizing factor in the quaternary structure of CcO and (2) association of subunit VIIa (possibly together with subunit III) is critical for functional CL binding and full electron-transfer activity of CcO. PMID:26284624

  1. Evolution of protein bound Maillard reaction end-products and free Amadori compounds in low lactose milk in presence of fructosamine oxidase I.

    PubMed

    Troise, Antonio Dario; Buonanno, Martina; Fiore, Alberto; Monti, Simona Maria; Fogliano, Vincenzo

    2016-12-01

    Thermal treatments and storage influence milk quality, particularly in low lactose milk as the higher concentration of reducing sugars can lead to the increased formation of the Maillard reaction products (MRPs). The control of the Amadori products (APs) formation is the key step to mitigate the Maillard reaction (MR) in milk. The use of fructosamine oxidases, (Faox) provided promising results. In this paper, the effects of Faox I were evaluated by monitoring the concentration of free and bound MRPs in low lactose milk during shelf life. Results showed that the enzyme reduced the formation of protein-bound MRPs down to 79% after six days at 37°C. Faox I lowered the glycation of almost all the free amino acids resulting effective on basic and polar amino acids. Data here reported corroborate previous findings on the potentiality of Faox enzymes in controlling the early stage of the MR in foods. PMID:27374589

  2. The Biological Effects of Bilirubin Photoisomers.

    PubMed

    Jasprova, Jana; Dal Ben, Matteo; Vianello, Eleonora; Goncharova, Iryna; Urbanova, Marie; Vyroubalova, Karolina; Gazzin, Silvia; Tiribelli, Claudio; Sticha, Martin; Cerna, Marcela; Vitek, Libor

    2016-01-01

    Although phototherapy was introduced as early as 1950's, the potential biological effects of bilirubin photoisomers (PI) generated during phototherapy remain unclear. The aim of our study was to isolate bilirubin PI in their pure forms and to assess their biological effects in vitro. The three major bilirubin PI (ZE- and EZ-bilirubin and Z-lumirubin) were prepared by photo-irradiation of unconjugated bilirubin. The individual photoproducts were chromatographically separated (TLC, HPLC), and their identities verified by mass spectrometry. The role of Z-lumirubin (the principle bilirubin PI) on the dissociation of bilirubin from albumin was tested by several methods: peroxidase, fluorescence quenching, and circular dichroism. The biological effects of major bilirubin PI (cell viability, expression of selected genes, cell cycle progression) were tested on the SH-SY5Y human neuroblastoma cell line. Lumirubin was found to have a binding site on human serum albumin, in the subdomain IB (or at a close distance to it); and thus, different from that of bilirubin. Its binding constant to albumin was much lower when compared with bilirubin, and lumirubin did not affect the level of unbound bilirubin (Bf). Compared to unconjugated bilirubin, bilirubin PI did not have any effect on either SH-SY5Y cell viability, the expression of genes involved in bilirubin metabolism or cell cycle progression, nor in modulation of the cell cycle phase. The principle bilirubin PI do not interfere with bilirubin albumin binding, and do not exert any toxic effect on human neuroblastoma cells. PMID:26829016

  3. Bilirubin Binding Capacity in the Preterm Neonate.

    PubMed

    Amin, Sanjiv B

    2016-06-01

    Total serum/plasma bilirubin (TB), the biochemical measure currently used to evaluate and manage hyperbilirubinemia, is not a useful predictor of bilirubin-induced neurotoxicity in premature infants. Altered bilirubin-albumin binding in premature infants limits the usefulness of TB in premature infants. In this article, bilirubin-albumin binding, a modifying factor for bilirubin-induced neurotoxicity, in premature infants is reviewed. PMID:27235205

  4. Ligand-Bound GeneSwitch Causes Developmental Aberrations in Drosophila that Are Alleviated by the Alternative Oxidase

    PubMed Central

    Andjelković, Ana; Kemppainen, Kia K.; Jacobs, Howard T.

    2016-01-01

    Culture of Drosophila expressing the steroid-dependent GeneSwitch transcriptional activator under the control of the ubiquitous α-tubulin promoter was found to produce extensive pupal lethality, as well as a range of dysmorphic adult phenotypes, in the presence of high concentrations of the inducing drug RU486. Prominent among these was cleft thorax, seen previously in flies bearing mutant alleles of the nuclear receptor Ultraspiracle and many other mutants, as well as notched wings, leg malformations, and bristle abnormalities. Neither the α-tubulin-GeneSwitch driver nor the inducing drug on their own produced any of these effects. A second GeneSwitch driver, under the control of the daughterless promoter, which gave much lower and more tissue-restricted transgene expression, exhibited only mild bristle abnormalities in the presence of high levels of RU486. Coexpression of the alternative oxidase (AOX) from Ciona intestinalis produced a substantial shift in the developmental outcome toward a wild-type phenotype, which was dependent on the AOX expression level. Neither an enzymatically inactivated variant of AOX, nor GFP, or the alternative NADH dehydrogenase Ndi1 from yeast gave any such rescue. Users of the GeneSwitch system should be aware of the potential confounding effects of its application in developmental studies. PMID:27412986

  5. Photoacoustic microscopy of bilirubin in tissue phantoms

    NASA Astrophysics Data System (ADS)

    Zhou, Yong; Zhang, Chi; Yao, Da-Kang; Wang, Lihong V.

    2012-12-01

    Determining both bilirubin's concentration and its spatial distribution are important in disease diagnosis. Here, for the first time, we applied quantitative multiwavelength photoacoustic microscopy (PAM) to detect bilirubin concentration and distribution simultaneously. By measuring tissue-mimicking phantoms with different bilirubin concentrations, we showed that the root-mean-square error of prediction has reached 0.52 and 0.83 mg/dL for pure bilirubin and for blood-mixed bilirubin detection (with 100% oxygen saturation), respectively. We further demonstrated the capability of the PAM system to image bilirubin distribution both with and without blood. Finally, by underlaying bilirubin phantoms with mouse skins, we showed that bilirubin can be imaged with consistent accuracy down to >400 μm in depth. Our results show that PAM has potential for noninvasive bilirubin monitoring in vivo, as well as for further clinical applications.

  6. Inhibition of NADPH oxidase 1 activity and blocking the binding of cytosolic and membrane-bound proteins by honokiol inhibit migratory potential of melanoma cells.

    PubMed

    Prasad, Ram; Kappes, John C; Katiyar, Santosh K

    2016-02-16

    Overexpression of NADPH oxidase 1 (Nox1) in melanoma cells is often associated with increased migration/metastasis rate. To develop effective treatment options, we have examined the effect of honokiol, a phytochemical from Magnolia plant, on the migratory potential of human melanoma cell lines (A375, Hs294t, SK-Mel119 and SK-Mel28) and assessed whether Nox1 is the target. Using an in vitro cell migration assay, we observed that treatment of different melanoma cell lines with honokiol for 24 h resulted in a dose-dependent inhibition of cell migration that was associated with reduction in Nox1 expression and reduced levels of oxidative stress. Treatment of cells with N-acetyl-L-cysteine, an anti-oxidant, also inhibited the migration of melanoma cells. Treatment of cells with diphenyleneiodonium chloride, an inhibitor of Nox1, significantly decreased the migration ability of Hs294t and SK-Mel28 cells. Further, we examined the effect of honokiol on the levels of core proteins (p22phox and p47phox) of the NADPH oxidase complex. Treatment of Hs294t and SK-Mel28 cells with honokiol resulted in accumulation of the cytosolic p47phox protein and decreased levels of the membrane-bound p22phox protein, thus blocking their interaction and inhibiting Nox1 activation. Our in vivo bioluminescence imaging data indicate that oral administration of honokiol inhibited the migration/extravasation and growth of intravenously injected melanoma cells in internal body organs, such as liver, lung and kidney in nude mice, and that this was associated with an inhibitory effect on Nox1 activity in these internal organs/tissues. PMID:26760964

  7. Inhibition of NADPH oxidase 1 activity and blocking the binding of cytosolic and membrane-bound proteins by honokiol inhibit migratory potential of melanoma cells

    PubMed Central

    Prasad, Ram; Kappes, John C.; Katiyar, Santosh K.

    2016-01-01

    Overexpression of NADPH oxidase 1 (Nox1) in melanoma cells is often associated with increased migration/metastasis rate. To develop effective treatment options, we have examined the effect of honokiol, a phytochemical from Magnolia plant, on the migratory potential of human melanoma cell lines (A375, Hs294t, SK-Mel119 and SK-Mel28) and assessed whether Nox1 is the target. Using an in vitro cell migration assay, we observed that treatment of different melanoma cell lines with honokiol for 24 h resulted in a dose-dependent inhibition of cell migration that was associated with reduction in Nox1 expression and reduced levels of oxidative stress. Treatment of cells with N-acetyl-L-cysteine, an anti-oxidant, also inhibited the migration of melanoma cells. Treatment of cells with diphenyleneiodonium chloride, an inhibitor of Nox1, significantly decreased the migration ability of Hs294t and SK-Mel28 cells. Further, we examined the effect of honokiol on the levels of core proteins (p22phox and p47phox) of the NADPH oxidase complex. Treatment of Hs294t and SK-Mel28 cells with honokiol resulted in accumulation of the cytosolic p47phox protein and decreased levels of the membrane-bound p22phox protein, thus blocking their interaction and inhibiting Nox1 activation. Our in vivo bioluminescence imaging data indicate that oral administration of honokiol inhibited the migration/extravasation and growth of intravenously injected melanoma cells in internal body organs, such as liver, lung and kidney in nude mice, and that this was associated with an inhibitory effect on Nox1 activity in these internal organs/tissues. PMID:26760964

  8. Comparison in different species of biliary bilirubin-IX alpha conjugates with the activities of hepatic and renal bilirubin-IX alpha-uridine diphosphate glycosyltransferases.

    PubMed Central

    Fevery, J; Van de Vijver, M; Michiels, R; Heirwegh, K P

    1977-01-01

    The bilrubin-IXalpha conjugates in bile and the activities of bilirubin-IX alpha--UDP-glycosyltransferases in liver and kidney were determined for ten species of mammals and for the chicken. 1. In the mammalian species, bilirubin-IX alpha glucuronide was the predominant bile pigment. Excretion of neutral glycosides was unimportant, except in the cat, the mouse, the rabbit and the dog, where glucose and xylose represented 12--41% of total conjugating groups bound to bilirubin-IX alpha. In chicken bile, glucoside and glucuronide conjugates were of equal importance. They probably represent only a small fraction of the total bile pigment. 2. The transferase activities in liver showed pronounced species variation. This was also apparent with regard to activation by digitonin, pH optimum and relative activities of transferases acting on either UDP-glucuronic acid or neutral UDP-sugars. 3. Man, the dog, the cat and the rat excrete bilirubin-IX alpha largely as diconjugated derivatives. In general, diconjugated bilirubin-IX alpha could also be synthesized in vitro with liver homogenate, bilirubin-IX alpha and UDP-sugar. In contrast, for the other species examined, bilirubin pigments consisted predominantly of monoconjugated bilirubin-IX alpha. Synthesis in vitro with UDP-glucuronic acid, UDP-glucose or UDP-xylose as the sugar donor led exclusively to the formation of monoconjugated bilirubin-IX alpha. 4. The transferase activities in the kidney were restricted to the cortex and were important only for the rat and the dog. No activity at all could be detected for several species, including man. 5. Comparison of the transferase activities in liver with reported values of the maximal rate of excretion in bile suggests a close linkage between conjugation and biliary secretion of bilirubin-IX alpha. PMID:407905

  9. The Biological Effects of Bilirubin Photoisomers

    PubMed Central

    Jasprova, Jana; Dal Ben, Matteo; Vianello, Eleonora; Goncharova, Iryna; Urbanova, Marie; Vyroubalova, Karolina; Gazzin, Silvia; Tiribelli, Claudio; Sticha, Martin; Cerna, Marcela; Vitek, Libor

    2016-01-01

    Although phototherapy was introduced as early as 1950’s, the potential biological effects of bilirubin photoisomers (PI) generated during phototherapy remain unclear. The aim of our study was to isolate bilirubin PI in their pure forms and to assess their biological effects in vitro. The three major bilirubin PI (ZE- and EZ-bilirubin and Z-lumirubin) were prepared by photo-irradiation of unconjugated bilirubin. The individual photoproducts were chromatographically separated (TLC, HPLC), and their identities verified by mass spectrometry. The role of Z-lumirubin (the principle bilirubin PI) on the dissociation of bilirubin from albumin was tested by several methods: peroxidase, fluorescence quenching, and circular dichroism. The biological effects of major bilirubin PI (cell viability, expression of selected genes, cell cycle progression) were tested on the SH-SY5Y human neuroblastoma cell line. Lumirubin was found to have a binding site on human serum albumin, in the subdomain IB (or at a close distance to it); and thus, different from that of bilirubin. Its binding constant to albumin was much lower when compared with bilirubin, and lumirubin did not affect the level of unbound bilirubin (Bf). Compared to unconjugated bilirubin, bilirubin PI did not have any effect on either SH-SY5Y cell viability, the expression of genes involved in bilirubin metabolism or cell cycle progression, nor in modulation of the cell cycle phase. The principle bilirubin PI do not interfere with bilirubin albumin binding, and do not exert any toxic effect on human neuroblastoma cells. PMID:26829016

  10. Structure of the Acinetobacter baumannii Dithiol Oxidase DsbA Bound to Elongation Factor EF-Tu Reveals a Novel Protein Interaction Site

    PubMed Central

    Premkumar, Lakshmanane; Kurth, Fabian; Duprez, Wilko; Grøftehauge, Morten K.; King, Gordon J.; Halili, Maria A.; Heras, Begoña; Martin, Jennifer L.

    2014-01-01

    The multidrug resistant bacterium Acinetobacter baumannii is a significant cause of nosocomial infection. Biofilm formation, that requires both disulfide bond forming and chaperone-usher pathways, is a major virulence trait in this bacterium. Our biochemical characterizations show that the periplasmic A. baumannii DsbA (AbDsbA) enzyme has an oxidizing redox potential and dithiol oxidase activity. We found an unexpected non-covalent interaction between AbDsbA and the highly conserved prokaryotic elongation factor, EF-Tu. EF-Tu is a cytoplasmic protein but has been localized extracellularly in many bacterial pathogens. The crystal structure of this complex revealed that the EF-Tu switch I region binds to the non-catalytic surface of AbDsbA. Although the physiological and pathological significance of a DsbA/EF-Tu association is unknown, peptides derived from the EF-Tu switch I region bound to AbDsbA with submicromolar affinity. We also identified a seven-residue DsbB-derived peptide that bound to AbDsbA with low micromolar affinity. Further characterization confirmed that the EF-Tu- and DsbB-derived peptides bind at two distinct sites. These data point to the possibility that the non-catalytic surface of DsbA is a potential substrate or regulatory protein interaction site. The two peptides identified in this work together with the newly characterized interaction site provide a novel starting point for inhibitor design targeting AbDsbA. PMID:24860094

  11. Bilirubin chemistry and metabolism; harmful and protective aspects.

    PubMed

    Vítek, Libor; Ostrow, J Donald

    2009-01-01

    Unconjugated bilirubin (UCB), the principal mammalian bile pigment, is the end product of heme catabolism. Both belong to the superfamily of tetrapyrrolic compounds that serve multiple biological functions in animals and plants. Its six internal hydrogen bonds give UCB a unique structure responsible for its physico-chemical properties and biological effects. Like many weakly-polar, poorly-soluble compounds, UCB is transported in blood tightly bound to albumin, with less than 0.01% of total bilirubin circulating in an unbound form (free bilirubin, Bf). This fraction governs the diffusion of UCB into tissues, and therefore Bf is responsible for both its beneficial and toxic effects on cells. Although, UCB was long thought to be a non-functional waste product, recent studies have shown that the antioxidant effects of mildly elevated serum bilirubin levels, as well as activation of heme oxygenase, may protect against diseases associated with oxidative stress, such as atherosclerosis. By contrast, markedly elevated serum UCB levels may cause severe neurological damage, especially in neonates. The regulation of cellular UCB content, by its conjugation, oxidation, and export, are, therefore of paramount importance to cellular health. PMID:19754364

  12. Bilirubin Binding to PPARα Inhibits Lipid Accumulation.

    PubMed

    Stec, David E; John, Kezia; Trabbic, Christopher J; Luniwal, Amarjit; Hankins, Michael W; Baum, Justin; Hinds, Terry D

    2016-01-01

    Numerous clinical and population studies have demonstrated that increased serum bilirubin levels protect against cardiovascular and metabolic diseases such as obesity and diabetes. Bilirubin is a potent antioxidant, and the beneficial actions of moderate increases in plasma bilirubin have been thought to be due to the antioxidant effects of this bile pigment. In the present study, we found that bilirubin has a new function as a ligand for PPARα. We show that bilirubin can bind directly to PPARα and increase transcriptional activity. When we compared biliverdin, the precursor to bilirubin, on PPARα transcriptional activation to known PPARα ligands, WY 14,643 and fenofibrate, it showed that fenofibrate and biliverdin have similar activation properties. Treatment of 3T3-L1 adipocytes with biliverdin suppressed lipid accumulation and upregulated PPARα target genes. We treated wild-type and PPARα KO mice on a high fat diet with fenofibrate or bilirubin for seven days and found that both signal through PPARα dependent mechanisms. Furthermore, the effect of bilirubin on lowering glucose and reducing body fat percentage was blunted in PPARα KO mice. These data demonstrate a new function for bilirubin as an agonist of PPARα, which mediates the protection from adiposity afforded by moderate increases in bilirubin. PMID:27071062

  13. Bilirubin Binding to PPARα Inhibits Lipid Accumulation

    PubMed Central

    Stec, David E.; John, Kezia; Trabbic, Christopher J.; Luniwal, Amarjit; Hankins, Michael W.; Baum, Justin

    2016-01-01

    Numerous clinical and population studies have demonstrated that increased serum bilirubin levels protect against cardiovascular and metabolic diseases such as obesity and diabetes. Bilirubin is a potent antioxidant, and the beneficial actions of moderate increases in plasma bilirubin have been thought to be due to the antioxidant effects of this bile pigment. In the present study, we found that bilirubin has a new function as a ligand for PPARα. We show that bilirubin can bind directly to PPARα and increase transcriptional activity. When we compared biliverdin, the precursor to bilirubin, on PPARα transcriptional activation to known PPARα ligands, WY 14,643 and fenofibrate, it showed that fenofibrate and biliverdin have similar activation properties. Treatment of 3T3-L1 adipocytes with biliverdin suppressed lipid accumulation and upregulated PPARα target genes. We treated wild-type and PPARα KO mice on a high fat diet with fenofibrate or bilirubin for seven days and found that both signal through PPARα dependent mechanisms. Furthermore, the effect of bilirubin on lowering glucose and reducing body fat percentage was blunted in PPARα KO mice. These data demonstrate a new function for bilirubin as an agonist of PPARα, which mediates the protection from adiposity afforded by moderate increases in bilirubin. PMID:27071062

  14. Bilirubin and Its Carbohydrate Conjugates

    NASA Astrophysics Data System (ADS)

    Blanckaert, Norbert J. C.

    In mammals, the open tetrapyrrole bilirubin (structure 2, Fig. 1) is the principal degradation product of iron-protoporphyrin-IX (heme). The latter molecule is a tetrapyrrolic macrocycle and plays a critical role in aerobic metabolism by reversibly binding oxygen in hemoglobin and myoglobin, and by serving as the active site in oxidation reactions catalyzed by hemoprotein enzymes. Important cyclic tetrapyrroles in nature related to heme are chlorophylls, which contain magnesium and are derived from protoporphyrin-IX, and vitamin B12, a corrinoid derived from uroporphyrinogen-III.

  15. Clinical assessment of bilirubin-induced neurotoxicity in premature infants.

    PubMed

    Amin, Sanjiv B

    2004-10-01

    The clinical assessment of bilirubin-induced neurotoxicity in premature infants remains difficult in the absence of a gestational age-specific total or free (unbound) bilirubin level that predicts bilirubin-induced neurotoxicity. Because the total serum bilirubin concentration is an unreliable predictor of bilirubin-induced neurotoxicity in premature infants, alternative mean for predicting bilirubin-induced neurotoxicity in jaundiced preterm newborns is needed. Over the last few years, we have witnessed substantial gain in our knowledge involving usefulness of bilirubin-binding variables (total bilirubin, free bilirubin, bilirubin:albumin molar ratio) for clinical assessment of bilirubin-induced neurotoxicity in preterm infants. The knowledge gained has provided impetus for more clinical studies that are geared toward confirming the usefulness of free bilirubin as a predictor of bilirubin-induced neurotoxicity and identifying the gestational age-specific free bilirubin level that may increase the risk of bilirubin-induced neurotoxicity in premature infants. The paper has attempted to provide an overview of bilirubin-induced auditory toxicity along with the existing clinical evidence in favor of free bilirubin assay and usefulness of auditory brainstem evoked response for evaluation of bilirubin-induced neurotoxicity in premature infants. In addition, the author has described findings that suggest an association of apnea, a clinical manifestation, with acute bilirubin encephalopathy in premature infants. PMID:15686265

  16. An amperometric bilirubin biosensor based on a conductive poly-terthiophene-Mn(II) complex.

    PubMed

    Rahman, Md Aminur; Lee, Kyung-Sun; Park, Deog-Su; Won, Mi-Sook; Shim, Yoon-Bo

    2008-01-18

    An amperometric bilirubin biosensor was fabricated by complexing the Mn(II) ion with a conducting polymer and the final biosensor surface was coated with a thin polyethyleneimine (PEI) film containing an enzyme, ascorbate oxidase (AsOx). The complexation between poly-5,2'-5',2''-terthiophene-3-carboxylic acid (PolyTTCA) and Mn(II) through the formation of Mn-O bond was confirmed by XPS. The PolyTTCA-Mn(II) complex was also characterized using cyclic voltammetry. The PolyTTCA-Mn(II)/PEI-AsOx biosensor specifically detect bilirubin through the mediated electron transfer by the Mn(II) ion. To optimize the experimental condition, various experimental parameters such as pH, temperature, and applied potential were examined. A linear calibration plot for bilirubin was obtained between 0.1 microM and 50 microM with the detection limit of 40+/-3.8 nM. Interferences from other biological compounds, especially ascorbate and dopamine were efficiently minimized by coating the biosensor surface with PEI-AsOx. The bilirubin sensor exhibited good stability and fast response time (<5s). The applicability of this bilirubin sensor was tested in a human serum sample. PMID:17964773

  17. Interaction of bilirubin and indocyanine green with the binding and conjugation of sulfobromophthalein by rat liver cytosol proteins.

    PubMed

    Davis, D R; Yeary, R A

    1980-02-01

    The interaction of bilirubin and indocyanine green with sulfobromophthalein (BSP) binding and conjugation by rat liver cytosol proteins was studied. BSP bound to cytosol proteins X, ligandin and Z and the BSP-glutathione conjugate were isolated by sephadex gel chromatography. Neither bilirubin nor indocyanine green affected the binding of BSP to ligandin and Z protein. However, indocyanine green did significantly reduce BSP conjugation in both in vitro and in vivo experiments. Diethyl maleate significantly reduced liver glutathione levels and BSP conjugation. It is suggested that indocyanine gree competitively binds at the ligandin catalytic site whereas the primary binding site for bilirubin is probably a noncatalytic site. PMID:7367753

  18. Direct electrical communication between chemically modified enzymes and metal electrodes. 1. Electron transfer from glucose oxidase to metal electrodes via electron relays, bound covalently to the enzyme

    SciTech Connect

    Degani, Y.; Heller, A.

    1987-03-12

    Glucose-reduced glucose oxidase does not directly transfer electrons to conventional electrodes because the distance between its redox centers and the electrode surface exceeds, even on closest approach, the distance across which electrons are transferred at sufficient rates. Therefore, electrical communication between the redox centers of this enzyme and electrodes required either the presence, and diffusion to and from the enzyme's redox center, of O/sub 2/ and H/sub 2/O/sub 2/, or the presence of members of a redox couple, or the use of special electrodes like TTF/TCNQ. They show here that direct electrical communication between the redox center of a large enzyme molecule and a simple metal electrode can be established through chemical modification of the enzyme. When a sufficient number of electron-relaying centers are attached through covalent bonding to the protein of glucose oxidase, electrons are transferred from the enzyme's redox centers to relays that are closer to the periphery of the enzyme. Because some of the relays are located sufficiently close to the enzyme's surface, electrons are transferred at practical rates to the electrode. As a result, a glucose-concentration-dependent current flows in an electrochemical cell made with conventional electrodes when the electrolytic solution contains the relay-modified enzyme. Such a current does not flow when the solution contains the natural enzyme. Specifically, electrical communication is established between the FAD/FADH/sub 2/ centers of glucose oxidase and gold, platinum, or carbon electrodes through the covalent bonding of an average of 12 molecules of ferrocenecarboxylic acid per glucose oxidase molecule.

  19. Determination of bilirubin glucuronide and assay of glucuronyltransferase with bilirubin as acceptor

    PubMed Central

    Van Roy, F. P.; Heirwegh, K. P. M.

    1968-01-01

    1. Conjugated bilirubin is conveniently determined by coupling with the diazonium salt of ethyl anthranilate. 2. This method has been used in the development of assays for UDP-glucuronyltransferase (EC 2.4.1.17), with bilirubin as substrate, in rat liver homogenates, microsomal preparations and partly purified fractions. 3. Chromatographic analysis suggests that bilirubin monoglucuronide is the product of the enzyme systems studied. PMID:5660631

  20. Intermolecular interactions in the bilirubin-cholate-silica system

    NASA Astrophysics Data System (ADS)

    Vlasova, N. N.; Golovkova, L. P.; Severinovskaya, O. V.

    2007-06-01

    Bilirubin-cholate interactions in aqueous solutions were studied. The constants of binding of bilirubin with taurocholate dimers and taurodeoxycholate trimers were calculated. The adsorption of bilirubin and cholates on the surface of highly dispersed silica was studied. It was shown that taurine-conjugated cholates are poorly adsorbed from micellar solutions on the silica surface, the specific amount of bilirubin adsorbed decreases with increasing concentration of cholates in the solution, the affinity of free bilirubin for the silica surface is independent of the nature of the cholic acid, and that the affinity of cholate-bilirubin complexes for the silica surface is lower than the affinity of free bilirubin.

  1. Universal bilirubin screening for severe neonatal hyperbilirubinemia.

    PubMed

    Bhutani, V K; Vilms, R J; Hamerman-Johnson, L

    2010-10-01

    To reduce the incidence of severe neonatal hyperbilirubinemia affecting newborns with jaundice in the United States and to prevent kernicterus, there is a need to implement proven prevention strategies for severe neonatal hyperbilirubinemia as recommended in the 2004 American Academy of Pediatrics Guidelines for newborns >35 weeks gestational age. The purpose of universal predischarge bilirubin screening is to identify infants with bilirubin levels >75th percentile for age in hours and track those with rapid rates of bilirubin rise (>0.2 mg per 100 ml per h). Early identification has been reported to predict severe hyperbilirubinemia and allow for evidence-based targeted interventions. A systems approach is likely to reduce the preventable causes of acute bilirubin encephalopathy. To do so, highest priority should be given to (i) designating extreme hyperbilirubinemia (total serum bilirubin >427 μmol l(-1) or >25 mg per 100 ml) as a reportable condition by laboratories and health-care providers through public health mandates; (ii) implementation of Joint Commission's Sentinel Report for kernicterus; (iii) nursing outreach to communities for education of prospective parents; (iv) development of clinical pathways to monitor, evaluate and track infants with extreme hyperbilirubinemia; and (v) societal awareness. These efforts should be monitored by a state and national surveillance system in order to critically improve the timeliness and completeness of notifications and to allow evaluation and interventions at the policy and individual family level. PMID:20877410

  2. PPARα: A Master Regulator of Bilirubin Homeostasis.

    PubMed

    Bigo, Cyril; Kaeding, Jenny; El Husseini, Diala; Rudkowska, Iwona; Verreault, Mélanie; Vohl, Marie Claude; Barbier, Olivier

    2014-01-01

    Hypolipidemic fibrates activate the peroxisome proliferator-activated receptor (PPAR) α to modulate lipid oxidation and metabolism. The present study aimed at evaluating how 3 PPARα agonists, namely, fenofibrate, gemfibrozil, and Wy14,643, affect bilirubin synthesis and metabolism. Human umbilical vein epithelial cells (HUVEC) and coronary artery smooth muscle cells (CASMC) were cultured in the absence or presence of the 3 activators, and mRNA, protein, and/or activity levels of the bilirubin synthesizing heme oxygenase- (HO-) 1 and biliverdin reductase (BVR) enzymes were determined. Human hepatocytes (HH) and HepG2 cells sustained similar treatments, except that the expression of the bilirubin conjugating UDP-glucuronosyltransferase (UGT) 1A1 enzyme and multidrug resistance-associated protein (MRP) 2 transporter was analyzed. In HUVECs, gemfibrozil, fenofibrate, and Wy14,643 upregulated HO-1 mRNA expression without affecting BVR. Wy14,643 and fenofibrate also caused HO-1 protein accumulation, while gemfibrozil and fenofibrate favored the secretion of bilirubin in cell media. Similar positive regulations were also observed with the 3 PPARα ligands in CASMCs where HO-1 mRNA and protein levels were increased. In HH and HepG2 cells, both UGT1A1 and MRP2 transcripts were also accumulating. These observations indicate that PPARα ligands activate bilirubin synthesis in vascular cells and metabolism in liver cells. The clinical implications of these regulatory events are discussed. PMID:25147562

  3. Initial photochemistry of bilirubin probed by femtosecond spectroscopy.

    PubMed

    Zietz, Burkhard; Gillbro, Tomas

    2007-10-18

    Bilirubin is a breakdown product from heme catabolism, and reduced excretion of bilirubin can lead to jaundice. Phototherapy is the most common treatment for neonatal jaundice, a condition frequently encountered in newborn infants. Knowledge of the photochemistry of bilirubin, which is dominated by (ultra)fast components, is necessary for the profound understanding of the processes in phototherapy. Here, we report results from femtosecond fluorescence upconversion measurements on bilirubin and half-bilirubin model compounds, as well as pump-probe absorption measurements on bilirubin. A fast component of ca. 120 fs in the multiexponential fluorescence decay, being only visible in the bilirubin molecule, is interpreted as exciton localization within the molecular halves. The slower components of several hundreds of femtoseconds and a few picoseconds, occurring in bilirubin and the half-bilirubin model, are interpreted as relaxation to a (twisted) intermediate, which decays further with ca. 15 ps to the ground state. PMID:17927274

  4. Crystal Structures of Multicopper Oxidase CueO Bound to Copper(I) and Silver(I): Functional Role of a Methonine-Rich Sequence

    SciTech Connect

    Singh, Satish K.; Roberts, Sue A.; McDevitt, Sylvia F.; Weichsel, Andrzej; Wildner, Guenter F.; Grass, Gregor B.; Rensing, Christopher; Montfort, William R.

    2011-10-24

    The multicopper oxidase CueO oxidizes toxic Cu(I) and is required for copper homeostasis in Escherichia coli. Like many proteins involved in copper homeostasis, CueO has a methionine-rich segment that is thought to be critical for copper handling. How such segments function is poorly understood. Here, we report the crystal structure of CueO at 1.1 {angstrom} with the 45-residue methionine-rich segment fully resolved, revealing an N-terminal helical segment with methionine residues juxtaposed for Cu(I) ligation and a C-terminal highly mobile segment rich in methionine and histidine residues. We also report structures of CueO with a C500S mutation, which leads to loss of the T1 copper, and CueO with six methionines changed to serine. Soaking C500S CueO crystals with Cu(I), or wild-type CueO crystals with Ag(I), leads to occupancy of three sites, the previously identified substrate-binding site and two new sites along the methionine-rich helix, involving methionines 358, 362, 368, and 376. Mutation of these residues leads to a {approx}4-fold reduction in kcat for Cu(I) oxidation. Ag(I), which often appears with copper in nature, strongly inhibits CueO oxidase activities in vitro and compromises copper tolerance in vivo, particularly in the absence of the complementary copper efflux cus system. Together, these studies demonstrate a role for the methionine-rich insert of CueO in the binding and oxidation of Cu(I) and highlight the interplay among cue and cus systems in copper and silver homeostasis.

  5. Alteration of serum bilirubin level in schizophrenia.

    PubMed

    Semnani, Yousef; Nazemi, Farzad; Azariyam, Aileen; Ardakani, Mohammad Javad Ehsani

    2010-11-01

    Abstract Objective. Alteration of serum bilirubin level in acute episodes of psychosis in patients with schizophrenia has been reported but the pattern of this alteration is controversial. Methods. Patients diagnosed as schizophrenia (162, group S) or bipolar disorder (155, group B) entered the study. The control group consisted of 95 patients admitted to cardiac care unit who had no personal or family history of major psychiatric disorders. Pre- and post-admission levels of bilirubin were measured and compared within and between the groups. Patients were examined to exclude all other causes of hyperbilirubinemia. Group S and B participants were also evaluated using positive and negative syndrome subscale (PANSS) both at admission and discharge. Results. The mean admission bilirubin levels of all the groups were in the normal range (significantly higher in group S than groups B and C) and were affected by the score of general psychopathology subscale rather than the scores of positive symptoms subscale. Conclusions. Although bilirubin decreased in all three groups at discharge, the rate of decrease was significantly higher in group S. The reason for this is not clear and needs further study. PMID:24917437

  6. Mechanisms of bilirubin toxicity: clinical implications.

    PubMed

    Hansen, Thor Willy Ruud

    2002-12-01

    The basic mechanism of kernicterus and bilirubin encephalopathy has not been unequivocally determined. Much knowledge has been gained about phenomena that contribute to bilirubin neurotoxicity, and this knowledge has implications for clinical practice. Conditions that impact on blood-brain barrier function, increase brain blood flow, or impact on bilirubin metabolism, including its transport in serum, should be avoided, if possible. Such conditions include drugs and drug stabilizers that compete with bilirubin binding to albumin, or that inhibit P-glycoprotein in the blood-brain barrier, prematurity/immaturity, and clinically significant illness in the infant that involves hemolysis, respiratory and metabolic acidosis, infection, asphyxia, hypoxia and (perhaps) hyperoxia, and hyperosmolality. If these conditions are not avoidable then there should be a more aggressive approach to the treatment of hyperbilirubinemia. The limits of tolerance for hyperbilirubinemia varies among neonates and there are no tools to determine with certainty when a particular infant is approaching the danger zone. Neurological symptoms in a jaundiced infant require extreme vigilance, and, in most cases, immediate intervention. PMID:12516745

  7. Can Excess Bilirubin Levels Cause Learning Difficulties?

    ERIC Educational Resources Information Center

    Pretorius, E.; Naude, H.; Becker, P. J.

    2002-01-01

    Examined learning problems in South African sample of 7- to 14-year-olds whose mothers reported excessively high infant bilirubin shortly after the child's birth. Found that this sample had lowered verbal ability with the majority also showing impaired short-term and long-term memory. Findings suggested that impaired formation of astrocytes…

  8. Bilirubin is an Endogenous Antioxidant in Human Vascular Endothelial Cells.

    PubMed

    Ziberna, Lovro; Martelanc, Mitja; Franko, Mladen; Passamonti, Sabina

    2016-01-01

    Bilirubin is a standard serum biomarker of liver function. Inexplicably, it is inversely correlated with cardiovascular disease risk. Given the role of endothelial dysfunction in originating cardiovascular diseases, direct analysis of bilirubin in the vascular endothelium would shed light on these relationships. Hence, we used high-performance liquid chromatography coupled with thermal lens spectrometric detection and diode array detection for the determination of endogenous cellular IXα-bilirubin. To confirm the isomer IXα-bilirubin, we used ultra-performance liquid chromatography coupled with a high-resolution mass spectrometer using an electrospray ionization source, as well as tandem mass spectrometric detection. We measured bilirubin in both arterial and venous rat endothelium (0.9-1.5 pmol mg(-1) protein). In the human endothelial Ea.hy926 cell line, we demonstrated that intracellular bilirubin (3-5 pmol mg(-1) protein) could be modulated by either extracellular bilirubin uptake, or by up-regulation of heme oxygenase-1, a cellular enzyme related to endogenous bilirubin synthesis. Moreover, we determined intracellular antioxidant activity by bilirubin, with EC50 = 11.4 ± 0.2 nM, in the range of reported values of free serum bilirubin (8.5-13.1 nM). Biliverdin showed similar antioxidant properties as bilirubin. We infer from these observations that intra-endothelial bilirubin oscillates, and may thus be a dynamic factor of the endothelial function. PMID:27381978

  9. Bilirubin is an Endogenous Antioxidant in Human Vascular Endothelial Cells

    PubMed Central

    Ziberna, Lovro; Martelanc, Mitja; Franko, Mladen; Passamonti, Sabina

    2016-01-01

    Bilirubin is a standard serum biomarker of liver function. Inexplicably, it is inversely correlated with cardiovascular disease risk. Given the role of endothelial dysfunction in originating cardiovascular diseases, direct analysis of bilirubin in the vascular endothelium would shed light on these relationships. Hence, we used high-performance liquid chromatography coupled with thermal lens spectrometric detection and diode array detection for the determination of endogenous cellular IXα-bilirubin. To confirm the isomer IXα-bilirubin, we used ultra-performance liquid chromatography coupled with a high-resolution mass spectrometer using an electrospray ionization source, as well as tandem mass spectrometric detection. We measured bilirubin in both arterial and venous rat endothelium (0.9–1.5 pmol mg−1 protein). In the human endothelial Ea.hy926 cell line, we demonstrated that intracellular bilirubin (3–5 pmol mg−1 protein) could be modulated by either extracellular bilirubin uptake, or by up-regulation of heme oxygenase-1, a cellular enzyme related to endogenous bilirubin synthesis. Moreover, we determined intracellular antioxidant activity by bilirubin, with EC50 = 11.4 ± 0.2 nM, in the range of reported values of free serum bilirubin (8.5–13.1 nM). Biliverdin showed similar antioxidant properties as bilirubin. We infer from these observations that intra-endothelial bilirubin oscillates, and may thus be a dynamic factor of the endothelial function. PMID:27381978

  10. Crystal structure of the dithiol oxidase DsbA enzyme from proteus mirabilis bound non-covalently to an active site peptide ligand.

    PubMed

    Kurth, Fabian; Duprez, Wilko; Premkumar, Lakshmanane; Schembri, Mark A; Fairlie, David P; Martin, Jennifer L

    2014-07-11

    The disulfide bond forming DsbA enzymes and their DsbB interaction partners are attractive targets for development of antivirulence drugs because both are essential for virulence factor assembly in Gram-negative pathogens. Here we characterize PmDsbA from Proteus mirabilis, a bacterial pathogen increasingly associated with multidrug resistance. PmDsbA exhibits the characteristic properties of a DsbA, including an oxidizing potential, destabilizing disulfide, acidic active site cysteine, and dithiol oxidase catalytic activity. We evaluated a peptide, PWATCDS, derived from the partner protein DsbB and showed by thermal shift and isothermal titration calorimetry that it binds to PmDsbA. The crystal structures of PmDsbA, and the active site variant PmDsbAC30S were determined to high resolution. Analysis of these structures allows categorization of PmDsbA into the DsbA class exemplified by the archetypal Escherichia coli DsbA enzyme. We also present a crystal structure of PmDsbAC30S in complex with the peptide PWATCDS. The structure shows that the peptide binds non-covalently to the active site CXXC motif, the cis-Pro loop, and the hydrophobic groove adjacent to the active site of the enzyme. This high-resolution structural data provides a critical advance for future structure-based design of non-covalent peptidomimetic inhibitors. Such inhibitors would represent an entirely new antibacterial class that work by switching off the DSB virulence assembly machinery. PMID:24831013

  11. Crystal Structure of the Dithiol Oxidase DsbA Enzyme from Proteus Mirabilis Bound Non-covalently to an Active Site Peptide Ligand

    PubMed Central

    Kurth, Fabian; Duprez, Wilko; Premkumar, Lakshmanane; Schembri, Mark A.; Fairlie, David P.; Martin, Jennifer L.

    2014-01-01

    The disulfide bond forming DsbA enzymes and their DsbB interaction partners are attractive targets for development of antivirulence drugs because both are essential for virulence factor assembly in Gram-negative pathogens. Here we characterize PmDsbA from Proteus mirabilis, a bacterial pathogen increasingly associated with multidrug resistance. PmDsbA exhibits the characteristic properties of a DsbA, including an oxidizing potential, destabilizing disulfide, acidic active site cysteine, and dithiol oxidase catalytic activity. We evaluated a peptide, PWATCDS, derived from the partner protein DsbB and showed by thermal shift and isothermal titration calorimetry that it binds to PmDsbA. The crystal structures of PmDsbA, and the active site variant PmDsbAC30S were determined to high resolution. Analysis of these structures allows categorization of PmDsbA into the DsbA class exemplified by the archetypal Escherichia coli DsbA enzyme. We also present a crystal structure of PmDsbAC30S in complex with the peptide PWATCDS. The structure shows that the peptide binds non-covalently to the active site CXXC motif, the cis-Pro loop, and the hydrophobic groove adjacent to the active site of the enzyme. This high-resolution structural data provides a critical advance for future structure-based design of non-covalent peptidomimetic inhibitors. Such inhibitors would represent an entirely new antibacterial class that work by switching off the DSB virulence assembly machinery. PMID:24831013

  12. Association of cytochrome c with membrane-bound cytochrome c oxidase proceeds parallel to the membrane rather than in bulk solution.

    PubMed

    Spaar, Alexander; Flöck, Dagmar; Helms, Volkhard

    2009-03-01

    Electron transfer between the water-soluble cytochrome c and the integral membrane protein cytochrome c oxidase (COX) is the terminal reaction in the respiratory chain. The first step in this reaction is the diffusional association of cytochrome c toward COX, and it is still not completely clear whether cytochrome c diffuses in the bulk solution while encountering COX, or whether it prefers to diffuse laterally on the membrane surface. This is a rather crucial question, since in the latter case the association would be strongly dependent on the lipid composition and the presence of additional membrane proteins. We applied Brownian dynamics simulations to investigate the effect of an atomistically modeled dipalmitoyl phosphatidylcholine membrane on the association behavior of cytochrome c toward COX from Paracoccus denitrificans. We studied the negatively charged, physiological electron-transfer partner of COX, cytochrome c(552), and the positively charged horse-heart cytochrome c. As expected, both cytochrome c species prefer diffusion in bulk solution while associating toward COX embedded in a membrane, where the partial charges of the lipids were switched off, and the corresponding optimal association pathways largely overlap with the association toward fully solvated COX. Remarkably, after switching on the lipid partial charges, both cytochrome c species were strongly attracted by the inhomogeneous charge distribution caused by the zwitterionic lipid headgroups. This effect is particularly enhanced for horse-heart cytochrome c and is stronger at lower ionic strength. We therefore conclude that in the presence of a polar or even a charged membrane, cytochrome c diffuses laterally rather than in three dimensions. PMID:19254533

  13. Studies of the conformation of bilirubin and its dimethyl ester in dimethyl sulphoxide solutions by nuclear magnetic resonance.

    PubMed Central

    Kaplan, D; Navon, G

    1982-01-01

    The conformation of bilirubin and its dimethyl ester in dimethyl sulphoxide (DMSO) was investigated by n.m.r. spectroscopy. The chemical shifts of the pyrrole NH and Lactam protons of bilirubin and its dimethyl ester in DMSO indicate a strong interaction with the solvent. Inter-proton distances were calculated from nuclear Overhauser effects (NOE), selective and non-selective relaxation times (T1) and rotational correlation times taken from 13C relaxation times. The interproton distances indicate that the conformation of the skeleton of bilirubin and its dimethyl ester in DMSO is similar to that of bilirubin and mesobilirubin in the crystalline state and in chloroform solutions, except for a possible slight twist of the pyrrolenone rings about the methine bonds, which may be a consequence of solvation of the NH groups by DMSO. Unlike in chloroform solutions, no direct hydrogen-bonding occurs between the carboxylic acid and the lactam groups of bilirubin in DMSO, as shown by the absence of an NOE between these groups. The fast exchange of the pyrrole NH protons with 2H shows that no hydrogen-bonding occurs between these protons and the propionic residues, in line with their solvation by DMSO. From the above results, and from the slowness of the internal motion of the propionic residues of bilirubin and its dimethyl ester, it is concluded that these residues are tied to the skeleton via bound solvent molecules. PMID:6284124

  14. Bilirubin-Induced Neurotoxicity in the Preterm Neonate.

    PubMed

    Watchko, Jon F

    2016-06-01

    Bilirubin-induced neurotoxicity in preterm neonates remains a clinical concern. Multiple cellular and molecular cascades likely underlie bilirubin-induced neuronal injury, including plasma membrane perturbations, excitotoxicity, neuroinflammation, oxidative stress, and cell cycle arrest. Preterm newborns are particularly vulnerable secondary to central nervous system immaturity and concurrent adverse clinical conditions that may potentiate bilirubin toxicity. Acute bilirubin encephalopathy in preterm neonates may be subtle and manifest primarily as recurrent symptomatic apneic events. Low-bilirubin kernicterus continues to be reported in preterm neonates, and although multifactorial in nature, is often associated with marked hypoalbuminemia. PMID:27235209

  15. Functionalized Magnetic Fe3O4-β-Cyclodextran Nanoparticles for Efficient Removal of Bilirubin.

    PubMed

    Han, Lulu; Chu, Simin; Wei, Houliang; Ren, Jun; Xu, Li; Jia, Lingyun

    2016-06-01

    Bilirubin (BR), as a lipophilic toxin, can binds and deposits in various tissues, especially the brain tissue, leading to hepatic coma and even death. Magnetic nanoparticles adsorbent modified by β-cyclodextran (Fe3O4-β-CD) was developed to remove the BR from the plasma. Fe3O4-β-CD nanoparticles was prepared through Schiff base reaction between the polyethylenimine (PEI)-modified Fe3O4 and aldehyde-functionalized β-CD, and characterized by Fourier transform infrared (FTIR) spectra, X-ray diffraction (XRD), transmission electron microscopy (TEM), vibrating sample magnetometer (VSM) and dynamic light scattering (DLS). Under optimized conditions, the Fe3O4-β-CD adsorbent could adsorb 225.6 mg/g free BR in PBS and reach the adsorption equilibrium within 90 min mainly through hydrophobic interaction; Moreover, the adsorbent displayed better adsorption capability in a dialysis system for BSA-bound bilirubin, plasma bilirubin and total bile acid, and the removal rates of those were 66%, 31% and 41% respectively. Because of the advantages of fast separation and purification process, low preparation cost, good adsorption capability for plasma bilirubin, Fe3O4-β-CD may become an economical and promising absorbent of BR for clinical applications. PMID:27427594

  16. Hepatocyte cotransport of taurocholate and bilirubin glucuronides: Role of microtubules

    SciTech Connect

    Crawford, J.M.; Gollan, J.L. )

    1988-07-01

    Modulation of bile pigment excretion by bile salts has been attributed to modification of canalicular membrane transport or a physical interaction in bile. Based on the observation that a microtubule-dependent pathway is involved in the hepatocellular transport of bile salts, the authors investigated the possibility that bilirubin glucuronides are associated with bile salts during intracellular transport. Experiments were conducted in intact rats (basal) or after overnight biliary diversion and intravenous reinfusion of taurocholate (depleted/reinfused). All rats were pretreated with intravenous low-dose colchicine or its inactive isomer lumicolchicine. Biliary excretion of radiolabeled bilirubin glucuronides derived from tracer ({sup 14}C)bilirubin-({sup 3}H)bilirubin monoglucuronide (coinjected iv) was unchanged in basal rats but was consistently delayed in depleted/reinfused rats. This was accompanied by a significant shift toward bilirubin diglucuronide formation from both substrates. In basal Gunn rats, with deficient bilirubin glucuronidation, biliary excretion of intravenous ({sup 14}C)bilirubin monoglucuronide-({sup 3}H)bilirubin diglucuronide was unaffected by colchicine but was retarded in depleted/reinfused Gunn rats. Colchicine had no effect on the rate of bilirubin glucuronidation in vitro in rat liver microsomes. They conclude that a portion of the bilirubin glucuronides generated endogenously in hepatocytes or taken up directly from plasma may be cotransported with bile salts to the bile canalicular membrane via a microtubule-dependent mechanism.

  17. Preliminary development of a fiber optic sensor for measuring bilirubin.

    PubMed

    Babin, Steven M; Sova, Raymond M

    2014-01-01

    Preliminary development of a fiber optic bilirubin sensor is described, where an unclad sensing portion is used to provide evanescent wave interaction of the transmitted light with the chemical environment. By using a wavelength corresponding to a bilirubin absorption peak, the Beer-Lambert Law can be used to relate the concentration of bilirubin surrounding the sensing portion to the amount of absorbed light. Initial testing in vitro suggests that the sensor response is consistent with the results of bulk absorption measurements as well as the Beer-Lambert Law. In addition, it is found that conjugated and unconjugated bilirubin have different peak absorption wavelengths, so that two optical frequencies may potentially be used to measure both types of bilirubin. Future development of this device could provide a means of real-time, point-of-care monitoring of intravenous bilirubin in critical care neonates with hyperbilirubinemia. PMID:25057239

  18. Preliminary Development of a Fiber Optic Sensor for Measuring Bilirubin

    PubMed Central

    Babin, Steven M; Sova, Raymond M

    2014-01-01

    Preliminary development of a fiber optic bilirubin sensor is described, where an unclad sensing portion is used to provide evanescent wave interaction of the transmitted light with the chemical environment. By using a wavelength corresponding to a bilirubin absorption peak, the Beer–Lambert Law can be used to relate the concentration of bilirubin surrounding the sensing portion to the amount of absorbed light. Initial testing in vitro suggests that the sensor response is consistent with the results of bulk absorption measurements as well as the Beer–Lambert Law. In addition, it is found that conjugated and unconjugated bilirubin have different peak absorption wavelengths, so that two optical frequencies may potentially be used to measure both types of bilirubin. Future development of this device could provide a means of real-time, point-of-care monitoring of intravenous bilirubin in critical care neonates with hyperbilirubinemia. PMID:25057239

  19. Bilirubin UDP-glucuronyltransferase activity of wistar rat kidney.

    PubMed

    Foliot, A; Christoforov, B; Petite, J P; Etienne, J P; Housset, E; Dubois, M

    1975-08-01

    Wistar rat kidneys have been shown to possess a bilirubin glucuronyltransferase (BGT) activity capable of conjugating about 3/5 of the total pool of unconjugated bilirubin within 48 h of being grafted to Gunn rat hosts. Bilirubin conjugated by the kidney is taken up by the liver and excreted in the bile. Except when the bile duct is ligated, no conjugated bilirubin appears in the plasma or urine. Renal BGT activity is about 1/20th of the hepatic activity on a weight basis in Wistar rats. The Gunn rat's hyperbilirubinemia probably causes an induction of the renal enzyme since its activity doubles in 48 h. PMID:808968

  20. Bilirubin and its oxidation products damage brain white matter.

    PubMed

    Lakovic, Katarina; Ai, Jinglu; D'Abbondanza, Josephine; Tariq, Asma; Sabri, Mohammed; Alarfaj, Abdullah K; Vasdev, Punarjot; Macdonald, Robert Loch

    2014-11-01

    Brain injury after intracerebral hemorrhage (ICH) occurs in cortex and white matter and may be mediated by blood breakdown products, including hemoglobin and heme. Effects of blood breakdown products, bilirubin and bilirubin oxidation products, have not been widely investigated in adult brain. Here, we first determined the effect of bilirubin and its oxidation products on the structure and function of white matter in vitro using brain slices. Subsequently, we determined whether these compounds have an effect on the structure and function of white matter in vivo. In all, 0.5 mmol/L bilirubin treatment significantly damaged both the function and the structure of myelinated axons but not the unmyelinated axons in brain slices. Toxicity of bilirubin in vitro was prevented by dimethyl sulfoxide. Bilirubin oxidation products (BOXes) may be responsible for the toxicity of bilirubin. In in vivo experiments, unmyelinated axons were found more susceptible to damage from bilirubin injection. These results suggest that unmyelinated axons may have a major role in white-matter damage in vivo. Since bilirubin and BOXes appear in a delayed manner after ICH, preventing their toxic effects may be worth investigating therapeutically. Dimethyl sulfoxide or its structurally related derivatives may have a potential therapeutic value at antagonizing axonal damage after hemorrhagic stroke. PMID:25160671

  1. Bilirubin and its oxidation products damage brain white matter

    PubMed Central

    Lakovic, Katarina; Ai, Jinglu; D'Abbondanza, Josephine; Tariq, Asma; Sabri, Mohammed; Alarfaj, Abdullah K; Vasdev, Punarjot; Macdonald, Robert Loch

    2014-01-01

    Brain injury after intracerebral hemorrhage (ICH) occurs in cortex and white matter and may be mediated by blood breakdown products, including hemoglobin and heme. Effects of blood breakdown products, bilirubin and bilirubin oxidation products, have not been widely investigated in adult brain. Here, we first determined the effect of bilirubin and its oxidation products on the structure and function of white matter in vitro using brain slices. Subsequently, we determined whether these compounds have an effect on the structure and function of white matter in vivo. In all, 0.5 mmol/L bilirubin treatment significantly damaged both the function and the structure of myelinated axons but not the unmyelinated axons in brain slices. Toxicity of bilirubin in vitro was prevented by dimethyl sulfoxide. Bilirubin oxidation products (BOXes) may be responsible for the toxicity of bilirubin. In in vivo experiments, unmyelinated axons were found more susceptible to damage from bilirubin injection. These results suggest that unmyelinated axons may have a major role in white-matter damage in vivo. Since bilirubin and BOXes appear in a delayed manner after ICH, preventing their toxic effects may be worth investigating therapeutically. Dimethyl sulfoxide or its structurally related derivatives may have a potential therapeutic value at antagonizing axonal damage after hemorrhagic stroke. PMID:25160671

  2. Mitochondrial targeting of bilirubin regulatory enzymes: An adaptive response to oxidative stress

    SciTech Connect

    Muhsain, Siti Nur Fadzilah; Lang, Matti A.; Abu-Bakar, A'edah

    2015-01-01

    The intracellular level of bilirubin (BR), an endogenous antioxidant that is cytotoxic at high concentrations, is tightly controlled within the optimal therapeutic range. We have recently described a concerted intracellular BR regulation by two microsomal enzymes: heme oxygenase 1 (HMOX1), essential for BR production and cytochrome P450 2A5 (CYP2A5), a BR oxidase. Herein, we describe targeting of these enzymes to hepatic mitochondria during oxidative stress. The kinetics of microsomal and mitochondrial BR oxidation were compared. Treatment of DBA/2J mice with 200 mg pyrazole/kg/day for 3 days increased hepatic intracellular protein carbonyl content and induced nucleo-translocation of Nrf2. HMOX1 and CYP2A5 proteins and activities were elevated in microsomes and mitoplasts but not the UGT1A1, a catalyst of BR glucuronidation. A CYP2A5 antibody inhibited 75% of microsomal BR oxidation. The inhibition was absent in control mitoplasts but elevated to 50% after treatment. An adrenodoxin reductase antibody did not inhibit microsomal BR oxidation but inhibited 50% of mitochondrial BR oxidation. Ascorbic acid inhibited 5% and 22% of the reaction in control and treated microsomes, respectively. In control mitoplasts the inhibition was 100%, which was reduced to 50% after treatment. Bilirubin affinity to mitochondrial and microsomal CYP2A5 enzyme is equally high. Lastly, the treatment neither released cytochrome c into cytoplasm nor dissipated membrane potential, indicating the absence of mitochondrial membrane damage. Collectively, the observations suggest that BR regulatory enzymes are recruited to mitochondria during oxidative stress and BR oxidation by mitochondrial CYP2A5 is supported by mitochondrial mono-oxygenase system. The induced recruitment potentially confers membrane protection. - Highlights: • Pyrazole induces oxidative stress in the mouse liver. • Pyrazole-induced oxidative stress induces mitochondrial targeting of key bilirubin regulatory enzymes, HMOX1

  3. Incorporation of copper into lysyl oxidase.

    PubMed

    Kosonen, T; Uriu-Hare, J Y; Clegg, M S; Keen, C L; Rucker, R B

    1997-10-01

    Lysyl oxidase is a copper-dependent enzyme involved in extracellular processing of collagens and elastin. Although it is known that copper is essential for the functional activity of the enzyme, there is little information on the incorporation of copper. In the present study we examined the insertion of copper into lysyl oxidase using 67Cu in cell-free transcription/translation assays and in normal skin fibroblast culture systems. When a full-length lysyl oxidase cDNA was used as a template for transcription/translation reactions in vitro, unprocessed prolysyl oxidase appeared to bind copper. To examine further the post-translational incorporation of copper into lysyl oxidase, confluent skin fibroblasts were incubated with inhibitors of protein synthesis (cycloheximide, 10 microg/ml), glycosylation (tunicamycin, 10 microg/ml), protein secretion (brefeldin A, 10 microg/ml) and prolysyl oxidase processing (procollagen C-peptidase inhibitor, 2.5 microg/ml) together with 300 microCi of carrier-free 67Cu. It was observed that protein synthesis was a prerequisite for copper incorporation, but inhibition of glycosylation by tunicamycin did not affect the secretion of 67Cu as lysyl oxidase. Brefeldin A inhibited the secretion of 67Ci-labelled lysyl oxidase by 46%, but the intracellular incorporation of copper into lysyl oxidase was not affected. In addition, the inhibition of the extracellular proteolytic processing of prolysyl oxidase to lysyl oxidase had minimal effects on the secretion of protein-bound 67Cu. Our results indicate that, similar to caeruloplasmin processing [Sato and Gitlin (1991) J. Biol. Chem. 266, 5128-5134], copper is inserted into prolysyl oxidase independently of glycosylation. PMID:9355764

  4. Structural Insights into Sulfite Oxidase Deficiency

    SciTech Connect

    Karakas,E.; Wilson, H.; Graf, T.; Xiang, S.; Jaramillo-Busquets, S.; Rajagopalan, K.; Kisker, C.

    2005-01-01

    Sulfite oxidase deficiency is a lethal genetic disease that results from defects either in the genes encoding proteins involved in molybdenum cofactor biosynthesis or in the sulfite oxidase gene itself. Several point mutations in the sulfite oxidase gene have been identified from patients suffering from this disease worldwide. Although detailed biochemical analyses have been carried out on these mutations, no structural data could be obtained because of problems in crystallizing recombinant human and rat sulfite oxidases and the failure to clone the chicken sulfite oxidase gene. We synthesized the gene for chicken sulfite oxidase de novo, working backward from the amino acid sequence of the native chicken liver enzyme by PCR amplification of a series of 72 overlapping primers. The recombinant protein displayed the characteristic absorption spectrum of sulfite oxidase and exhibited steady state and rapid kinetic parameters comparable with those of the tissue-derived enzyme. We solved the crystal structures of the wild type and the sulfite oxidase deficiency-causing R138Q (R160Q in humans) variant of recombinant chicken sulfite oxidase in the resting and sulfate-bound forms. Significant alterations in the substrate-binding pocket were detected in the structure of the mutant, and a comparison between the wild type and mutant protein revealed that the active site residue Arg-450 adopts different conformations in the presence and absence of bound sulfate. The size of the binding pocket is thereby considerably reduced, and its position relative to the cofactor is shifted, causing an increase in the distance of the sulfur atom of the bound sulfate to the molybdenum.

  5. INDUCTION OF BILIRUBIN CLEARANCE BY THE CONSTITUTIVE ANDROSTANE RECEPTOR (CAR)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bilirubin clearance is one of the numerous important functions of the liver. Defects in this process result in jaundice, which is particularly common in neonates. Elevated bilirubin levels can be decreased by treatment with phenobarbital. Because the nuclear hormone receptor constitutive androstane ...

  6. Conjugated Bilirubin Triggers Anemia by Inducing Erythrocyte Death

    PubMed Central

    Lang, Elisabeth; Gatidis, Sergios; Freise, Noemi F; Bock, Hans; Kubitz, Ralf; Lauermann, Christian; Orth, Hans Martin; Klindt, Caroline; Schuier, Maximilian; Keitel, Verena; Reich, Maria; Liu, Guilai; Schmidt, Sebastian; Xu, Haifeng C; Qadri, Syed M; Herebian, Diran; Pandyra, Aleksandra A; Mayatepek, Ertan; Gulbins, Erich; Lang, Florian; Häussinger, Dieter; Lang, Karl S; Föller, Michael; Lang, Philipp A

    2015-01-01

    Hepatic failure is commonly associated with anemia, which may result from gastrointestinal bleeding, vitamin deficiency, or liver-damaging diseases, such as infection and alcohol intoxication. At least in theory, anemia during hepatic failure may result from accelerated clearance of circulating erythrocytes. Here we show that bile duct ligation (BDL) in mice leads to severe anemia despite increased reticulocyte numbers. Bilirubin stimulated suicidal death of human erythrocytes. Mechanistically, bilirubin triggered rapid Ca2+ influx, sphingomyelinase activation, formation of ceramide, and subsequent translocation of phosphatidylserine to the erythrocyte surface. Consistent with our in vitro and in vivo findings, incubation of erythrocytes in serum from patients with liver disease induced suicidal death of erythrocytes in relation to their plasma bilirubin concentration. Consistently, patients with hyperbilirubinemia had significantly lower erythrocyte and significantly higher reticulocyte counts compared to patients with low bilirubin levels. Conclusion: Bilirubin triggers suicidal erythrocyte death, thus contributing to anemia during liver disease. (Hepatology 2015;61:275–284) PMID:25065608

  7. Bilirubin Nanoparticles as a Nanomedicine for Anti-inflammation Therapy.

    PubMed

    Lee, Yonghyun; Kim, Hyungjun; Kang, Sukmo; Lee, Jinju; Park, Jinho; Jon, Sangyong

    2016-06-20

    Despite the high potency of bilirubin as an endogenous anti-inflammatory compound, its clinical translation has been hampered because of its insolubility in water. Bilirubin-based nanoparticles that may overcome this critical issue are presented. A polyethylene glycol compound (PEG) was covalently attached to bilirubin, yielding PEGylated bilirubin (PEG-BR). The PEG-BR self-assembled into nanoscale particles with a size of approximately 110 nm, termed bilirubin nanoparticles (BRNPs). BRNPs are highly efficient hydrogen peroxide scavengers, thereby protecting cells from H2 O2 -induced cytotoxicity. In a murine model of ulcerative colitis, intravenous injection of BRNPs showed preferential accumulation of nanoparticles at the sites of inflammation and significantly inhibited the progression of acute inflammation in the colon. Taken together, BRNPs show potential for use as a therapeutic nanomedicine in various inflammatory diseases. PMID:27144463

  8. In Cellulo Mapping of Subcellular Localized Bilirubin.

    PubMed

    Park, Jong-Seok; Nam, Eunju; Lee, Hye-Kyeong; Lim, Mi Hee; Rhee, Hyun-Woo

    2016-08-19

    Bilirubin (BR) is a de novo synthesized metabolite of human cells. However, subcellular localization of BR in the different organelles of human cells has been largely unknown. Here, utilizing UnaG as a genetically encoded fluorescent BR sensor, we report the existence of relatively BR-enriched and BR-depleted microspaces in various cellular organelles of live cells. Our studies indicate that (i) the cytoplasmic facing membrane of the endoplasmic reticulum (ER) and the nucleus are relatively BR-enriched spaces and (ii) mitochondrial intermembrane space and the ER lumen are relatively BR-depleted spaces. Thus, we demonstrate a relationship between such asymmetrical BR distribution in the ER membrane and the BR metabolic pathway. Furthermore, our results suggest plausible BR-transport and BR-regulating machineries in other cellular compartments, including the nucleus and mitochondria. PMID:27232847

  9. Trans-Cutaneous Bilirubinometery versus Serum Bilirubin in Neonatal Jaundice.

    PubMed

    Mahram, Manoochehr; Oveisi, Sonia; Jaberi, Najmeh

    2015-12-01

    Hyperbilirubinemia is a common problem in neonates and causes serious complications. Thus, serial measurements of bilirubin should be done. This assessment is done through two methods of laboratory measurement in serum sample and transcutaneous bilirubinometer. This descriptive study compared transcutaneous bilirubin assessment and laboratory serum bilirubin. Bilirubin level was assessed among 256 neonates admitted to the Qods Children's Hospital in Qazvin- Iran, because of neonatal indirect jaundice, through two methods of transcutaneous bilirubinometery from two sites of forehead and sternum and laboratory measurement of bilirubin in serum. The cases were non-hemolytic icteric term neonates weighing 2500 gram or more and had not received phototherapy or other treatments. Neonates with hemolytic forms of jaundice, sepsis and suspicious to metabolic disorders were excluded. Assessments by means of KJ-8000 transcutaneous bilirubinometer from two sites of forehead and sternum and through laboratory measurement of serum bilirubin were registered and analyzed. The results of the current study showed that there was a correlation of 0.82 between serum bilirubin and transcutaneous forehead bilirubin assessment and for the used device sensitivity of 0.844; specificity of 0.842, Youden Index of 0.709 and Shortest of 0.042 for a cut-off of 12.4 in bilirubin of participants. Furthermore, Likelihood Ratio positive and negative (LR) were 5.665 and 0.164, respectively and diagnostic Odds Ratio (LR+/LR-) was 34.56. Transcutaneous bilirubinometery can be considered as a reliable tool to assess bilirubin for the screening of neonatal jaundice in term neonates. PMID:26749233

  10. [Does bilirubin interfere with capillary electrophoresis of serum proteins?].

    PubMed

    Hellara, Ilhem; Fekih, Ons; Triki, Sonia; Elmay, Ahlem; Neffati, Fadoua; Najjar, Mohamed Fadhel

    2014-01-01

    Capillary electrophoresis of serum proteins is a fast, reliable and simple technique, but many interference exist. The objective of our work is to study the interference of bilirubin on this technique; 70 icteric sera were analysed on Capillarys ™ (Sebia). A second electrophoresis was performed on 40 samples after bilirubin photodegradation. The bilirubin and serum proteins were determinated respectively by Jendrassik and Grof and biuret methods on Konélab 20i ™ (Thermo Electron Corporation). We found abnormal spreading of the albumin fraction of the anode side wich constitute sometimes an isolated fraction in the traditional area of pre-albumin migration. This fraction varies from 2.0 ± 2.0% (0.0 to 7.3%) or 0.98 ± 1.53 g/L (0 to 5.3 g/L) and it seems to be related to the direct bilirubin since, following overloading sera with a solution of bilirubin, no further fraction was recovered. An average decrease of bilirubin after photodegradation of 58 ± 17% (26-89%) is followed by a decrease in the same order 64 ± 38% (10-100%) of the additional fraction. Acetate cellulose electrophoresis of the same samples showed no variation. The high bilirubin levels seem modify slightly the electrophoretic profile. However the impact of the interference on the interpretation of electrophoretic trace is negligible. PMID:24492101

  11. Association of abnormal plasma bilirubin with aggressive hepatocellular carcinoma phenotype.

    PubMed

    Carr, Brian I; Guerra, Vito; Giannini, Edoardo G; Farinati, Fabio; Ciccarese, Francesca; Ludovico Rapaccini, Gian; Di Marco, Maria; Benvegnù, Luisa; Zoli, Marco; Borzio, Franco; Caturelli, Eugenio; Chiaramonte, Maria; Trevisani, Franco

    2014-04-01

    Cirrhosis-related abnormal liver function is associated with predisposition to hepatocellular carcinoma (HCC). It features in several HCC classification systems and is an HCC prognostic factor. The aim of the present study was to examine the phenotypic tumor differences in HCC patients with normal or abnormal plasma bilirubin levels. A 2,416-patient HCC cohort was studied and dichotomized into normal and abnormal plasma bilirubin groups. Their HCC characteristics were compared for tumor aggressiveness features, namely, blood alpha-fetoprotein (AFP) levels, tumor size, presence of portal vein thrombosis (PVT) and tumor multifocality. In the total cohort, elevated bilirubin levels were associated with higher AFP levels, increased PVT and multifocality, and lower survival, despite similar tumor sizes. When different tumor size terciles were compared, similar results were found, even among patients with small tumors. A multiple logistic regression model for PVT or tumor multifocality showed increased odds ratios for elevated levels of gamma glutamyl transpeptidase (GGTP), bilirubin, and AFP and for larger tumor sizes. We conclude that HCC patients with abnormal bilirubin levels had worse prognosis than patients with normal bilirubin. They also had an increased incidence of PVT and tumor multifocality, and higher AFP levels, in patients with both small and larger tumors. The results show an association between bilirubin levels and indices of HCC aggressiveness. PMID:24787296

  12. Acute Alcohol Consumption Elevates Serum Bilirubin, an Endogenous Antioxidant

    PubMed Central

    O’Malley, Stephanie S.; Gueorguieva, Ralitza; Wu, Ran; Jatlow, Peter I.

    2015-01-01

    Background Moderate alcohol consumption has been associated with both negative and favorable effects on health. The mechanisms responsible for reported favorable effects remain unclear. Higher (not necessarily elevated) concentrations of serum bilirubin, an antioxidant, have also been associated with reduced risk of cardiovascular disease and all-cause mortality. This study tests the hypothesis that single dose alcohol consumption elevates bilirubin providing a potential link between these observations. Methods 18 healthy individuals (8 cigarette smokers) were administered alcohol, calibrated to achieve blood concentrations of 20, 80 and 120 mg/dL, in random order in 3 laboratory sessions separated by a week. Each session was preceded by and followed by 5–7 days of alcohol abstinence. Serum bilirubin was measured at 7:45 am prior to drinking, at 2 pm, and at 7:45 the next morning. Mixed effects regression models compared baseline and 24 hr. post-drinking bilirubin concentrations. Results Total serum bilirubin (sum of indirect and direct) concentration increased significantly after drinking from baseline to 24 hours in non-smokers (from Mean=0.38, SD=0.24 to Mean=0.51 SD=0.30, F(1, 32.2) =24.24, p<.0001) but not in smokers (from Mean=0.25, SD=0.12 to Mean=0.26, SD=0.15, F(1, 31.1) =0.04, p=0.84). In nonsmokers the indirect bilirubin concentration and the ratio of indirect (unconjugated) to direct (conjugated) bilirubin also increased significantly. Conclusions Alcohol consumption leads to increases in serum bilirubin in nonsmokers. Considering the antioxidant properties of bilirubin, our findings suggest one possible mechanism for the reported association between alcohol consumption and reduced risk of some disorders that could be tested in future longitudinal studies. PMID:25707709

  13. Bilirubin Neurotoxicity in Preterm Infants: Risk and Prevention

    PubMed Central

    Bhutani, Vinod K.; Wong, Ronald J.

    2013-01-01

    Hemolytic conditions in preterm neonates, including Rhesus (Rh) disease, can lead to mortality and long-term impairments due to bilirubin neurotoxicity. Universal access to Rh immunoprophylaxis, coordinated perinatal-neonatal care, and effective phototherapy has virtually eliminated the risk of kernicterus in many countries. In the absence of jaundice due to isoimmunization and without access to phototherapy or exchange transfusion (in 1955), kernicterus was reported at 10.1%, 5.5%, and 1.2% in babies <30, 31-32, and 33-34 wks gestational age, respectively. Phototherapy initiated at 24±12 hr effectively prevented hyperbilirubinemia in infants <2,000 g even in the presence of hemolysis. This approach (in 1985) reduced exchange transfusions from 23.9% to 4.8%. Now with 3 decades of experience in implementing effective phototherapy, the need for exchange transfusions has virtually been eliminated. However, bilirubin neurotoxicity continues to be associated with prematurity alone. The ability to better predict this risk, other than birthweight and gestation, has been elusive. Objective tests such as total bilirubin, unbound or free bilirubin, albumin levels, and albumin-bilirubin binding, together with observations of concurrent hemolysis, sepsis, and rapid rate of bilirubin rise have been considered, but their individual or combined predictive utility has yet to be refined. The disruptive effects of immaturity, concurrent neonatal disease, cholestasis, use of total parenteral nutrition or drugs that alter bilirubin-binding abilities augment the clinical risk of neurotoxicity. Current management options rely on the “fine-tuning” of each infant's exposure to beneficial antioxidants and avoidance of silent neurotoxic properties of bilirubin navigated within the safe spectrum of operational thresholds demarcated by experts. PMID:24049745

  14. Bilirubin as an endogenous modulator of neurotrophin redox signaling.

    PubMed

    Mancuso, Cesare; Capone, Caterina; Ranieri, Sofia Chiatamone; Fusco, Salvatore; Calabrese, Vittorio; Eboli, Maria Luisa; Preziosi, Paolo; Galeotti, Tommaso; Pani, Giovambattista

    2008-08-01

    Bilirubin is neurotoxic upon excess accumulation in the brain, but it also plays important physiological roles related to its antioxidant properties. Here we report that exposure of PC12 and primary rat cerebellar granule neurons to bilirubin (0.5-10 microM) drastically decreases nerve growth factor (NGF)/brain-derived neurotrophic factor signaling to Akt and extracellular signal-regulated kinases (ERKs), indicating a direct interference of the molecule with crucial prosurvival signaling pathways. This effect likely involves the scavenging capacity of bilirubin, the latter being able to inhibit, in PC12 cells, accumulation of intracellular reactive oxygen species and phosphorylation of Akt and ERKs in response to extracellular hydrogen peroxide. Interestingly, in the absence of exogenous growth factor, bilirubin elicited the phosphorylation of ERKs and of the cAMP responsive element binding (CREB) transcription factor, a signature of NGF-dependent survival signaling. These growth factor-like signaling effects were paralleled by the induction of the neuronal nitric oxide synthase (nNOS) and generation of nitric oxide (NO). Pharmacological dissection of the signaling cascade triggered by bilirubin revealed that phosphorylation of ERKs requires NO signaling through soluble guanylyl cyclase, and, further upstream, influx of extracellular calcium is necessary for nNOS induction and NO release, likely through calcium-dependent phosphorylation of CREB. Importantly, the cascade elicited by bilirubin through NO and ERK is cytoprotective, as revealed by exacerbated bilirubin toxicity in cultures treated by either NOS or MEK inhibitors. Taken together, these observations indicate an important action of bilirubin on redox signaling by neurotrophins, with either inhibitory or agonistic effects based on growth factor availability. PMID:18338802

  15. Hepatic Conversion of Bilirubin Monoglucuronide to Diglucuronide in Uridine Diphosphate-Glucuronyl Transferase-Deficient Man and Rat by Bilirubin Glucuronoside Glucuronosyltransferase

    PubMed Central

    Chowdhury, J. Roy; Jansen, P. L. M.; Fischberg, E. B.; Daniller, A.; Arias, I. M.

    1978-01-01

    The microsomal enzyme uridine diphosphate (UDP) glucuronate glucuronyltransferase (E.C. 2.4.1.17) catalyzes formation of bilirubin mono-glucuronide from bilirubin and UDPglucuronic acid. Bilirubin glucuronoside glucuronosyltransferase (E.C. 2.4.1.95), an enzyme concentrated in plasma membrane-enriched fractions of rat liver, converts bilirubin monoglucuronide to bilirubin diglucuronide. Bilirubin glucuronoside glucuronosyltransferase activity was studied in homogenates of liver biopsy specimens obtained from patients with the Crigler-Najjar syndrome (Type I) and in subcellular liver fractions of rats homozygous for UDP glucuronate glucuronyltransferase deficiency (Gunn strain). In patients with the Crigler-Najjar syndrome (Type I) and in Gunn rats, hepatic UDPglucuronate glucuronyltransferase activity was not measurable; however, bilirubin glucuronoside glucuronosyltransferase activity was similar to that in normal controls. The subcellular distribution of bilirubin glucuronoside glucuronosyltransferase activity in Gunn rat liver was similar to the distribution observed in normal Wistar rat liver. When bilirubin monoglucuronide was infused intravenously into Gunn rats, 29±5% of the conjugated bilirubin excreted in bile was bilirubin diglucuronide. After transplantation of normal Wistar rat kidney, which contained UDPglucuronate glucuronyltransferase activity, in Gunn rats, the serum bilirubin concentration decreased by 80% in 4 days. The major route of bilirubin removal was biliary excretion of conjugated bilirubin, approximately 70% of which was bilirubin diglucuronide. Although patients with the Crigler-Najjar syndrome (Type I) and Gunn rats lack UDP glucuronate glucuronyltransferase, their livers enzymatically convert bilirubin monoglucuronide to diglucuronide in vitro. Conversion in bilirubin monoglucuronide to diglucuronide was demonstrated in Gunn rats in vivo. PMID:96142

  16. Influence of hemoglobin on non-invasive optical bilirubin sensing

    NASA Astrophysics Data System (ADS)

    Jiang, Jingying; Gong, Qiliang; Zou, Da; Xu, Kexin

    2012-03-01

    Since the abnormal metabolism of bilirubin could lead to diseases in the human body, especially the jaundice which is harmful to neonates. Traditional invasive measurements are difficult to be accepted by people because of pain and infection. Therefore, the real-time and non-invasive measurement of bilirubin is of great significance. However, the accuracy of currently transcutaneous bilirubinometry(TcB) is generally not high enough, and affected by many factors in the human skin, mostly by hemoglobin. In this talk, absorption spectra of hemoglobin and bilirubin have been collected and analyzed, then the Partial Least Squares (PLS) models have been built. By analyzing and comparing the Correlation and Root Mean Square Error of Prediction(RMSEP), the results show that the Correlation of bilirubin solution model is larger than that of the mixture solution added with hemoglobin, and its RMSEP value is smaller than that of mixture solution. Therefore, hemoglobin has influences on the non-invasive optical bilirubin sensing. In next step, it is necessary to investigate how to eliminate the influence.

  17. Mitochondrial targeting of bilirubin regulatory enzymes: An adaptive response to oxidative stress.

    PubMed

    Muhsain, Siti Nur Fadzilah; Lang, Matti A; Abu-Bakar, A'edah

    2015-01-01

    The intracellular level of bilirubin (BR), an endogenous antioxidant that is cytotoxic at high concentrations, is tightly controlled within the optimal therapeutic range. We have recently described a concerted intracellular BR regulation by two microsomal enzymes: heme oxygenase 1 (HMOX1), essential for BR production and cytochrome P450 2A5 (CYP2A5), a BR oxidase. Herein, we describe targeting of these enzymes to hepatic mitochondria during oxidative stress. The kinetics of microsomal and mitochondrial BR oxidation were compared. Treatment of DBA/2J mice with 200mgpyrazole/kg/day for 3days increased hepatic intracellular protein carbonyl content and induced nucleo-translocation of Nrf2. HMOX1 and CYP2A5 proteins and activities were elevated in microsomes and mitoplasts but not the UGT1A1, a catalyst of BR glucuronidation. A CYP2A5 antibody inhibited 75% of microsomal BR oxidation. The inhibition was absent in control mitoplasts but elevated to 50% after treatment. An adrenodoxin reductase antibody did not inhibit microsomal BR oxidation but inhibited 50% of mitochondrial BR oxidation. Ascorbic acid inhibited 5% and 22% of the reaction in control and treated microsomes, respectively. In control mitoplasts the inhibition was 100%, which was reduced to 50% after treatment. Bilirubin affinity to mitochondrial and microsomal CYP2A5 enzyme is equally high. Lastly, the treatment neither released cytochrome c into cytoplasm nor dissipated membrane potential, indicating the absence of mitochondrial membrane damage. Collectively, the observations suggest that BR regulatory enzymes are recruited to mitochondria during oxidative stress and BR oxidation by mitochondrial CYP2A5 is supported by mitochondrial mono-oxygenase system. The induced recruitment potentially confers membrane protection. PMID:25478736

  18. Bilirubin-Induced Audiologic Injury in Preterm Infants.

    PubMed

    Olds, Cristen; Oghalai, John S

    2016-06-01

    Although hyperbilirubinemia is extremely common among neonates and is usually mild and transient, it sometimes leads to bilirubin-induced neurologic damage (BIND). The auditory pathway is highly sensitive to the effects of elevated total serum/plasma bilirubin (TB) levels, with damage manifesting clinically as auditory neuropathy spectrum disorder. Compared to full-term neonates, preterm neonates are more susceptible to BIND and suffer adverse effects at lower TB levels with worse long-term outcomes. Furthermore, although standardized guidelines for management of hyperbilirubinemia exist for term and late preterm neonates, similar guidelines for neonates less than 35 weeks gestational age are limited. PMID:27235210

  19. [Micelle-mediated extraction for concentrating conjugated bilirubin in urine].

    PubMed

    Matsudo, T; Saitoh, T; Matsubara, C

    2001-02-01

    An extraction method based on the phase separation of aqueous micellar solutions of n-octyl-beta-D-thioglucoside (OTG) was applied to the concentrating conjugated bilirubin in urine. The analyte in sample solutions could be efficiently concentrated into a small volume of surfactant-rich phase, while hydrophilic matrix components including urinary protein, ascorbic acid, and saccharide remained in the aqueous phase. The concentrated OTG negligibly affected the diazo reaction and the subsequent spectrophotometric detection. Conjugated bilirubin was successfully determined in the concentration range from 0.05 microgram/ml to 5 micrograms/ml with a 96-well microplate reader absorption spectrophotometer. PMID:11218735

  20. Galactosaemia: an unusual cause of chronic bilirubin encephalopathy.

    PubMed

    Sahoo, Tanushree; Thukral, Anu; Agarwal, Ramesh; Sankar, Mari Jeeva

    2015-01-01

    Galactosaemia is a disorder of galactose metabolism in which raised levels of galactose and galactose-1-phosphate damage various organs. Although galactosaemia is a common metabolic liver disease in childhood, it is a rare cause of neonatal hyperbilirubinemia requiring intervention. We report an unusual case of neonatal galactosaemia that at presentation had features of acute bilirubin encephalopathy requiring exchange transfusion and at discharge had features of chronic bilirubin encephalopathy. This case report emphasises the need for timely suspicion and diagnosis of this disease for prevention of chronic morbidity. PMID:25618877

  1. Neonatal jaundice: a critical review of the role and practice of bilirubin analysis.

    PubMed

    Kirk, Jean M

    2008-09-01

    Neonatal jaundice is common, and usually harmless, because of physiological jaundice or breast-feeding. In some neonates unconjugated bilirubin concentration, coupled with other risk factors, is sufficient to allow free bilirubin to cross the blood-brain barrier and cause kernicterus. Another subgroup of infants is jaundiced because of elevated conjugated bilirubin; a marker for a number of pathological conditions. Bilirubin measurement must identify those infants at risk. Transcutaneous bilirubin measurement is increasingly used in healthy infants, especially before early discharge or at home, to assess the need for laboratory bilirubin measurement. Transcutaneous measurements are not covered by laboratory quality assessment schemes. Guidelines on management of neonatal jaundice utilize age in hours and other risk factors to define bilirubin action thresholds, which may be as low as 100 micromol/L for sick premature infants, whereas early discharged babies may only present after bilirubin concentrations are extremely high. Hence, there is a requirement for accurate total bilirubin measurement from <100 to >500 micromol/L, with sufficient precision to assess the rate of bilirubin change with time. Babies presenting with late jaundice always require conjugated bilirubin measurement. It is of concern that many total and direct bilirubin automated kit methods suffer from haemolysis interference, while use of in-house methods or modification of commercial methods has virtually disappeared. External quality assessment has a vital role in providing data on different methods' performance, including accuracy, precision and susceptibility to interference. Laboratories should consider whether their adult bilirubin methods are suitable for neonates. PMID:18753416

  2. Developmental onset of bilirubin-induced neurotoxicity involves Toll-like receptor 2-dependent signaling in humanized UDP-glucuronosyltransferase1 mice.

    PubMed

    Yueh, Mei-Fei; Chen, Shujuan; Nguyen, Nghia; Tukey, Robert H

    2014-02-21

    Biological and signaling events that connect developmentally induced hyperbilirubinemia to bilirubin-induced neurological dysfunction (BIND) and CNS toxicity in humans are poorly understood. In mammals, UDP-glucuronosyltransferase 1A1 (UGT1A1) is the sole enzyme responsible for bilirubin glucuronidation, a rate-limiting step necessary for bilirubin metabolism and clearance. Humanized mice that express the entire UGT1 locus (hUGT1) and the UGT1A1 gene, develop neonatal hyperbilirubinemia, with 8-10% of hUGT1 mice succumbing to CNS damage, a phenotype that is presented by uncontrollable seizures. We demonstrate that neuroinflammation and reactive gliosis are prominent features of bilirubin brain toxicity, and a disturbed redox status resulting from activation of NADPH oxidase is an important contributing mechanism found in BIND. Using knock-out mice and primary brain cells, we connect a key pattern recognition receptor, Toll-like receptor 2 (TLR2), to hyperbilirubinemia-induced signaling. We illustrate a requirement for TLR2 signaling in regulating gliosis, proinflammatory mediators, and oxidative stress when neonatal mice encounter severe hyperbilirubinemia. TLR2-mediated gliosis strongly correlates with pronounced neuroinflammation in the CNS with up-regulation of TNFα, IL-1β, and IL-6, creating a pro-inflammatory CNS environment. Gene expression and immunohistochemistry staining show that hUGT1/Tlr2(-/-) mice fail to activate glial cells, proinflammatory cytokines, and stress response genes. In addition, bilirubin-induced apoptosis was significantly enhanced by blocking TLR2 signaling indicating its anti-apoptotic property. Consequently, a higher neonatal death rate (57.1%) in hUGT1/Tlr2(-/-) mice was observed when compared with hUGT1 mice (8.7%). These results suggest that TLR2 signaling and microglia neuroinflammation are linked to a repair and/or protection mode against BIND. PMID:24403077

  3. Nanofibrous polymeric beads from aramid fibers for efficient bilirubin removal.

    PubMed

    Peng, Zihang; Yang, Ye; Luo, Jiyue; Nie, Chuanxiong; Ma, Lang; Cheng, Chong; Zhao, Changsheng

    2016-08-16

    Polymer based hemoperfusion has been developed as an effective therapy to remove the extra bilirubin from patients. However, the currently applied materials suffer from either low removal efficiency or poor blood compatibility. In this study, we report the development of a new class of nanofibrous absorbent that exhibited high bilirubin removal efficiency and good blood compatibility. The Kevlar nanofiber was prepared by dissolving micron-sized Kevlar fiber in proper solvent, and the beads were prepared by dropping Kevlar nanofiber solutions into ethanol. Owing to the nanofiborous structure of the Kevlar nanofiber, the beads displayed porous structures and large specific areas, which would facilitate the adsorption of toxins. In the adsorption test, it was noticed that the beads possessed an adsorption capacity higher than 40 mg g(-1) towards bilirubin. In plasma mimetic solutions, the beads still showed high bilirubin removal efficiency. Furthermore, after incorporating with carbon nanotubes, the beads were found to have increased adsorption capacity for human degradation waste. Moreover, the beads showed excellent blood compatibility in terms of a low hemolysis ratio, prolonged clotting times, suppressed coagulant activation, limited platelet activation, and inhibited blood related inflammatory activation. Additionally, the beads showed good compatibility with endothelial cells. In general, the Kevlar nanofiber beads, which integrated with high adsorption capacity, good blood compatibility and low cytotoxicity, may have great potential for hemoperfusion and some other applications in biomedical fields. PMID:27481656

  4. Biphasic Effect of Rifampicin on Bilirubin- A Case Report.

    PubMed

    Gopi, Manigandan; Seshadri, Mandalam Subramanian

    2016-04-01

    Drug induced hepatitis is a major problem which a physician encounters in his clinical practice. In view of increasing incidence of tuberculosis in our country a large number of infected individuals are started on Antituberculous (ATT) drugs and rifampicin is invariably part of the regimen. One of the major adverse effects of ATT drugs is drug- induced hepatitis which is characterized by elevation of liver enzymes and bilirubin. Hepatotoxicity is usually idiosyncratic or dose-dependent. Rifampicin causes transient elevation of transaminases in 10-20 percent of individuals and this does not warrant dose adjustments of the drug. Rarely rifampicin can lead to severe hepatitis with hyperbilirubinaemia and marked elevations of SGOT and SGPT and in some patients this can be fatal. The exact mechanism of Rifampicin induced hepatotoxicity is not known but it is postulated to be due to idiosyncratic reaction to rifampicin metabolites which may be directly toxic or induce an immunologically mediated liver injury. Rarely rifampicin may cause hyperbilirubinaemia without enzyme elevation. Here we report a patient with bilateral pulmonary tuberculosis who developed transient severe indirect hyperbilirubinaemia on rifampicin. On review of relevant literature we find that rifampicin can have a biphasic effect on bilirubin, an initial increase in indirect bilirubin and later normalization of bilirubin. We have reported this case because of its rarity in clinical practice. PMID:27190870

  5. Biphasic Effect of Rifampicin on Bilirubin- A Case Report

    PubMed Central

    Seshadri, Mandalam Subramanian

    2016-01-01

    Drug induced hepatitis is a major problem which a physician encounters in his clinical practice. In view of increasing incidence of tuberculosis in our country a large number of infected individuals are started on Antituberculous (ATT) drugs and rifampicin is invariably part of the regimen. One of the major adverse effects of ATT drugs is drug- induced hepatitis which is characterized by elevation of liver enzymes and bilirubin. Hepatotoxicity is usually idiosyncratic or dose-dependent. Rifampicin causes transient elevation of transaminases in 10-20 percent of individuals and this does not warrant dose adjustments of the drug. Rarely rifampicin can lead to severe hepatitis with hyperbilirubinaemia and marked elevations of SGOT and SGPT and in some patients this can be fatal. The exact mechanism of Rifampicin induced hepatotoxicity is not known but it is postulated to be due to idiosyncratic reaction to rifampicin metabolites which may be directly toxic or induce an immunologically mediated liver injury. Rarely rifampicin may cause hyperbilirubinaemia without enzyme elevation. Here we report a patient with bilateral pulmonary tuberculosis who developed transient severe indirect hyperbilirubinaemia on rifampicin. On review of relevant literature we find that rifampicin can have a biphasic effect on bilirubin, an initial increase in indirect bilirubin and later normalization of bilirubin. We have reported this case because of its rarity in clinical practice. PMID:27190870

  6. Solar Irradiation of Bilirubin: An Experiment in Photochemical Oxidation

    ERIC Educational Resources Information Center

    Pillay A. E.; Salih, F. M.

    2006-01-01

    An experiment in photochemical oxidation, which deals with bilirubin, a well-known light-sensitive biological compound that is pedagogically ideal for photochemical experiments at tertiary institutes, is presented. The experiment would benefit students in chemistry who eventually branch out into the health sciences or biochemistry.

  7. The pharmacological features of bilirubin: the question of the century.

    PubMed

    Zahir, Farhana; Rabbani, Gulam; Khan, Rizwan Hasan; Rizvi, Shamim J; Jamal, Mohammad Sarwar; Abuzenadah, Adel M

    2015-09-01

    This review looks at the toxicity and metabolism of bilirubin in terms of its pharmacological potential. Its role has gained importance as more research has revealed the functional significance and interrelationship between the gasotransmitters nitric oxide and carbon monoxide. The biological actions of bilirubin have mostly been characterized in the high micromolar range where toxic effects occur. However, it could also prove to be an important cytoprotector for brain tissue, which is inherently less equipped for antioxidant defense. Plasma bilirubin levels negatively correlate to a number of disease states. Higher levels of bilirubin that are still within the normal range provide a protective effect to the body. The effects on various disorders could be tested using controlled pharmacological upregulation of the molecule with animal models. At nanomolar concentrations, considerable benefits have been obtained when the molecule was delivered pharmacologically under in vitro or in vivo test conditions, particularly in neurodegenerative disorders and after tissue or organ transplantation. The induction of heme oxygenase-1 (HMOX-1) via the activation of nuclear factor erythroid 2-related factor or the use of bile pigments in the harvesting of diseased tissue are novel applications, and like every new therapy, should be used with caution. HMOX-1 is tissue specific, and in exceptional states, such as schizophrenia and specific types of renal disorder, the same therapy may have disastrous effects. PMID:26208389

  8. An overview on alcohol oxidases and their potential applications.

    PubMed

    Goswami, Pranab; Chinnadayyala, Soma Sekhar R; Chakraborty, Mitun; Kumar, Adepu Kiran; Kakoti, Ankana

    2013-05-01

    Alcohol oxidases (Alcohol: O₂ Oxidoreductase; EC 1.1.3.x) are flavoenzymes that catalyze the oxidation of alcohols to the corresponding carbonyl compounds with a concomitant release of hydrogen peroxide. Based on substrate specificity, alcohol oxidases may be categorized broadly into four different groups namely, (a) short chain alcohol oxidase (SCAO), (b) long chain alcohol oxidase (LCAO), (c) aromatic alcohol oxidase (AAO), and (d) secondary alcohol oxidase (SAO). The sources reported for these enzymes are mostly limited to bacteria, yeast, fungi, plant, insect, and mollusks. However, the quantum of reports for each category of enzymes considerably varies across these sources. The enzymes belonging to SCAO and LCAO are intracellular in nature, whereas AAO and SAO are mostly secreted to the medium. SCAO and LCAO are invariably reported as multimeric proteins with very high holoenzyme molecular masses, but the molecular characteristics of these enzymes are yet to be clearly elucidated. One of the striking features of the alcohol oxidases that make them distinct from the widely known alcohol dehydrogenase is the avidly bound cofactor to the redox center of these enzymes that obviate the need to supplement cofactor during the catalytic reaction. These flavin-based redox enzymes have gained enormous importance in the development of various industrial processes and products primarily for developing biosensors and production of various industrially useful carbonyl compounds. The present review provides an overview on alcohol oxidases from different categories focusing research on these oxidases during the last decade along with their potential industrial applications. PMID:23525937

  9. Ultrafast deactivation of bilirubin: dark intermediates and two-photon isomerization.

    PubMed

    Carreira-Blanco, Carlos; Singer, Patrick; Diller, Rolf; Luis Pérez Lustres, J

    2016-03-01

    Bilirubin is a neurotoxic product responsible for neonatal jaundice, which is generally treated by phototherapy. The photoreaction involves ultrafast internal conversion via an elusive intermediate and Z-E isomerization with minor yield (less than 3% in solution). The structure of the intermediate remains unclear. Here, the combination of UV-vis and mid-IR ultrafast transient absorption spectroscopy reports a comprehensive picture of the mechanism and provides essential structural information about the intermediate species. Thus, spectral dynamics during the earliest ps unveils a wavepacket travelling from the Franck-Condon region to the crossing point with a dark state. The latter shows a tighter molecular skeleton than the ground state and decays with 15 ps time constant. Remarkably, the relative contribution of a non-decaying component increases linearly with pump energy, suggesting that Z-E isomerization could also be triggered by two-photon excitation. Implications for the photochemistry of protein-bound open tetrapyrroles are discussed. PMID:26887629

  10. Distant Determination of Bilirubin Distribution in Skin by Multi-Spectral Imaging

    NASA Astrophysics Data System (ADS)

    Saknite, I.; Jakovels, D.; Spigulis, J.

    2011-01-01

    For mapping the bilirubin distribution in bruised skin the multi-spectral imaging technique was employed, which made it possible to observe temporal changes of the bilirubin content in skin photo-types II and III. The obtained results confirm the clinical potential of this technique for skin bilirubin diagnostics.

  11. Twenty-five years of progress in bilirubin metabolism (1952-77).

    PubMed Central

    Billing, B H

    1978-01-01

    This review deals with the development of our understanding of the chemistry of bilirubin and its glucuronide derivatives during the years 1952-1977. It examines the relation between haem metabolism and bilirubin formation and our present knowledge of hepatic transport of bilirubin. The heterogeneity of familial hyperbilirubinaemia is discussed. PMID:98394

  12. Multistimuli-Responsive Bilirubin Nanoparticles for Anticancer Therapy.

    PubMed

    Lee, Yonghyun; Lee, Soyoung; Lee, Dong Yun; Yu, Byeongjun; Miao, Wenjun; Jon, Sangyong

    2016-08-26

    Although stimuli-responsive materials hold potential for use as drug-delivery carriers for treating cancers, their clinical translation has been limited. Ideally, materials used for the purpose should be biocompatible and nontoxic, provide "on-demand" drug release in response to internal or external stimuli, allow large-scale manufacturing, and exhibit intrinsic anticancer efficacy. We present multistimuli-responsive nanoparticles formed from bilirubin, a potent endogenous antioxidant that possesses intrinsic anticancer and anti-inflammatory activity. Exposure of the bilirubin nanoparticles (BRNPs) to either reactive oxygen species (ROS) or external laser light causes rapid disruption of the BRNP nanostructure as a result of a switch in bilirubin solubility, thereby releasing encapsulated drugs. In a xenograft tumor model, BRNPs loaded with the anticancer drug doxorubicin (DOX@BRNPs), when combined with laser irradiation of 650 nm, significantly inhibited tumor growth. This study suggests that BRNPs may be used as a drug-delivery carrier as well as a companion medicine for effectively treating cancers. PMID:27485478

  13. Bilirubin oxidation products (BOXes): synthesis, stability and chemical characteristics

    PubMed Central

    Wurster, W. L.; Pyne-Geithman, G. J.; Peat, I. R.; Clark, J. F.

    2009-01-01

    Summary Bilirubin oxidation products (BOXes) have been a subject of interest in neurosurgery because they are purported to be involved in subarachnoid hemorrhage induced cerebral vasospasm. There is a growing body of information concerning their putative role in vasospasm; however, there is a dearth of information concerning the chemical and biochemical characteristics of BOXes. A clearer understanding of the synthesis, stability and characteristics of BOXes will be important for a better understanding of the role of BOXes post subarachnoid hemorrhage. We used hydrogen peroxide to oxidize bilirubin and produce BOXes. BOXes were extracted and analyzed using conventional methods such as HPLC and mass spectrometry. Characterization of the stability BOXes demonstrates that light can photodegrade BOXes with a t1/2 of up to 10 h depending upon conditions. Mixed isomers of BOXes have an apparent extinction coefficient of ε = 6985, and a λmax of 310 nm. BOXes are produced by the oxidation of bilirubin, yielding a mixture of isomers: 4-methyl-5-oxo-3-vinyl-(1,5-dihydropyrrol-2-ylidene)acetamide (BOX A) and 3-methyl-5-oxo-4-vinyl-(1,5-dihydropyrrol-2-ylidene)acetamide (BOX B). The BOXes are photodegraded by ambient light and can be analyzed spectrophotometrically with their extinction coefficient as well as with HPLC or mass spectrometry. Their small molecular weight and photodegradation may have made them difficult to characterize in previous studies. PMID:18456996

  14. Bilirubin binding with liver cystatin induced structural and functional changes.

    PubMed

    Mustafa, Mir Faisal; Bano, Bilqees

    2014-05-01

    Cysteine proteinases and their inhibitors play a significant role in the proteolytic environment of the cells. Inhibitors of cysteine proteinases regulate the activity of these enzymes helping in checking the degdration activity of cathepsins. The bilirubin secreated by liver cells can bind to cystatin present in the liver resulting in its functional inactivation, which may further lead to the increase in cathepsins level causing liver cirrhosis. In case of some pathophysiological conditions excess bilirubin gets accumulated e.g. in presence of Fasciola hepatica (liver fluke) in mammals and humans, leading to liver cirrhosis and possibly jaundice or normal blockade of bile duct causing increased level of bilirubin in blood. Protease-cystatin imbalance causes disease progression. In the present study, Bilirubin (BR) and liver cystatin interaction was studied to explore the cystatin inactivation and structural alteration. The binding interaction was studied by UV-absorption, FT-IR and fluorescence spectroscopy. The quenching of protein fluorescence confirmed the binding of BR with buffalo liver cystatin (BLC). Stern-Volmer analysis of BR-BLC system indicates the presence of static component in the quenching mechanism and the number of binding sites to be close to 1. The fluorescence data proved that the fluorescence quenching of liver cystatin by BR was the result of BR-cystatin complex formation. FTIR analysis of BR-Cystatin complex revealed change in the secondary structure due to perturbation in the microenvironment further confirmed by the decreased caseinolytic activity of BLC against papain. Fluorescence measurements also revealed quenching of fluorescence and shift in peak at different time intervals and at varying pH values. Photo-illumination of BR-cystatin complex causes change in the surrounding environment of liver cystatin as indicated by red-shift. The binding constant for BR-BLC complex was found to be 9.279 × 10(4) M(-1). The cystatin binding with

  15. Association of bilirubin and malondialdehyde levels with retinopathy in type 2 diabetes mellitus

    PubMed Central

    Dave, Apoorva; Kalra, Pramila; Gowda, B. H. Rakshitha; Krishnaswamy, Malavika

    2015-01-01

    Introduction: Bilirubin as an antioxidant and malondialdehyde (MDA) as an oxidant have been shown to be associated with various complications of type 2 diabetes mellitus (DM). Aims and Objectives: The aim was to measure the levels of serum bilirubin and MDA in type 2 DM patients with and without diabetic retinopathy (DR) and to correlate them with severity of DR. Materials and Methods: A total number of 120 subjects out of which 40 were controls without type 2 DM and the rest 80 were type 2 DM patients were included in the study. Of those 80 diabetics, 44 patients did not have DR and 36 patients had DR. Results: The total bilirubin, direct bilirubin, indirect bilirubin were higher in controls as compared to cases (P = 0.017, 0.033, 0.024). Serum MDA levels were found to be higher in diabetics as compared to controls (P = 0.00). The values of all the three parameters, that is, total bilirubin, direct bilirubin and indirect bilirubin were lower in patients with retinopathy as compared to those without retinopathic changes (P = 0.00, 0.020, and 0.007). Subjects were assigned to quartiles based on serum total bilirubin concentration. The prevalence of DR was significantly lower among persons with the highest bilirubin quartile compared to those with the lowest quartile. The severity of DR was inversely proportional to the total bilirubin levels (P = 0.001). The multiple logistic regression analysis showed total bilirubin to be associated with prevalence of DR (P = 0.035). Conclusions: The levels of total bilirubin were significantly lower in patients with DR and also in the late stages of retinopathy as compared to those without retinopathy and in controls but MDA levels did not show any association with DR. PMID:25932393

  16. Unconjugated Bilirubin and an Increased Proportion of Bilirubin Monoconjugates in the Bile of Patients with Gilbert's Syndrome and Crigler-Najjar Disease

    PubMed Central

    Fevery, Johan; Blanckaert, Norbert; Heirwegh, Karel P. M.; Préaux, Anne-Marie; Berthelot, Pierre

    1977-01-01

    Bilirubin pigments were studied in the bile of 20 normal adults, 25 patients with Gilbert's syndrome, 9 children with Crigler-Najjar disease, and 6 patients with hemolysis, to determine how a deficiency of hepatic bilirubin UDP-glucuronosyltransferase would affect the end products of bilirubin biotransformation. In the bile from patients with Gilbert's syndrome, a striking increase was found in the proportion of bilirubin monoconjugates (48.6±9.8% of total conjugates) relative to that in normal bile (27.2±7.8%). This increase was even more pronounced in children with Crigler-Najjar disease, in whom, even in the most severe cases, glucuronide could always be demonstrated in the bile. Furthermore, unconjugated bilirubin-IXα was unquestionably present in the bile of these children and amounted to 30-57% of their total bilirubin pigments (<1% in the controls). It was not possible to predict from the biliary bilirubin composition whether a child would respond to phenobarbital therapy or not. Bile composition was normal in patients with hemolysis, except when there was associated deficiency of hepatic glucuronosyltransferase. Therefore, the observed alterations were not a simple consequence of unconjugated hyperbilirubinemia. The present findings suggest that Crigler-Najjar disease represents a more pronounced expression than Gilbert's syndrome of a common biochemical defect. Hepatic bilirubin UDP-glucuronosyltransferase deficiency leads to decreased formation of diconjugates with an ensuing increase in the proportion of bilirubin monoconjugates in bile; in the most severe cases, an elevated content of biliary unconjugated bilirubin is also found. PMID:409736

  17. Bilirubin release induced by tumor necrosis factor in combination with galactosamine is toxic to mice.

    PubMed

    Van Molle, W; Libert, C

    2003-08-01

    Application of tumor necrosis factor (TNF) in combination with galactosamine (GalN) in mice causes severe apoptosis of hepatocytes, resulting in complete destruction of the liver. Administration of high levels of unconjugated bilirubin and abnormally high production of unconjugated bilirubin have been reported to cause liver damage and are associated with several human pathologies. Serum alanine aminotransferase as well as total and direct bilirubin levels in mice were determined. Bilirubin levels are shown to significantly increase after a challenge with TNF/GalN in mice. Pretreatment with a heme oxygenase-1 inhibitor significantly prevents this release in bilirubin and offers significant protection against TNF/GalN-induced lethality. A correlation between the release of unconjugated bilirubin and the toxicity accompanied with this release is provided. PMID:12906872

  18. Heme/copper terminal oxidases

    SciTech Connect

    Ferguson-Miller, S.; Babcock, G.T.

    1996-11-01

    Spatially well-organized electron-transfer reactions in a series of membrane-bound redox proteins form the basis for energy conservation in both photosynthesis and respiration. The membrane-bound nature of the electron-transfer processes is critical, as the free energy made available in exergonic redox chemistry is used to generate transmembrane proton concentration and electrostatic potential gradients. These gradients are subsequently used to drive ATP formation, which provides the immediate energy source for constructive cellular processes. The terminal heme/copper oxidases in respiratory electron-transfer chains illustrate a number of the thermodynamic and structural principles that have driven the development of respiration. This class of enzyme reduces dioxygen to water, thus clearing the respiratory system of low-energy electrons so that sustained electron transfer and free-energy transduction can occur. By using dioxygen as the oxidizing substrate, free-energy production per electron through the chain is substantial, owing to the high reduction potential of O{sub 2} (0.815 V at pH 7). 122 refs.

  19. 21 CFR 862.1115 - Urinary bilirubin and its conjugates (nonquantitative) test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1115 Urinary bilirubin and its conjugates (nonquantitative)...

  20. 21 CFR 862.1113 - Bilirubin (total and unbound) in the neonate test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1113 Bilirubin (total and unbound) in the neonate test system....

  1. 21 CFR 862.1115 - Urinary bilirubin and its conjugates (nonquantitative) test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1115 Urinary bilirubin and its conjugates (nonquantitative)...

  2. 21 CFR 862.1113 - Bilirubin (total and unbound) in the neonate test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1113 Bilirubin (total and unbound) in the neonate test system....

  3. 21 CFR 862.1113 - Bilirubin (total and unbound) in the neonate test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1113 Bilirubin (total and unbound) in the neonate test system....

  4. 21 CFR 862.1115 - Urinary bilirubin and its conjugates (nonquantitative) test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1115 Urinary bilirubin and its conjugates (nonquantitative)...

  5. 21 CFR 862.1113 - Bilirubin (total and unbound) in the neonate test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1113 Bilirubin (total and unbound) in the neonate test system....

  6. 21 CFR 862.1115 - Urinary bilirubin and its conjugates (nonquantitative) test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1115 Urinary bilirubin and its conjugates (nonquantitative)...

  7. 21 CFR 862.1113 - Bilirubin (total and unbound) in the neonate test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1113 Bilirubin (total and unbound) in the neonate test system....

  8. 21 CFR 862.1115 - Urinary bilirubin and its conjugates (nonquantitative) test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry Test Systems § 862.1115 Urinary bilirubin and its conjugates (nonquantitative)...

  9. Bilirubin Increases Insulin Sensitivity by Regulating Cholesterol Metabolism, Adipokines and PPARγ Levels.

    PubMed

    Liu, Jinfeng; Dong, Huansheng; Zhang, Yong; Cao, Mingjun; Song, Lili; Pan, Qingjie; Bulmer, Andrew; Adams, David B; Dong, Xiao; Wang, Hongjun

    2015-01-01

    Obesity can cause insulin resistance and type 2 diabetes. Moderate elevations in bilirubin levels have anti-diabetic effects. This study is aimed at determining the mechanisms by which bilirubin treatment reduces obesity and insulin resistance in a diet-induced obesity (DIO) mouse model. DIO mice were treated with bilirubin or vehicle for 14 days. Body weights, plasma glucose, and insulin tolerance tests were performed prior to, immediately, and 7 weeks post-treatment. Serum lipid, leptin, adiponectin, insulin, total and direct bilirubin levels were measured. Expression of factors involved in adipose metabolism including sterol regulatory element-binding protein (SREBP-1), insulin receptor (IR), and PPARγ in liver were measured by RT-PCR and Western blot. Compared to controls, bilirubin-treated mice exhibited reductions in body weight, blood glucose levels, total cholesterol (TC), leptin, total and direct bilirubin, and increases in adiponectin and expression of SREBP-1, IR, and PPARγ mRNA. The improved metabolic control achieved by bilirubin-treated mice was persistent: at two months after treatment termination, bilirubin-treated DIO mice remained insulin sensitive with lower leptin and higher adiponectin levels, together with increased PPARγ expression. These results indicate that bilirubin regulates cholesterol metabolism, adipokines and PPARγ levels, which likely contribute to increased insulin sensitivity and glucose tolerance in DIO mice. PMID:26017184

  10. Unconjugated bilirubin mediates heme oxygenase-1-induced vascular benefits in diabetic mice.

    PubMed

    Liu, Jian; Wang, Li; Tian, Xiao Yu; Liu, Limei; Wong, Wing Tak; Zhang, Yang; Han, Quan-Bin; Ho, Hing-Man; Wang, Nanping; Wong, Siu Ling; Chen, Zhen-Yu; Yu, Jun; Ng, Chi-Fai; Yao, Xiaoqiang; Huang, Yu

    2015-05-01

    Heme oxygenase-1 (HO-1) exerts vasoprotective effects. Such benefit in diabetic vasculopathy, however, remains unclear. We hypothesize that bilirubin mediates HO-1-induced vascular benefits in diabetes. Diabetic db/db mice were treated with hemin (HO-1 inducer) for 2 weeks, and aortas were isolated for functional and molecular assays. Nitric oxide (NO) production was measured in cultured endothelial cells. Hemin treatment augmented endothelium-dependent relaxations (EDRs) and elevated Akt and endothelial NO synthase (eNOS) phosphorylation in db/db mouse aortas, which were reversed by the HO-1 inhibitor SnMP or HO-1 silencing virus. Hemin treatment increased serum bilirubin, and ex vivo bilirubin treatment improved relaxations in diabetic mouse aortas, which was reversed by the Akt inhibitor. Biliverdin reductase silencing virus attenuated the effect of hemin. Chronic bilirubin treatment improved EDRs in db/db mouse aortas. Hemin and bilirubin reversed high glucose-induced reductions in Akt and eNOS phosphorylation and NO production. The effect of hemin but not bilirubin was inhibited by biliverdin reductase silencing virus. Furthermore, bilirubin augmented EDRs in renal arteries from diabetic patients. In summary, HO-1-induced restoration of endothelial function in diabetic mice is most likely mediated by bilirubin, which preserves NO bioavailability through the Akt/eNOS/NO cascade, suggesting bilirubin as a potential therapeutic target for clinical intervention of diabetic vasculopathy. PMID:25475440

  11. CHARACTERISTICS OF POLYPHENOL OXIDASES

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polyphenol oxidase (PPO, EC 1.14.18.1 or EC 1.10.3.1) catalyzes the oxidation of o-diphenols to o-quinones. Highly reactive o-quinones couple with phenolics and specific amino acids on proteins to form the characteristic browning products in many wounded fruits, vegetables, and leaf tissues of plant...

  12. Reproducibility and intragastric variation of duodenogastric reflux using ambulatory gastric bilirubin monitoring.

    PubMed

    Manifold, D K; Anggiansah, A; Marshall, R E; Owen, W J

    2001-01-01

    Duodenogastric reflux has long been considered to be important in the pathogenesis of many gastric disorders that exhibit regional variation within the stomach. Ambulatory gastric bilirubin monitoring is a new technique and, although extensively validated, reproducibility and gastric regional variation have not been specifically addressed. Fourteen patients with symptoms of gastroesophageal reflux and 12 healthy subjects underwent 24-h ambulatory gastric bilirubin monitoring with the bilirubin sensor in the upper stomach. Gastric bilirubin monitoring with two simultaneous bilirubin probes, one in the upper stomach and the other in the antrum, was performed on a separate occasion. Gastric bilirubin exposure in the initial and repeat studies showed a good correlation (R = 0.60, P < 0.01). Gastric bilirubin exposure in the upper stomach and the antrum showed a high degree of correlation (R = 0.90, P < 0.01). In conclusion, reproducible results are obtained with ambulatory gastric bilirubin monitoring and duodenogastric reflux does not exhibit significant regional variation within the stomach. PMID:11270798

  13. Bilirubin content and 4-nitrophenol glucuronosyltransferase activity in Gunn rat liver.

    PubMed

    Celier, C; Foliot, A

    1984-04-01

    The purpose of this study was to determine whether the hepatic content of bilirubin could influence liver 4-nitrophenolglucuronosyltransferase (4-NP-GT) in the Gunn rat. In animals fed on a 45% lipid diet, compared with rats fed on a normal lipid diet (3%), the bilirubin content of the hepatic microsomal fraction decreased and the bilirubin/protein ratio was reduced. 4-NP-GT activities were comparable in both groups. Administration of clofibrate to Gunn rats greatly enhanced the bilirubin content of liver microsomal fraction. Since this treatment raised the microsomal protein content, the bilirubin/protein ratio was not modified. No significant change in 4-NP-GT was noted. After bilirubin perfusion in Gunn and Wistar rats, no change was observed in hepatic monooxygenase activities or in 4-NP-GT, although the bilirubin/protein ratio was dramatically increased in the microsomal fraction. From these results the low activity of liver 4-NP-GT in Gunn rats does not seem directly related to the hepatic content of bilirubin. PMID:6421531

  14. Measurement of serum bilirubin and its mono- and diconjugates: application to patients with hepatobiliary disease.

    PubMed Central

    Scharschmidt, B F; Blanckaert, N; Farina, F A; Kabra, P M; Stafford, B E; Weisiger, R A

    1982-01-01

    A technique has recently been described by Blanckaert and his colleagues that specifically and accurately quantifies unconjugated bilirubin, diconjugated bilirubin, and the C-8 and C-12 isomers of monoconjugated bilirubin. This technique has now been used to determine the distribution pattern of bilirubin and its ester conjugates in 91 sera from 65 patients with hepatobiliary disease, and the results were compared with two conventional diazo assays. Both diazo assays yielded values for total bilirubin concentration that were markedly and unpredictably higher than those obtained by the new technique, and the direct-reacting fraction by diazo assay showed little or no agreement with the fraction of total ester conjugates determined by the new method. Previous studies using the new method had shown that bilirubin conjugates are undetectable in sera from healthy adults or individuals with Gilbert's syndrome, but they were found in 89 of the 91 present patient sera. The fraction of total serum bilirubin represented by C-8 monoconjugates, C-12 monoconjugates, diconjugates, and total ester conjugates was higher in patients with biliary obstruction than in those with parenchymal liver disease, but extensive overlap between groups prevented determination of these conjugated species from being diagnostically useful. Overall, bilirubin ester conjugates in serum consisted of 30% C-8 monoconjugates, 37% C-12 monoconjugates, and 33% diconjugates, while urine contained predominantly diconjugates. PMID:7095558

  15. Relationship of Bilirubin Levels in Infancy to Later Intellectual Development. Interim Report No. 20.

    ERIC Educational Resources Information Center

    Rubin, Rosalyn A.; And Others

    The relationship of bilirubin (a red bile pigment that is sometimes found in the urine and occurs in the blood and tissues in jaundice) in infancy to later intellectual development was investigated in 241 infants with moderately elevated and high bilirubin levels. Ss were administered motor, psycholinguistic, and intelligence tests at age 8…

  16. Hepatocellular uptake of sulfobromophthalein and bilirubin is selectively inhibited by an antibody to the liver plasma membrane sulfobromophthalein/bilirubin binding protein.

    PubMed Central

    Stremmel, W; Berk, P D

    1986-01-01

    To clarify sulfobromophthalein (BSP) and bilirubin uptake mechanisms, isolated rat hepatocytes were incubated with [35S]BSP. The initial uptake velocity (V0), determined from the first, linear portion of the cumulative uptake curve, was saturable (Michaelis constant [Km] = 6.2 +/- 0.5 microM; Vmax = 638 +/- 33 pmol X min-1 per 10(5) hepatocytes), maximal at 37 degrees C and pH 7.4, and competitively inhibited by bilirubin, but not by taurocholate, cholate, or oleate. Preloading with unlabeled BSP led to trans-stimulation of V0. Sodium substitution or pretreatment of hepatocytes with ouabain or metabolic inhibitors had no effect on V0; trypsin reduced V0 by 39% (P less than 0.001). A rabbit antiserum to the rat liver plasma membrane (LPM)-BSP/bilirubin binding protein selectively reduced V0 of 5 microM [35S]BSP and [14C]bilirubin by 41 and 42%, respectively (P less than 0.01); uptakes of [3H]oleate, [3H]cholate and [3H]taurocholate were not affected. Hence, the LPM-BSP/bilirubin binding protein plays a role in the carrier-mediated uptake of BSP and bilirubin by hepatocytes. PMID:3745441

  17. Revalidation and rationale for high pKa values of unconjugated bilirubin

    PubMed Central

    Ostrow, J Donald; Mukerjee, Pasupati

    2007-01-01

    Background Our prior solvent partition analysis, published in 1992, yielded pKa values for unconjugated bilirubin of about 8.1 and 8.4, but these results have been challenged and studies by other methods have suggested pKa values below 5.0. Methods We repeated our published solvent partition studies, using 14C-unconjugated bilirubin highly purified by extraction of residual labeled impurities from CHCl3 into an aqueous buffer, pH 7.0. Partition ratios at six pH values from 5.0 to 9.0 were determined by radioassay and compared with our prior values obtained by diazo assay. Results At pH values ranging from 4.8 to 9.2, stable aqueous/chloroform 14C-partition ratios did not differ significantly from our published partition ratios based on diazo assay. Conclusion These results support the high pKa values of unconjugated bilirubin, above 8.0, derived from our earlier solvent partition study. In both studies, our measurements were based on the rapid analysis of clearly under-saturated solutions of highly-purified bilirubin over a wide pH range, using properly purified and preserved solvents. No previous direct estimate of the aqueous pKa values of unconjugated bilirubin meets all these preconditions. Three theoretical factors acting in combination, each related to the unique, extensive internal H-bonding of the -COOH groups, are proposed to support high pKa values of unconjugated bilirubin in water: a) donation of an H-bond from the -OH moiety of the -COOH group, which is broken on ionization; b) hindered solvation of the -COO- group after ionization; and c) restricted rotation of the -COO- and -COOH groups. Our findings and rationale rebut methodological and theoretical criticisms leveled against our prior work. High pKa values for unconjugated bilirubin dictate that: a) bilirubin diacid, which readily diffuses across membranes and can cause neurotoxicity, is the dominant unbound bilirubin species of unconjugated bilirubin in plasma at physiological pH; b) at the near

  18. Metabolism of bilirubin by human cytochrome P450 2A6

    SciTech Connect

    Abu-Bakar, A'edah; Arthur, Dionne M.; Wikman, Anna S.; Rahnasto, Minna; Juvonen, Risto O.; Vepsäläinen, Jouko; Raunio, Hannu; Ng, Jack C.; Lang, Matti A.

    2012-05-15

    The mouse cytochrome P450 (CYP) 2A5 has recently been shown to function as hepatic “Bilirubin Oxidase” (Abu-Bakar, A., et al., 2011. Toxicol. Appl. Pharmacol. 257, 14–22). To date, no information is available on human CYP isoforms involvement in bilirubin metabolism. In this paper we provide novel evidence for human CYP2A6 metabolising the tetrapyrrole bilirubin. Incubation of bilirubin with recombinant yeast microsomes expressing the CYP2A6 showed that bilirubin inhibited CYP2A6-dependent coumarin 7-hydroxylase activity to almost 100% with an estimated K{sub i} of 2.23 μM. Metabolite screening by a high-performance liquid chromatography/electrospray ionisation mass spectrometry indicated that CYP2A6 oxidised bilirubin to biliverdin and to three other smaller products with m/z values of 301, 315 and 333. Molecular docking analyses indicated that bilirubin and its positively charged intermediate interacted with key amino acid residues at the enzyme's active site. They were stabilised at the site in a conformation favouring biliverdin formation. By contrast, the end product, biliverdin was less fitting to the active site with the critical central methylene bridge distanced from the CYP2A6 haem iron facilitating its release. Furthermore, bilirubin treatment of HepG2 cells increased the CYP2A6 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 CYP2A6 compared to cells treated only with CHX. Collectively, the observations indicate that the CYP2A6 may function as human “Bilirubin Oxidase” where bilirubin is potentially a substrate and a regulator of the enzyme. -- Highlights: ► Human CYP2A6 interacts with bilirubin with a high affinity. ► Bilirubin docking to the CYP2A6 active site is more stable than biliverdin docking. ► Recombinant CYP2A6 microsomes metabolised bilirubin to biliverdin. ► Bilirubin increased the hepatic CYP2

  19. Hepatic bilirubin UDP-glucuronyltransferase in patients with sickle cell anemia.

    PubMed

    Maddrey, W C; Cukier, J O; Maglalang, A C; Boitnott, J K; Odell, G B

    1978-02-01

    In sickle cell anemia the shortened survival of red blood cells presents the liver with an augmented load of bilirubin for hepatic clearance. To determine the effects of this excessive bilirubin load on the microsomal conjugating enzyme, hepatic bilirubin UDP-glucuronyltransferase, levels of this enzyme were measured in liver biopsies from patients with sickle cell anemia and several comparison groups. UDP-glucuronyltransferase activity in 14 patients with sickle cell anemia was 2-fold greater (P less than 0.005) than in 14 nonjaundiced comparison patients without liver disease. The elevated UDP-glucuronyltransferase activity in sickle cell anemia was similar to that found in 10 patients who chronically ingested drugs (barbiturates or estrogens) known to increase UDP-glucuronyltransferase activity. These observations suggest enhanced conjugation of bilirubin in patients with sickle cell anemia may result from substrate (bilirubin) induction of UDP-glycuronyltransferase. PMID:413760

  20. Metabolism of heme and bilirubin in rat and human small intestinal mucosa.

    PubMed Central

    Hartmann, F; Bissell, D M

    1982-01-01

    Formation of heme, bilirubin, and bilirubin conjugates has been examined in mucosal cells isolated from the rat upper small intestine. Intact, viable cells were prepared by enzymatic dissociation using a combined vascular and luminal perfusion and incubated with an isotopically labeled precursor, delta-amino-[2,3-3H]levulinic acid. Labeled heme and bile pigment were formed with kinetics similar to those exhibited by hepatocytes. Moreover, the newly formed bilirubin was converted rapidly to both mono- and diglucuronide conjugates. In addition, cell-free extracts of small intestinal mucosa from rats or humans exhibited a bilirubin-UDP-glucuronyl transferase activity that was qualitatively similar to that present in liver. The data suggest that the small intestinal mucosa normally contributes to bilirubin metabolism. PMID:6806320

  1. Comparison of transcutaneous and total serum bilirubin measurement in Turkish newborns.

    PubMed

    Şimşek, Fatih Mehmet; Narter, Fatma; Ergüven, Müferet

    2014-01-01

    Severe neonatal hyperbilirubinemia can be prevented by screening for neonatal jaundice. Transcutaneous bilirubin (TcB) measurement is a noninvasive method for screening neonates. The aim of this study was to examine the correlation between TcB measurement (using the JM-103 bilirubinometer) and total serum bilirubin (TSB) measurement. To our knowledge, this is the first study evaluating the usefulness of the JM-103 bilirubinometer in Turkish neonates. Two hundred and fifty healthy infants in our well-baby nurseries and follow-up clinic with a gestational age of ≥36 weeks who were ≤15 days old were enrolled in this study. TcB measurements were taken usinng the JM-103; almost simultaneously, TSB was checked using a spectrophotometric method. The mean±SD TSB level was 11.2±4.6 mg/dl (range, 0.9-27.0 mg/dl); 17.2% of cases had TSB>15 mg/dl. There was good correlation between transcutaneous bilirubin and total serum bilirubin measurements (Pearson's correlation coefficient 0.87 for TcB from the forehead, 0.88 for TcB from the sternum; p<0.001). The transcutaneous bilirubin measurement tended to underestimate the value with increasing discrepancy at higher TSB values. The mean difference between serum bilirubin and transcutaneous (from the sternum and forehead) bilirubin values was significantly lower in cases not requiring phototherapy than in those requiring phototherapy [2.6 mg/dl (sternum) vs 4.8 mg/dl, 2.9 mg/dl (forehead) vs. 5.2 mg/dl, respectively; p<0.001] Although the JM-103 bilirubinometer tends to underestimate serum bilirubin, especially in patients with high bilirubin levels, it is a suitable screening tool to identify jaundiced infants that require a serum bilirubin check and may reduce the need for TSB measurements. PMID:26388591

  2. Activity of glucose oxidase functionalized onto magnetic nanoparticles

    PubMed Central

    Kouassi, Gilles K; Irudayaraj, Joseph; McCarty, Gregory

    2005-01-01

    Background Magnetic nanoparticles have been significantly used for coupling with biomolecules, due to their unique properties. Methods Magnetic nanoparticles were synthesized by thermal co-precipitation of ferric and ferrous chloride using two different base solutions. Glucose oxidase was bound to the particles by direct attachment via carbodiimide activation or by thiophene acetylation of magnetic nanoparticles. Transmission electron microscopy was used to characterize the size and structure of the particles while the binding of glucose oxidase to the particles was confirmed using Fourier transform infrared spectroscopy. Results The direct binding of glucose oxidase via carbodiimide activity was found to be more effective, resulting in bound enzyme efficiencies between 94–100% while thiophene acetylation was 66–72% efficient. Kinetic and stability studies showed that the enzyme activity was more preserved upon binding onto the nanoparticles when subjected to thermal and various pH conditions. The overall activity of glucose oxidase was improved when bound to magnetic nanoparticles Conclusion Binding of enzyme onto magnetic nanoparticles via carbodiimide activation is a very efficient method for developing bioconjugates for biological applications PMID:15762994

  3. A Conserved Steroid Binding Site in Cytochrome c Oxidase

    SciTech Connect

    Qin, Ling; Mills, Denise A.; Buhrow, Leann; Hiser, Carrie; Ferguson-Miller, Shelagh

    2010-09-02

    Micromolar concentrations of the bile salt deoxycholate are shown to rescue the activity of an inactive mutant, E101A, in the K proton pathway of Rhodobacter sphaeroides cytochrome c oxidase. A crystal structure of the wild-type enzyme reveals, as predicted, deoxycholate bound with its carboxyl group at the entrance of the K path. Since cholate is a known potent inhibitor of bovine oxidase and is seen in a similar position in the bovine structure, the crystallographically defined, conserved steroid binding site could reveal a regulatory site for steroids or structurally related molecules that act on the essential K proton path.

  4. Influence of Phosphatidylcholine and Calcium on Self-Association and Bile Salt Mixed Micellar Binding of the Natural Bile Pigment, Bilirubin Ditaurate.

    PubMed

    Neubrand, Michael W; Carey, Martin C; Laue, Thomas M

    2015-11-17

    Recently [Neubrand, M. W., et al. (2015) Biochemistry 54, 1542-1557], we determined a concentration-dependent monomer-dimer-tetramer equilibrium in aqueous bilirubin ditaurate (BDT) solutions and explored the nature of high-affinity binding of BDT monomers with monomers and micelles of the common taurine-conjugated bile salts (BS). We now investigate, employing complementary physicochemical methods, including fluorescence emission spectrophotometry and quasi-elastic light scattering spectroscopy, the influence of phosphatidylcholine (PC), the predominant phospholipid of bile and calcium, the major divalent biliary cation, on these self-interactions and heterointeractions. We have used short-chain, lyso and long-chain PC species as models and contrasted our results with those of parallel studies employing unconjugated bilirubin (UCB) as the fully charged dianion. Both bile pigments interacted with the zwitterionic headgroup of short-chain lecithins, forming water-soluble (BDT) and insoluble ion-pair complexes (UCB), respectively. Upon micelle formation, BDT monomers apparently remained at the headgroup mantle of short-chain PCs, but the ion pairs with UCB became internalized within the micelle's hydrophobic core. BDT interacted with the headgroups of unilamellar egg yolk (EY) PC vesicles; however, with the simultaneous addition of CaCl2, a reversible aggregation took place, but not vesicle fusion. With mixed EYPC/BS micelles, BDT became bound to the hydrophilic surface (as with simple BS micelles), and in turn, both BDT and BS bound calcium, but not other divalent cations. The calcium complexation of BDT and BS was enhanced strongly with increases in micellar EYPC, suggesting calcium-mediated cross-bridging of hydrophilic headgroups at the micelle's surface. Therefore, the physicochemical binding of BDT to BS in an artificial bile medium is influenced not only by BS species and concentration but also by long-chain PCs and calcium ions that exert a specific rather

  5. Outward Bound.

    ERIC Educational Resources Information Center

    Outward Bound, Inc., Andover, MA.

    The Outward Bound concept was developed in Germany and Great Britain with the saving of human life as the ultimate goal. Courses are designed to help students discover their true physical and mental limits through development of skills including emergency medical aid, firefighting, search and rescue, mountaineering, and sailing. Five Outward Bound…

  6. Enterohepatic circulation of nonconjugated bilirubin in rats fed with human milk

    SciTech Connect

    Alonso, E.M.; Whitington, P.F.; Whitington, S.H.; Rivard, W.A.; Given, G. )

    1991-03-01

    To test the hypothesis that enhanced intestinal absorption of bilirubin may contribute to prolonged nonconjugated hyperbilirubinemia in human milk-fed infants, we studied a cross-section of 36 healthy infants and mothers. Milk from mothers and serum from infants were collected at 16.3 +/- 2.4 days. Milk was studied for its effect on the absorption of bilirubin labeled with carbon 14 in rats and compared with buffer and iron-fortified infant formula (Similac With Iron). The percentage of a 1 mg bilirubin dose absorbed by the rat was 25.29 +/- 4.0% when it was administered into the duodenum with buffer, 4.67 +/- 2.4% with Similac formula, and 7.7 +/- 2.9% with human milk. Linear regression analysis, using the infant's serum nonconjugated bilirubin level as the dependent variable and the percentage of (14C)bilirubin absorbed by the rat with the corresponding mother's milk as the independent variable, revealed a significant correlation (r = 0.40; p = 0.016). Inspection of the data suggested that absorptive permissiveness correlated closely with infant serum bilirubin values greater than 24 mumol/L (1.4 mg/dl) (r = 0.55; p = 0.007), whereas in those with bilirubin values less than or equal to 24 mumol/L, there was no apparent correlation. Milk was also analyzed for beta-glucuronidase, nonesterified fatty acids, and the ability to inhibit glucuronosyltransferase activity of rat liver microsomes in vitro, none of which correlated with the infant's serum bilirubin. These data support the theory that enhanced intestinal absorption of bilirubin contributes to the jaundice associated with breast-feeding.

  7. Tumour suppressor protein p53 regulates the stress activated bilirubin oxidase cytochrome P450 2A6.

    PubMed

    Hu, Hao; Yu, Ting; Arpiainen, Satu; Lang, Matti A; Hakkola, Jukka; Abu-Bakar, A'edah

    2015-11-15

    Human cytochrome P450 (CYP) 2A6 enzyme has been proposed to play a role in cellular defence against chemical-induced oxidative stress. The encoding gene is regulated by various stress activated transcription factors. This paper demonstrates that p53 is a novel transcriptional regulator of the gene. Sequence analysis of the CYP2A6 promoter revealed six putative p53 binding sites in a 3kb proximate promoter region. The site closest to transcription start site (TSS) is highly homologous with the p53 consensus sequence. Transfection with various stepwise deletions of CYP2A6-5'-Luc constructs--down to -160bp from the TSS--showed p53 responsiveness in p53 overexpressed C3A cells. However, a further deletion from -160 to -74bp, including the putative p53 binding site, totally abolished the p53 responsiveness. Electrophoretic mobility shift assay with a probe containing the putative binding site showed specific binding of p53. A point mutation at the binding site abolished both the binding and responsiveness of the recombinant gene to p53. Up-regulation of the endogenous p53 with benzo[α]pyrene--a well-known p53 activator--increased the expression of the p53 responsive positive control and the CYP2A6-5'-Luc construct containing the intact p53 binding site but not the mutated CYP2A6-5'-Luc construct. Finally, inducibility of the native CYP2A6 gene by benzo[α]pyrene was demonstrated by dose-dependent increases in CYP2A6 mRNA and protein levels along with increased p53 levels in the nucleus. Collectively, the results indicate that p53 protein is a regulator of the CYP2A6 gene in C3A cells and further support the putative cytoprotective role of CYP2A6. PMID:26343999

  8. The alternative oxidases: simple oxidoreductase proteins with complex functions.

    PubMed

    Young, Luke; Shiba, Tomoo; Harada, Shigeharu; Kita, Kiyoshi; Albury, Mary S; Moore, Anthony L

    2013-10-01

    The alternative oxidases are membrane-bound monotopic terminal electron transport proteins found in all plants and in some agrochemically important fungi and parasites including Trypansoma brucei, which is the causative agent of trypanosomiasis. They are integral membrane proteins and reduce oxygen to water in a four electron process. The recent elucidation of the crystal structure of the trypanosomal alternative oxidase at 2.85 Å (1 Å=0.1 nm) has revealed salient structural features necessary for its function. In the present review we compare the primary and secondary ligation spheres of the alternative oxidases with other di-iron carboxylate proteins and propose a mechanism for the reduction of oxygen to water. PMID:24059524

  9. Bilirubin as a Determinant for Altered Neurogenesis, Neuritogenesis, and Synaptogenesis

    PubMed Central

    Fernandes, Adelaide; Falcão, Ana Sofia; Abranches, Elsa; Bekman, Evguenia; Henrique, Domingos; Lanier, Lorene M.; Brites, Dora

    2009-01-01

    Elevated levels of serum unconjugated bilirubin (UCB) in the first weeks of life may lead to long-term neurologic impairment. We previously reported that an early exposure of developing neurons to UCB, in conditions mimicking moderate to severe neonatal jaundice, leads to neuritic atrophy and cell death. Here, we have further analyzed the effect of UCB on nerve cell differentiation and neuronal development, addressing how UCB may affect the viability of undifferentiated neural precursor cells and their fate decisions, as well as the development of hippocampal neurons in terms of dendritic and axonal elongation and branching, the axonal growth cone morphology, and the establishment of dendritic spines and synapses. Our results indicate that UCB reduces the viability of proliferating neural precursors, decreases neurogenesis without affecting astrogliogenesis, and increases cellular dysfunction in differentiating cells. In addition, an early exposure of neurons to UCB decreases the number of dendritic and axonal branches at 3 and 9 days in vitro (DIV), and a higher number of neurons showed a smaller growth cone area. UCB-treated neurons also reveal a decreased density of dendritic spines and synapses at 21 DIV. Such deleterious role of UCB in neuronal differentiation, development, and plasticity may compromise the performance of the brain in later life. PMID:19449315

  10. Crystallization of Mitochondrial Cytochrome Oxidase

    NASA Astrophysics Data System (ADS)

    Ozawa, Takayuki; Tanaka, Masashi; Wakabayashi, Takashi

    1982-12-01

    Cytochrome c oxidase (ferrocytochrome c:oxygen oxidoreductase, EC 1.9.3.1) was purified from beef heart mitochondria. By washing the oxidase with detergent on a hydrophobic interaction column, phospholipids were depleted to the level of 1 mol of cardiolipin per mol of heme a. Hydrophobic impurities and partially denatured oxidase were separated from the intact oxidase on an affinity column with cytochrome c as the specific ligand. The final preparation of the oxidase contained seven distinct polypeptides. The molecular weight of the oxidase was estimated to be 130,000 from its specific heme a and copper content and from the subunit composition. Crystals of the oxidase were obtained by slow removal of the detergent from the buffer in which the oxidase was dissolved. The needle-shaped crystals were 100 μ m in average length and 5 μ m in width, and they strongly polarized visible light. Electron diffraction patterns were obtained with an unstained glutaraldehyde-fixed single crystal by electron microscopy using 1,000-kV electrons. From electron micrographs and the diffraction patterns of the crystal, it was concluded that the crystal is monoclinic in the space group P21, with unit cell dimensions a = 92 angstrom, b = 84 angstrom, and c = 103 angstrom, and α =β 90 degrees, γ = 126 degrees.

  11. Solvation and crystal effects in bilirubin studied by NMR spectroscopy and density functional theory.

    PubMed

    Rohmer, Thierry; Matysik, Jörg; Mark, Franz

    2011-10-27

    The open-chain tetrapyrrole compound bilirubin was investigated in chloroform and dimethyl sulfoxide solutions by liquid-state NMR and as solid by (1)H, (13)C, and (15)N magic-angle spinning (MAS) solid-state NMR spectroscopy. Density functional theory (DFT) calculations were performed to interpret the data, using the B3LYP exchange-correlation functional to optimize geometries and to compute NMR chemical shieldings by the gauge-including atomic orbital method. The dependence of geometries and chemical shieldings on the size of the basis sets was investigated for the reference molecules tetramethylsilane, NH(3), and H(2)O, and for bilirubin as a monomer and in clusters consisting of up to six molecules. In order to assess the intrinsic errors of the B3LYP approximation in calculating NMR shieldings, complete basis set estimates were obtained for the nuclear shielding values of the reference molecules. The experimental liquid-state NMR data of bilirubin are well reproduced by a monomeric bilirubin molecule using the 6-311+G(2d,p) basis set for geometry optimization and for calculating chemical shieldings. To simulate the bilirubin crystal, a hexameric model was required. It was constructed from geometry-optimized monomers using information from the X-ray structure of bilirubin to fix the monomeric entities in space and refined by partial optimization. Combining experimental (1)H-(13)C and (1)H-(15)N NMR correlation spectroscopy and density functional theory, almost complete sets of (1)H, (13)C, and (15)N chemical shift assignments were obtained for both liquid and solid states. It is shown that monomeric bilirubin in chloroform solution is formed by 3-vinyl anti conformers, while bilirubin crystals are formed by 3-vinyl syn conformers. This conformational change leads to characteristic differences between the liquid- and solid-state NMR resonances. PMID:21846145

  12. Association of Serum Bilirubin with SYNTAX Score and Future Cardiovascular Events in Patients Undergoing Coronary Intervention

    PubMed Central

    Chang, Chun-Chin; Hsu, Chien-Yi; Huang, Po-Hsun; Chiang, Chia-Hung; Huang, Shao-Sung; Leu, Hsin-Bang; Huang, Chin-Chou; Chen, Jaw-Wen; Lin, Shing-Jong

    2016-01-01

    Background Bilirubin has emerged as an important endogenous antioxidant molecule, and increasing evidence shows that bilirubin may protect against atherosclerosis. The SYNTAX score has been developed to assess the severity and complexity of coronary artery disease. The aim of this study was to evaluate whether serum bilirubin levels are associated with SYNTAX scores and whether they could be used to predict future cardiovascular events in patients undergoing coronary intervention. Methods Serum bilirubin levels and other blood parameters in patients with at least 12-h fasting states were determined. The primary endpoint was any composite cardiovascular event within 1 year, including death, nonfatal myocardial infarction, and target-vessel revascularization. Results In total, 250 consecutive patients with stable coronary artery disease (mean age 70 ± 13) who had received coronary intervention were enrolled. All study subjects were divided into two groups: group 1 was defined as high SYNTAX score (> 22), and group 2 was defined as low SYNTAX score (≤ 22). Total bilirubin levels were significantly lower in the high SYNTAX score group than in the low SYNTAX score group (0.51 ± 0.22 vs. 0.72 ± 0.29 mg/dl, p < 0.001). By multivariate analysis, serum total bilirubin levels were identified as an independent predictor for high SYNTAX score (adjusted odds ratio: 0.28, 95% confidence interval 0.04-0.42; p = 0.004). Use of the Kaplan-Meier analysis demonstrated a significant difference in 1-year cardiovascular events between high (> 0.8 mg/dl), medium (> 0.5, ≤ 0.8 mg/dl), and low (≤ 0.5 mg/dl) bilirubin levels (log-rank test p = 0.011). Conclusions Serum bilirubin level is associated with SYNTAX score and predicts future cardiovascular events in patients undergoing coronary intervention. PMID:27471354

  13. The origins and kinetics of bilirubin in dogs with hepatobiliary and haemolytic diseases.

    PubMed

    Rothuizen, J; van den Brom, W E; Fevery, J

    1992-05-01

    In 35 dogs with spontaneous hepatobiliary liver disease the kinetics and the sources of bilirubin were quantified. The disorders were extrahepatic bile duct obstruction (n = 4), fulminant hepatitis (n = 2), (sub)acute hepatitis (n = 5), chronic active hepatitis (CAH) with cirrhosis (n = 6), hepatic lymphosarcoma (n = 5), centrizonal necrosis secondary to haemolytic anaemia (n = 6) and other (n = 2). The plasma disappearance of [3H]bilirubin was analyzed with a two-compartment model in all dogs. The ratio early labeled/late labeled bilirubin was determined by measuring the incorporation of [14C]glycine into erythrocyte haem and faecal stercobilin. By introducing this relation in the model analysis the bilirubin production rates from erythrocyte destruction (PE), ineffective erythropoiesis (PI) and hepatic haemoprotein (PL) could be quantified. Total bilirubin turnover was increased in both primary haemolytic disease and most cases of hepatobiliary disease. Erythrocyte survival was reduced in all cases but one. The bilirubin clearance was impaired to 30-50% of the normal value in most cases of hepatobiliary disease and also in primary haemolysis. In dogs with fulminant hepatitis, and cirrhosis with or without CAH, the clearance rates were reduced to values below 15% of normal. In these dogs both an impaired clearance and an increased production were important determinants of hyperbilirubinaemia. In other cases plasma bilirubin was primarily determined by increased production. These clearances and production rates were similar in haemolysis and in many cases of primary hepatobiliary disease. The hepatic haemoprotein turnover was quite variable in all subgroups, ranging from 1-74% of the total bilirubin turnover.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:1506635

  14. Bilirubin as a potential causal factor in type 2 diabetes risk: a Mendelian randomization study

    PubMed Central

    Abbasi, Ali; Deetman, Petronella E.; Corpeleijn, Eva; Gansevoort, Ron T.; Gans, Rijk O.B.; Hillege, Hans L.; van der Harst, Pim; Stolk, Ronald P.; Navis, Gerjan; Alizadeh, Behrooz Z.; Bakker, Stephan J.L.

    2014-01-01

    Circulating bilirubin, a natural antioxidant, is associated with decreased risk of type 2 diabetes (T2D), but the nature of the relationship remains unknown. We performed Mendelian randomization in a prospective cohort of 3,381 participants free of diabetes at baseline (aged 28-75 years; women, 52.6%). We used rs6742078 located in UDP-glucuronosyltransferase (UGT1A1) locus as instrumental variable (IV) to study a potential causal effect of serum total bilirubin on T2D risk. T2D developed in a total of 210 (6.2%) participants during a median follow-up of 7.8 years. In adjusted analyses, rs6742078, which explained 19.5% of bilirubin variation, was strongly associated with total bilirubin (a 0.68-SD increase in bilirubin levels per T allele; P<1×10−122) and was also associated with T2D risk (OR 0.69 [95%CI, 0.54-0.90]; P=0.006). Per 1-SD increase in log-transformed bilirubin levels, we observed a 25% (OR 0.75 [95%CI, 0.62-0.92]; P=0.004) lower risk of T2D. In Mendelian randomization analysis, the causal risk reduction for T2D was estimated to be 42% (causal ORIVestimation per 1-SD increase in log-transformed bilirubin 0.58 [95%CI, 0.39-0.84]; P=0.005), which was comparable to the observational estimate (Durbin-Wu-Hausman chi-square test Pfor difference =0.19). These novel results provide evidence that elevated bilirubin is causally associated with risk of T2D and support its role as a protective determinant. PMID:25368098

  15. Extended mathematical model for "in vivo" quantification of the interaction betweeen atazanavir and bilirubin.

    PubMed

    Lozano, Roberto; Domeque, Nieves; Apesteguia, Alberto-Fermín

    2014-02-01

    The objective of the present work was to conduct an "in vivo" analysis of the atazanavir-bilirubin interaction. We developed a new mathematical approach to PK/PDPK models for competitive interaction based on the Michaelis-Menten equation, which was applied to patients with polymorphisms in the gene for UDP-glucuronosyltransferase 1A1 (UGT1A1). Atazanavir is known to induce concentration-dependent increases in bilirubin plasma levels. Thus, we employed our mathematical model to analyse rises in steady state atazanavir and bilirubin concentrations, ultimately plotting a nomogram for detection of suboptimal atazanavir exposure. Application of our model revealed that an absolute value or a steady state increase in bilirubin falling below 3.8Φ µmol/L (where Φ is a correction factor, =1 for UGT1A1 wild type and ≠1 for UGT1A1 variants) could be used to predict suboptimal atazanavir exposure and treatment failure. Thus, we have successfully established a new mathematical approach for pharmacodynamic-pharmacokinetic modelling of the interaction between atazanavir and bilirubin, as it relates to genetic variants of UGT1A1. Taken together, our findings indicate that bilirubin plasma levels represent a valuable marker of atazanavir exposure. PMID:24243081

  16. Quantitative assessment of the multiple processes responsible for bilirubin homeostasis in health and disease

    PubMed Central

    Levitt, David G; Levitt, Michael D

    2014-01-01

    Serum bilirubin measurements are commonly obtained for the evaluation of ill patients and to screen for liver disease in routine physical exams. An enormous research effort has identified the multiple mechanisms involved in the production and metabolism of conjugated (CB) and unconjugated bilirubin (UB). While the qualitative effects of these mechanisms are well understood, their expected quantitative influence on serum bilirubin homeostasis has received less attention. In this review, each of the steps involved in bilirubin production, metabolism, hepatic cell uptake, and excretion is quantitatively examined. We then attempt to predict the expected effect of normal and defective function on serum UB and CB levels in health and disease states including hemolysis, extra- and intrahepatic cholestasis, hepatocellular diseases (eg, cirrhosis, hepatitis), and various congenital defects in bilirubin conjugation and secretion (eg, Gilbert’s, Dubin–Johnson, Crigler–Najjar, Rotor syndromes). Novel aspects of this review include: 1) quantitative estimates of the free and total UB and CB in the plasma, hepatocyte, and bile; 2) detailed discussion of the important implications of the recently recognized role of the hepatic OATP transporters in the maintenance of CB homeostasis; 3) discussion of the differences between the standard diazo assay versus chromatographic measurement of CB and UB; 4) pharmacokinetic implications of the extremely high-affinity albumin binding of UB; 5) role of the enterohepatic circulation in physiologic jaundice of newborn and fasting hyperbilirubinemia; and 6) insights concerning the clinical interpretation of bilirubin measurements. PMID:25214800

  17. Surface-modified anodic aluminum oxide membrane with hydroxyethyl celluloses as a matrix for bilirubin removal.

    PubMed

    Xue, Maoqiang; Ling, Yisheng; Wu, Guisen; Liu, Xin; Ge, Dongtao; Shi, Wei

    2013-01-01

    Microporous anodic aluminum oxide (AAO) membranes were modified by 3-glycidoxypropyltrimethoxysilane to produce terminal epoxy groups. These were used to covalently link hydroxyethyl celluloses (HEC) to amplify reactive groups of AAO membrane. The hydroxyl groups of HEC-AAO composite membrane were further modified with 1,4-butanediol diglycidyl ether to link arginine as an affinity ligand. The contents of HEC and arginine of arginine-immobilized HEC-AAO membrane were 52.1 and 19.7mg/g membrane, respectively. As biomedical adsorbents, the arginine-immobilized HEC-AAO membranes were tested for bilirubin removal. The non-specific bilirubin adsorption on the unmodified HEC-AAO composite membranes was 0.8mg/g membrane. Higher bilirubin adsorption values, up to 52.6mg/g membrane, were obtained with the arginine-immobilized HEC-AAO membranes. Elution of bilirubin showed desorption ratio was up to 85% using 0.3M NaSCN solution as the desorption agent. Comparisons equilibrium and dynamic capacities showed that dynamic capacities were lower than the equilibrium capacities. In addition, the adsorption mechanism of bilirubin and the effects of temperature, initial concentration of bilirubin, albumin concentration and ionic strength on adsorption were also investigated. PMID:23290920

  18. NADPH oxidase mediates glucolipotoxicity-induced beta cell dysfunction--clinical implications.

    PubMed

    McCarty, Mark F; Barroso-Aranda, Jorge; Contreras, Francisco

    2010-03-01

    An impairment of glucose-stimulated insulin secretion--reflecting decreased glucokinase expression--and a moderate decrease in beta cell mass attributable to increased apoptosis, constitute the key features of beta cell failure in type 2 diabetes. Oxidative stress, provoked by prolonged exposure to excessive levels of glucose and/or fatty acids (glucolipotoxicity), appears to be a key mediator of these defects. Oxidant-provoked JNK activation induces nuclear export of the PDX-1 transcription factor, required for expression of glucokinase and other beta cell proteins. Conversely, increases in cAMP induced by incretin hormones promote the nuclear importation of PDX-1, counteracting the diabetogenic impact of oxidant stress; this may explain the utility of measures that slow dietary carbohydrate absorption for diabetes prevention. The ability of oxidative stress to boost apoptosis in beta cells is poorly understood, but may also entail JNK activation. Recent work establishes a phagocyte-type NADPH oxidase as the chief source of glucotoxicity-mediated oxidative stress in beta cells. Since bilirubin is now known to function physiologically as an inhibitor of NADPH oxidase, and phycocyanobilin (PCB) derived from spirulina likewise can inhibit this enzyme complex, supplemental PCB may have utility in the prevention and control of diabetes, and Gilbert syndrome, associated with chronically elevated free bilirubin, may be associated with decreased diabetes risk. PMID:19576699

  19. Bilirubin scavenges chloramines and inhibits myeloperoxidase-induced protein/lipid oxidation in physiologically relevant hyperbilirubinemic serum.

    PubMed

    Boon, A C; Hawkins, C L; Coombes, J S; Wagner, K H; Bulmer, A C

    2015-09-01

    Hypochlorous acid (HOCl), an oxidant produced by myeloperoxidase (MPO), induces protein and lipid oxidation, which is implicated in the pathogenesis of atherosclerosis. Individuals with mildly elevated bilirubin concentrations (i.e., Gilbert syndrome; GS) are protected from atherosclerosis, cardiovascular disease, and related mortality. We aimed to investigate whether exogenous/endogenous unconjugated bilirubin (UCB), at physiological concentrations, can protect proteins/lipids from oxidation induced by reagent and enzymatically generated HOCl. Serum/plasma samples supplemented with exogenous UCB (≤250µM) were assessed for their susceptibility to HOCl and MPO/H2O2/Cl(-) oxidation, by measuring chloramine, protein carbonyl, and malondialdehyde (MDA) formation. Serum/plasma samples from hyperbilirubinemic Gunn rats and humans with GS were also exposed to MPO/H2O2/Cl(-) to: (1) validate in vitro data and (2) determine the relevance of endogenously elevated UCB in preventing protein and lipid oxidation. Exogenous UCB dose-dependently (P<0.05) inhibited HOCl and MPO/H2O2/Cl(-)-induced chloramine formation. Albumin-bound UCB efficiently and specifically (3.9-125µM; P<0.05) scavenged taurine, glycine, and N-α-acetyllysine chloramines. These results were translated into Gunn rat and GS serum/plasma, which showed significantly (P<0.01) reduced chloramine formation after MPO-induced oxidation. Protein carbonyl and MDA formation was also reduced after MPO oxidation in plasma supplemented with UCB (P<0.05; 25 and 50µM, respectively). Significant inhibition of protein and lipid oxidation was demonstrated within the physiological range of UCB, providing a hypothetical link to protection from atherosclerosis in hyperbilirubinemic individuals. These data demonstrate a novel and physiologically relevant mechanism whereby UCB could inhibit protein and lipid modification by quenching chloramines induced by MPO-induced HOCl. PMID:26057938

  20. NADPH Oxidase and Neurodegeneration

    PubMed Central

    Hernandes, Marina S; Britto, Luiz R G

    2012-01-01

    NADPH oxidase (Nox) is a unique, multi-protein, electron transport system that produces large amounts of superoxide via the reduction of molecular oxygen. Nox-derived reactive oxygen species (ROS) are known to be involved in a variety of physiological processes, including host defense and signal transduction. However, over the past decade, the involvement of (Nox)-dependent oxidative stress in the pathophysiology of several neurodegenerative diseases has been increasingly recognized. ROS produced by Nox proteins contribute to neurodegenerative diseases through distinct mechanisms, such as oxidation of DNA, proteins, lipids, amino acids and metals, in addition to activation of redox-sensitive signaling pathways. In this review, we discuss the recent literature on Nox involvement in neurodegeneration, focusing on Parkinson and Alzheimer diseases. PMID:23730256

  1. Dexmedetomidine Attenuates Bilirubin-Induced Lung Alveolar Epithelial Cell Death In Vitro and In Vivo*

    PubMed Central

    Cui, Jian; Zhao, Hailin; Yi, Bin; Zeng, Jing; Lu, Kaizhi

    2015-01-01

    Objective: To investigate bilirubin-induced lung alveolar epithelial cell injury together with the protection afforded by dexmedetomidine. Design: Prospective, randomized, controlled study. Setting: Research laboratory. Subjects: Sprague Dawley rats. Interventions: Alveolar epithelial A549 cell lines were cultured and received bilirubin (from 0 to 160 μM) to explore the protective pathway of dexmedetomidine on bilirubin-induced alveolar epithelial cell injury assessed by immunochemistry and flow cytometry. Sprague-Dawley rats were subjected to common bile duct ligation surgery to explore the protective effect of dexmedetomidine on hyperbilirubinemia-induced alveolar epithelial cell injury and respiratory failure in comparison with the Sham (subjected to the surgery procedure but without bile duct ligation) or dexmedetomidine control (only received intraperitoneal injection of dexmedetomidine). Measurements and Main Results: In vitro, dexmedetomidine reversed the collapse of mitochondrial membrane potential (Δψm), upregulation of cytochrome C, B cell leukemia 2 associated X protein, and cleaved-caspase 3 and 9 in A549 epithelial cells with bilirubin challenge. Furthermore, dexmedetomidine reversed the arrest of cell cycle and the downregulation of the transforming growth factorβ, phosphorylated mammalian target of rapamycin, and p42/44 mitogen-activated protein kinase induced by bilirubin. In vivo, pulmonary edema and inflammation were found after common bile duct ligation. Bilirubin and Paco2 were significantly increased, and oxygen (Pao2) was significantly decreased in the blood of common bile duct ligation rats from the postsurgery day 7 to day 21 when compared with those in the sham controls, respectively (p < 0.01). Daily intraperitoneal injection of dexmedetomidine significantly alleviated the lung edema and injury and prevented respiratory failure. Conclusion: Our data both in vitro and in vivo demonstrated that dexmedetomidine protected alveolar

  2. Serum total bilirubin levels and coronary heart disease--Causal association or epiphenomenon?

    PubMed

    Kunutsor, Setor K

    2015-12-01

    Observational epidemiological evidence supports a linear inverse and independent association between serum total bilirubin levels and coronary heart disease (CHD) risk, but whether this association is causal remains to be ascertained. A Mendelian randomization approach was employed to test whether serum total bilirubin is causally linked to CHD. The genetic variant rs6742078--well known to specifically modify levels of serum total bilirubin and accounting for up to 20% of the variance in circulating serum total bilirubin levels--was used as an instrumental variable. In pooled analysis of estimates reported from published genome-wide association studies, every copy of the T allele of rs6742078 was associated with 0.42 standard deviation (SD) higher levels of serum total bilirubin (95% confidence interval, 0.40 to 0.43). Based on combined data from the Coronary Artery Disease Genome wide Replication and Meta-analyses and the Coronary Artery Disease (C4D) Genetics Consortium involving a total of 36,763 CHD cases and 76,997 controls, the odds ratio for CHD per copy of the T allele was 1.01 (95% confidence interval, 0.99 to 1.04). The odds ratio of CHD for a 1 SD genetically elevated serum total bilirubin level was 1.03 (95% confidence interval, 0.98 to 1.09). The current findings casts doubt on a strong causal association of serum total bilirubin levels with CHD. The inverse associations demonstrated in observational studies may be driven by biases such as unmeasured confounding and/or reverse causation. However, further research in large-scale consortia is needed. PMID:26408227

  3. Development of a System Model for Non-Invasive Quantification of Bilirubin in Jaundice Patients

    NASA Astrophysics Data System (ADS)

    Alla, Suresh K.

    Neonatal jaundice is a medical condition which occurs in newborns as a result of an imbalance between the production and elimination of bilirubin. Excess bilirubin in the blood stream diffuses into the surrounding tissue leading to a yellowing of the skin. An optical system integrated with a signal processing system is used as a platform to noninvasively quantify bilirubin concentration through the measurement of diffuse skin reflectance. Initial studies have lead to the generation of a clinical analytical model for neonatal jaundice which generates spectral reflectance data for jaundiced skin with varying levels of bilirubin concentration in the tissue. The spectral database built using the clinical analytical model is then used as a test database to validate the signal processing system in real time. This evaluation forms the basis for understanding the translation of this research to human trials. The clinical analytical model and signal processing system have been successful validated on three spectral databases. First spectral database is constructed using a porcine model as a surrogate for neonatal skin tissue. Samples of pig skin were soaked in bilirubin solutions of varying concentrations to simulate jaundice skin conditions. The resulting skins samples were analyzed with our skin reflectance systems producing bilirubin concentration values that show a high correlation (R2 = 0.94) to concentration of the bilirubin solution that each porcine tissue sample is soaked in. The second spectral database is the spectral measurements collected on human volunteers to quantify the different chromophores and other physical properties of the tissue such a Hematocrit, Hemoglobin etc. The third spectral database is the spectral data collected at different time periods from the moment a bruise is induced.

  4. Bilirubin modulated cytokines, growth factors and angiogenesis to improve cutaneous wound healing process in diabetic rats.

    PubMed

    Ram, Mahendra; Singh, Vishakha; Kumawat, Sanjay; Kant, Vinay; Tandan, Surendra Kumar; Kumar, Dinesh

    2016-01-01

    Bilirubin has shown cutaneous wound healing potential in some preliminary studies. Here we hypothesize that bilirubin facilitates wound healing in diabetic rats by modulating important healing factors/candidates and antioxidant parameters in a time-dependent manner. Diabetes was induced in male Wistar rats by streptozotocin. In all diabetic rats wounds were created under pentobarbitone anesthesia. All the rats were divided into two groups, of which one (control) was treated with ointment base and other with bilirubin ointment (0.3%). Wound closer measurement and tissue collection were done on days 3, 7, 14 and 19 post-wounding. The relative expressions of hypoxia inducible factor-1 alpha (HIF-1α), vascular endothelial growth factor (VEGF), stromal cell-derived factor-1 alpha (SDF-1α), transforming growth factor- beta1 (TGF-β1()), tumor necrosis factor-α (TNF-α) and interlukin-10 (IL-10) mRNA and proteins and the mRNA of interlukin-1 beta (IL-1β) and matrix metalloprteinase-9 (MMP-9) were determined in the wound tissues. CD-31 staining and collagen content were evaluated by immunohistochemistry and picrosirius red staining, respectively. Histopathological changes were assessed by H&E staining. The per cent wound closer was significantly higher from day 7 onwards in bilirubin-treated rats. HIF-1α, VEGF, SDF-1α, TGF-β1, IL-10 mRNA and protein levels were significantly higher on days 3, 7 and 14 in bilirubin-treated rats. The mRNA expression and protein level of TNF-α and the mRNA of IL-1β and MMP-9 were progressively and markedly reduced in bilirubin-treated rats. The collagen deposition and formation of blood vessels were greater in bilirubin-treated rats. Bilirubin markedly facilitated cutaneous wound healing in diabetic rats by modulating growth factors, cytokines, neovasculogenesis and collagen contents to the wound site. Topical application of bilirubin ointment might be of great use in cutaneous wound healing in diabetic patients. PMID:26679676

  5. Physiological Antioxidative Network of the Bilirubin System in Aging and Age-Related Diseases

    PubMed Central

    Kim, Sung Young; Park, Sang Chul

    2012-01-01

    Oxidative stress is detrimental to life process and is particularly responsible for aging and age-related diseases. Thus, most organisms are well equipped with a spectrum of biological defense mechanisms against oxidative stress. The major efficient antioxidative mechanism is the glutathione system, operating a redox cycling mechanism for glutathione utilization, which consists of glutathione and its peroxidase and reductase. However, this system is mainly effective for hydrophilic oxidants, while lipophilic oxidants require another scavenging system. Since many age-related pathological conditions are related to lipid peroxidation, especially in association with the aging process, the physiological role of the scavenging system for lipophilic oxidants should be considered. In this regard, the biliverdin to bilirubin conversion pathway, via biliverdin reductase (BVR), is suggested to be another major protective mechanism that scavenges lipophilic oxidants because of the lipophilic nature of bilirubin. The efficiency of this bilirubin system might be potentiated by operation of the intertwined bicyclic systems of the suggested redox metabolic cycle of biliverdin and bilirubin and the interactive control cycle of BVR and heme oxygenase. In order to combat oxidative stress, both antioxidative systems against hydrophilic and lipophilic oxidants are required to work cooperatively. In this regard, the roles of the bilirubin system in aging and age-related diseases are reassessed in this review, and their interacting networks are evaluated. PMID:22457648

  6. Potential Cardiovascular Risk Protection of Bilirubin in End-Stage Renal Disease Patients under Hemodialysis

    PubMed Central

    do Sameiro-Faria, Maria; Kohlova, Michaela; Ribeiro, Sandra; Rocha-Pereira, Petronila; Teixeira, Laetitia; Nascimento, Henrique; Reis, Flávio; Miranda, Vasco; Bronze-da-Rocha, Elsa; Quintanilha, Alexandre; Belo, Luís; Costa, Elísio; Santos-Silva, Alice

    2014-01-01

    We evaluated the potential cardiovascular risk protection of bilirubin in hemodialysis (HD) patients. An enlarged set of studies were evaluated in 191 HD patients, including hematological study, lipid profile, iron metabolism, nutritional, inflammatory markers, and dialysis adequacy. The TA duplication screening in the UDP-glucuronosyltransferase 1 A1 (UGT1A1) promoter region was also performed. The UGT1A1 genotype frequencies in HD patients were 49.2%, 42.4%, and 8.4% for 6/6, 6/7, and 7/7 genotypes, respectively. Although no difference was found in UGT1A1 genotype distribution between the three tertiles of bilirubin, significant differences were found with increasing bilirubin levels, namely, a decrease in platelet, leukocyte, and lymphocyte counts, transferrin, oxidized low-density lipoprotein (ox-LDL), ox-LDL/low-density lipoprotein cholesterol ratio, apolipoprotein (Apo) A, Apo B, and interleukin-6 serum levels and a significant increased concentration of hemoglobin, hematocrit, erythrocyte count, iron, transferrin saturation, Apo A/Apo B ratio, adiponectin, and paraoxonase 1 serum levels. After adjustment for age these results remained significant. Our data suggest that higher bilirubin levels are associated with beneficial effects in HD patients, by improving lipid profile and reducing the inflammatory grade, which might contribute to increase in iron availability. These results suggest a potential cardiovascular risk protection of bilirubin in HD patients. PMID:25276769

  7. Pro-healing effects of bilirubin in open excision wound model in rats.

    PubMed

    Ahanger, Azad A; Leo, Marie D; Gopal, Anu; Kant, Vinay; Tandan, Surendra K; Kumar, Dinesh

    2016-06-01

    Bilirubin, a by-product of heme degradation, has an important role in cellular protection. Therefore, we speculated that bilirubin could be of potential therapeutic value in wound healing. To validate the hypothesis, we used a full-thickness cutaneous wound model in rats. Bilirubin (30 mg/kg) was administered intraperitoneally every day for 9 days. The surface area of the wound was measured on days 0, 2, 4, 7 and 10 after the creation of the wound. The granulation tissue was collected on day 10 post-wounding for analysing various parameters of wound healing. Bilirubin treatment accelerated wound contraction and increased hydroxyproline and glucosamine contents. mRNA expression of pro-inflammatory factors such as intercellular cell adhesion molecule-1 (ICAM-1) and tumour necrosis factor-α (TNF-α) were down-regulated and that of anti-inflammatory cytokine interleukin-10 (IL-10) was up-regulated. The findings suggest that bilirubin could be a new agent for enhancing cutaneous wound healing. PMID:24947136

  8. A Genome-Wide Association Study of Total Bilirubin and Cholelithiasis Risk in Sickle Cell Anemia

    PubMed Central

    Milton, Jacqueline N.; Sebastiani, Paola; Solovieff, Nadia; Hartley, Stephen W.; Bhatnagar, Pallav; Arking, Dan E.; Dworkis, Daniel A.; Casella, James F.; Barron-Casella, Emily; Bean, Christopher J.; Hooper, W. Craig; DeBaun, Michael R.; Garrett, Melanie E.; Soldano, Karen; Telen, Marilyn J.; Ashley-Koch, Allison; Gladwin, Mark T.; Baldwin, Clinton T.; Steinberg, Martin H.; Klings, Elizabeth S.

    2012-01-01

    Serum bilirubin levels have been associated with polymorphisms in the UGT1A1 promoter in normal populations and in patients with hemolytic anemias, including sickle cell anemia. When hemolysis occurs circulating heme increases, leading to elevated bilirubin levels and an increased incidence of cholelithiasis. We performed the first genome-wide association study (GWAS) of bilirubin levels and cholelithiasis risk in a discovery cohort of 1,117 sickle cell anemia patients. We found 15 single nucleotide polymorphisms (SNPs) associated with total bilirubin levels at the genome-wide significance level (p value <5×10−8). SNPs in UGT1A1, UGT1A3, UGT1A6, UGT1A8 and UGT1A10, different isoforms within the UGT1A locus, were identified (most significant rs887829, p = 9.08×10−25). All of these associations were validated in 4 independent sets of sickle cell anemia patients. We tested the association of the 15 SNPs with cholelithiasis in the discovery cohort and found a significant association (most significant p value 1.15×10−4). These results confirm that the UGT1A region is the major regulator of bilirubin metabolism in African Americans with sickle cell anemia, similar to what is observed in other ethnicities. PMID:22558097

  9. Bilirubin Levels and Thrombus Burden in Patients With ST-Segment Elevation Myocardial Infarction.

    PubMed

    Hamur, Hikmet; Duman, Hakan; Bakirci, Eftal Murat; Kucuksu, Zafer; Demirelli, Selami; Kalkan, Kamuran; Degirmenci, Husnu

    2016-07-01

    We investigated whether serum bilirubin level (a marker of heme oxygenase activity) is a predictor of thrombus burden in patients with acute myocardial infarction. Patients (n = 229; male 72.9%; mean age 63 ± 13.4 years) who were admitted with ST-segment elevation myocardial infarction (STEMI) were enrolled. Patients were divided into 2 groups. Group 1 was defined as low thrombus burden and group 2 was defined as high thrombus burden. Patients with high thrombus burden had higher total bilirubin levels (14.4 [4.3-22.9] vs 7.7 [2.4-20.3] µmol/L, P ≤ .001), (0.84 [0.25-1.34] vs 0.45 [0.14-1.19] mg/dL P ≤ .001) and direct bilirubin levels (3.1 [2.1-8.4] vs 1.7 [0.5-6.5] µmol/L, P ≤ .001), (0.18 [0.03-0.49] vs 0.10 [0.03-0.38] mg/dL, P ≤ .001). At multivariate analysis, total bilirubin (odds ratio: 1.05, 95% confidence interval: 1.03-1.08, P ≤ .001) was the independent predictor of high thrombus burden. In conclusion, total bilirubin level is independently associated with high thrombus burden in patients with STEMI. PMID:26339042

  10. Efficacy of Human Adipose Tissue-Derived Stem Cells on Neonatal Bilirubin Encephalopathy in Rats.

    PubMed

    Amini, Naser; Vousooghi, Nasim; Hadjighassem, Mahmoudreza; Bakhtiyari, Mehrdad; Mousavi, Neda; Safakheil, Hosein; Jafari, Leila; Sarveazad, Arash; Yari, Abazar; Ramezani, Sara; Faghihi, Faezeh; Joghataei, Mohammad Taghi

    2016-05-01

    Kernicterus is a neurological syndrome associated with indirect bilirubin accumulation and damages to the basal ganglia, cerebellum and brain stem nuclei particularly the cochlear nucleus. To mimic haemolysis in a rat model such that it was similar to what is observed in a preterm human, we injected phenylhydrazine in 7-day-old rats to induce haemolysis and then infused sulfisoxazole into the same rats at day 9 to block bilirubin binding sites in the albumin. We have investigated the effectiveness of human adiposity-derived stem cells as a therapeutic paradigm for perinatal neuronal repair in a kernicterus animal model. The level of total bilirubin, indirect bilirubin, brain bilirubin and brain iron was significantly increased in the modelling group. There was a significant decreased in all severity levels of the auditory brainstem response test in the two modelling group. Akinesia, bradykinesia and slip were significantly declined in the experience group. Apoptosis in basal ganglia and cerebellum were significantly decreased in the stem cell-treated group in comparison to the vehicle group. All severity levels of the auditory brainstem response tests were significantly decreased in 2-month-old rats. Transplantation results in the substantial alleviation of walking impairment, apoptosis and auditory dysfunction. This study provides important information for the development of therapeutic strategies using human adiposity-derived stem cells in prenatal brain damage to reduce potential sensori motor deficit. PMID:26818600

  11. In vitro and in vivo studies on adlay-derived seed extracts: phenolic profiles, antioxidant activities, serum uric acid suppression, and xanthine oxidase inhibitory effects.

    PubMed

    Zhao, Mouming; Zhu, Dashuai; Sun-Waterhouse, Dongxiao; Su, Guowan; Lin, Lianzhu; Wang, Xiao; Dong, Yi

    2014-08-01

    This study aimed to explore the potential of polished adlay, brown adlay, adlay bran, and adlay hull to prevent and treat hyperuricemia. Brown adlay extract effectively decreased the serum uric acid levels of oxonate-induced hyperuricemic rats. Free and bound phenolic extracts from these materials contained significant amounts of phenolics, with free phenolics dominated by chlorogenic acid and p-coumaric acid while bound phenolics dominated by p-coumaric acid and ferulic acid. Free and bound phenolics of adlay bran exhibited significant xanthine oxidase inhibition activities, 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activities, oxygen radical absorbance capacities, and superoxide radical scavenging activities. Adlay bran phenolics could be effective xanthine oxidase inhibitors and radical scavengers. p-Coumaric acid is a xanthine oxidase inhibitor with strong superoxide radical scavenging activity. However, ferulic acid is a xanthine oxidase inhibitor with weak superoxide radical scavenging activity. Chlorogenic acid is a superoxide radical scavenger with weak xanthine oxidase inhibitory activity. PMID:25029106

  12. Magnetic Resonance Imaging of Bilirubin Encephalopathy: Current Limitations and Future Promise

    PubMed Central

    Wisnowski, Jessica L.; Panigrahy, Ashok; Painter, Michael J.; Watchko, Jon F.

    2014-01-01

    Infants with chronic bilirubin encephalopathy often demonstrate abnormal bilateral, symmetric, high-signal intensity on T2-weighted magnetic resonance imaging of the globus pallidus and subthalamic nucleus, consistent with the neuropathology of kernicterus. Early magnetic resonance imaging of at risk infants, while frequently showing increased T1-signal in these regions, may give false positive findings due to the presence of myelin in these structures. Advanced magnetic resonance imaging including diffusion-weighted imaging, magnetic resonance spectroscopy, and diffusion tensor imaging with tractography may shed new insights into the pathogenesis of bilirubin-induced brain injury and the neural basis of long-term disability in infants and children with chronic bilirubin encephalopathy. PMID:25267277

  13. Neonatal bilirubin management as an implementation example of interdisciplinary continuum of care tools.

    PubMed

    Thornton, Sidney N; Thompson, Bryce S; Millar, Jean A; Eggert, Larry D; Wilcox, Adam B

    2007-01-01

    Management of newborn bilirubin spans the inpatient and outpatient continuum of care. Intermountain Healthcare has developed and implemented a web-based tool for managing bilirubin that follows newborn patients across care settings and providers with a consistent plan of care. The underlying model for the tool is derived from published guidelines. The model divides the time-sensitive data into risk zones and associates each zone with the appropriate order set for follow-up care. The tool integrates Intermountain's Help2 infrastructure for authoring terms, guidelines, and order sets, with alerts, results, and data entry within the context of the care model. Implementation of the bilirubin management tool is shown to improve communication, ease workflow, and improve guideline compliance. Lessons learned from the implementation include recommendations for handling point-of-care laboratory data and managing archival views, which are insightful to health networks managing longitudinal data. PMID:18693932

  14. Role of bilirubin overproduction in revealing Gilbert's syndrome: is dyserythropoiesis an important factor?

    PubMed Central

    Metreau, J M; Yvart, J; Dhumeaux, D; Berthelot, P

    1978-01-01

    Gilbert's syndrome was diagnosed in 37 patients with unconjugated hyperbilirubinaemia without overt haemolysis or structural liver abnormality, who had a marked reduction in hepatic bilirubin UDP-glucuronosyltransferase activity (B-GTA) (as compared with that of 23 normal subjects). No significant correlation existed in these patients between serum bilirubin level and the values of B-GTA, thus suggesting that factors other than a low B-GTA must influence the degree of hyperbilirubinaemia in Gilbert's syndrome. Studies of 51Cr erythrocyte survival and 59Fe kinetics in 10 unselected patients demonstrated slight haemolysis in eight, whereas mild ineffective erythropoiesis was suggested in all from a low 24-hour incorporation of radioactive iron into circulating red cells. This overproduction of bilirubin resulting from mild haemolysis and perhaps dyserythropoiesis might reflect only an extreme degree of the normal situation. It certainly contributes to the hyperbilirubinaemia of Gilbert's syndrome and may play a major role in the manifestation of this condition. PMID:101425

  15. Bilirubin Encephalopathy in a Domestic Shorthair Cat With Increased Osmotic Fragility and Cholangiohepatitis.

    PubMed

    Contreras, E T; Giger, U; Malmberg, J L; Quimby, J M; Schaffer, P A

    2016-05-01

    A 7-month-old female domestic shorthair cat was diagnosed with chronic regenerative hemolytic anemia characterized by increased osmotic fragility of unknown etiology. At 13 months of age, the cat was evaluated for acute collapse. The cat was icteric with severe hyperbilirubinemia but no hematocrit changes. Severe obtundation and lateral recumbency progressed to tetraparesis and loss of proprioception in all 4 limbs, and a cerebellar or brainstem lesion was suspected. Postmortem examination revealed suppurative cholangiohepatitis and acute neuronal necrosis in the nuclei of the brainstem and cerebellum, consistent with bilirubin encephalopathy. This is the first known occurrence of cholangiohepatitis and bilirubin encephalopathy in an adult cat with chronic hemolytic anemia. Although rare, bilirubin encephalopathy should be considered a possible sequela to hyperbilirubinemia in adult patients. It remains unknown whether increased osmotic fragility was related to the cholangiohepatopathy. PMID:26354310

  16. Separation by thin-layer chromatography and structure elucidation of bilirubin conjugates isolated from dog bile.

    PubMed Central

    Heirwegh, K P; Fevery, J; Michiels, R; van Hees, G P; Compernolle, F

    1975-01-01

    1. A system for separation of bile pigments by t.l.c. and for their structure elucidation is presented. Separated bile pigments are characterized by t.l.c. of derived dipyrrolic azopigments. 2. At the tetrapyrrolic stage hydrolysis in strongly alkaline medium followed by t.l.c. demonstrates the presence of bilirubin-IIIalpha, -IXalpha and -XIIIalpha and allows assessment of their relative amounts. 3. Most structural information is derived from analysis of dipyrrolic azopigments. Such derivatives, obtained by treatment of separated bile pigments with diazotized ethyl anthranilate, were separated and purified by t.l.c. Micro methods showed (a) the nature of the dipyrrolic aglycone, (b) the nature of the bonds connecting aglycone to a conjugating group, (c) the ratio of vinyl/isovinyl isomers present in the aglycone and, (d) the nature of the conjugating groups (by suitable derivative formation and t.l.c. with reference to known compounds). 4. In bile of normal dogs at least 20 tetrapyrrolic, diazo-positive bile pigments could be recognized. Except for two pigments the tetrapyrrolic nucleus corresponded predominantly to bilirubin-IXalpha. All conjugated pigments had their conjugating groups connected in ester linkage to the tetrapyrrolic aglycone, Apart from bilirubin-IXalpha, monoconjugates and homogeneous and mixed diconjugates of bilirubin were demonstrated; conjugating groups of major importance were xylose, glucose and glucuronic acid. 5. Bilirubin isomer determination on native bile and isolated bile pigments, and dipyrrole-exchange assays with [14C8]bilirubin indicated (a) that the conjugates pre-exist in bile, and (b) that no significant dipyrrole exchange occurs during isolation of the pigments. PMID:1156357

  17. Sensitizing effect of Z,Z-bilirubin IXα and its photoproducts on enzymes in model solutions

    NASA Astrophysics Data System (ADS)

    Plavskii, V. Yu.; Mostovnikov, V. A.; Tret'yakova, A. I.; Mostovnikova, G. R.

    2008-05-01

    In model systems, we have studied side effects which may be induced by light during phototherapy of hyperbilirubinemia (jaundice) in newborn infants, with the aim of reducing the Z,Z-bilirubin IXα (Z,Z-BR IXα) level. We have shown that the sensitizing effect of Z,Z-BR IXα, localized at strong binding sites of the human serum albumin (HSA) macromolecule, is primarily directed at the amino acid residues of the carrier protein and does not involve the molecules of the enzyme (lactate dehydrogenase (LDH)) present in the buffer solution. The detected photodynamic damage to LDH is due to sensitization by bilirubin photoisomers, characterized by lower HSA association constants and located (in contrast to native Z,Z-BR IXα) on the surface of the HSA protein globule. Based on study of the spectral characteristics of the photoproducts of Z,Z-BR IXα and comparison of their accumulation kinetics in solution and the enzyme photo-inactivation kinetics, we concluded that the determining role in sensitized damage to LDH is played by lumirubin. The photosensitization effect depends on the wavelength of the radiation used for photoconversion of bilirubin. When (at the beginning of exposure) we make sure that identical numbers of photons are absorbed by the pigment in the different spectral ranges, the side effect is minimal for radiation corresponding to the long-wavelength edge of the bilirubin absorption band. We have shown that for a bilirubin/HSA concentration ratio >2 (when some of the pigment molecules are sorbed on the surface of the protein globule), the bilirubin can act as a photosensitizing agent for the enzyme present in solution. We discuss methods for reducing unfavorable side effects of light on the body of newborn infants during phototherapy of hyperbilirubinemia.

  18. Reduction of bilirubin by targeting human heme oxygenase-1 through siRNA.

    PubMed

    Xia, Zhen-Wei; Li, Chun-E; Jin, You-Xin; Shi, Yi; Xu, Li-Qing; Zhong, Wen-Wei; Li, Yun-Zhu; Yu, Shan-Chang; Zhang, Zi-Li

    2007-04-01

    Neonatal hyperbilirubinemia is a common clinical condition caused mainly by the increased production and decreased excretion of bilirubin. Current treatment is aimed at reducing the serum levels of bilirubin. Heme oxygenase-1 (HO-1) is a rate-limiting enzyme that generates bilirubin. In this study we intended to suppress HO-1 using the RNA interference technique. Small interfering RNA (siRNA)-A, -B, and -C were designed based on human HO-1 (hHO-1) mRNA sequences. siRNA was transfected into a human hepatic cell line (HL-7702). hHO-1 transcription and protein levels were then determined. In addition, the inhibitory effect of siRNA on hHO-1 was assessed in cells treated with hemin or transfected with an hHO-1 plasmid. siRNA-C showed the most potent suppressive effect on hHO-1. This inhibition is dose and time dependent. Compared with control, both hemin and hHO-1 plasmids up-regulated hHO-1 expression in HL-7702 cells. However, the up-regulation was significantly attenuated by siRNA-C. Furthermore, the decrease in hHO-1 activity was coincident with the suppression of its transcription. Finally, siRNA-C was shown to reduce hHO-1 enzymatic activity and bilirubin levels. Thus, this study provides a novel therapeutic rationale by blocking bilirubin formation via siRNA for preventing and treating neonatal hyperbilirubinemia and bilirubin encephalopathy at an early clinical stage. PMID:17392485

  19. A Hypothesis for Using Pathway Genetic Load Analysis for Understanding Complex Outcomes in Bilirubin Encephalopathy.

    PubMed

    Riordan, Sean M; Bittel, Douglas C; Le Pichon, Jean-Baptiste; Gazzin, Silvia; Tiribelli, Claudio; Watchko, Jon F; Wennberg, Richard P; Shapiro, Steven M

    2016-01-01

    Genetic-based susceptibility to bilirubin neurotoxicity and chronic bilirubin encephalopathy (kernicterus) is still poorly understood. Neonatal jaundice affects 60-80% of newborns, and considerable effort goes into preventing this relatively benign condition from escalating into the development of kernicterus making the incidence of this potentially devastating condition very rare in more developed countries. The current understanding of the genetic background of kernicterus is largely comprised of mutations related to alterations of bilirubin production, elimination, or both. Less is known about mutations that may predispose or protect against CNS bilirubin neurotoxicity. The lack of a monogenetic source for this risk of bilirubin neurotoxicity suggests that disease progression is dependent upon an overall decrease in the functionality of one or more essential genetically controlled metabolic pathways. In other words, a "load" is placed on key pathways in the form of multiple genetic variants that combine to create a vulnerable phenotype. The idea of epistatic interactions creating a pathway genetic load (PGL) that affects the response to a specific insult has been previously reported as a PGL score. We hypothesize that the PGL score can be used to investigate whether increased susceptibility to bilirubin-induced CNS damage in neonates is due to a mutational load being placed on key genetic pathways important to the central nervous system's response to bilirubin neurotoxicity. We propose a modification of the PGL score method that replaces the use of a canonical pathway with custom gene lists organized into three tiers with descending levels of evidence combined with the utilization of single nucleotide polymorphism (SNP) causality prediction methods. The PGL score has the potential to explain the genetic background of complex bilirubin induced neurological disorders (BIND) such as kernicterus and could be the key to understanding ranges of outcome severity in

  20. A Novel Perspective on the Biology of Bilirubin in Health and Disease.

    PubMed

    Gazzin, Silvia; Vitek, Libor; Watchko, Jon; Shapiro, Steven M; Tiribelli, Claudio

    2016-09-01

    Unconjugated bilirubin (UCB) is known to be one of the most potent endogenous antioxidant substances. While hyperbilirubinemia has long been recognized as an ominous sign of liver dysfunction, recent data strongly indicate that mildly elevated bilirubin (BLB) levels can be protective against an array of diseases associated with increased oxidative stress. These clinical observations are supported by new discoveries relating to the role of BLB in immunosuppression and inhibition of protein phosphorylation, resulting in the modulation of intracellular signaling pathways in vascular biology and cancer, among others. Collectively, the evidence suggests that targeting BLB metabolism could be considered a potential therapeutic approach to ameliorate a variety of conditions. PMID:27515064

  1. A Hypothesis for Using Pathway Genetic Load Analysis for Understanding Complex Outcomes in Bilirubin Encephalopathy

    PubMed Central

    Riordan, Sean M.; Bittel, Douglas C.; Le Pichon, Jean-Baptiste; Gazzin, Silvia; Tiribelli, Claudio; Watchko, Jon F.; Wennberg, Richard P.; Shapiro, Steven M.

    2016-01-01

    Genetic-based susceptibility to bilirubin neurotoxicity and chronic bilirubin encephalopathy (kernicterus) is still poorly understood. Neonatal jaundice affects 60–80% of newborns, and considerable effort goes into preventing this relatively benign condition from escalating into the development of kernicterus making the incidence of this potentially devastating condition very rare in more developed countries. The current understanding of the genetic background of kernicterus is largely comprised of mutations related to alterations of bilirubin production, elimination, or both. Less is known about mutations that may predispose or protect against CNS bilirubin neurotoxicity. The lack of a monogenetic source for this risk of bilirubin neurotoxicity suggests that disease progression is dependent upon an overall decrease in the functionality of one or more essential genetically controlled metabolic pathways. In other words, a “load” is placed on key pathways in the form of multiple genetic variants that combine to create a vulnerable phenotype. The idea of epistatic interactions creating a pathway genetic load (PGL) that affects the response to a specific insult has been previously reported as a PGL score. We hypothesize that the PGL score can be used to investigate whether increased susceptibility to bilirubin-induced CNS damage in neonates is due to a mutational load being placed on key genetic pathways important to the central nervous system's response to bilirubin neurotoxicity. We propose a modification of the PGL score method that replaces the use of a canonical pathway with custom gene lists organized into three tiers with descending levels of evidence combined with the utilization of single nucleotide polymorphism (SNP) causality prediction methods. The PGL score has the potential to explain the genetic background of complex bilirubin induced neurological disorders (BIND) such as kernicterus and could be the key to understanding ranges of outcome severity

  2. Age-dependent pattern of cerebellar susceptibility to bilirubin neurotoxicity in vivo in mice

    PubMed Central

    Bortolussi, Giulia; Baj, Gabriele; Vodret, Simone; Viviani, Giulia; Bittolo, Tamara; Muro, Andrés F.

    2014-01-01

    Neonatal jaundice is caused by high levels of unconjugated bilirubin. It is usually a temporary condition caused by delayed induction of UGT1A1, which conjugates bilirubin in the liver. To reduce bilirubin levels, affected babies are exposed to phototherapy (PT), which converts toxic bilirubin into water-soluble photoisomers that are readily excreted out. However, in some cases uncontrolled hyperbilirubinemia leads to neurotoxicity. To study the mechanisms of bilirubin-induced neurological damage (BIND) in vivo, we generated a mouse model lacking the Ugt1a1 protein and, consequently, mutant mice developed jaundice as early as 36 hours after birth. The mutation was transferred into two genetic backgrounds (C57BL/6 and FVB/NJ). We exposed mutant mice to PT for different periods and analyzed the resulting phenotypes from the molecular, histological and behavioral points of view. Severity of BIND was associated with genetic background, with 50% survival of C57BL/6‑Ugt1−/− mutant mice at postnatal day 5 (P5), and of FVB/NJ-Ugt1−/− mice at P11. Life-long exposure to PT prevented cerebellar architecture alterations and rescued neuronal damage in FVB/NJ-Ugt1−/− but not in C57BL/6-Ugt1−/− mice. Survival of FVB/NJ-Ugt1−/− mice was directly related to the extent of PT treatment. PT treatment of FVB/NJ-Ugt1−/− mice from P0 to P8 did not prevent bilirubin-induced reduction in dendritic arborization and spine density of Purkinje cells. Moreover, PT treatment from P8 to P20 did not rescue BIND accumulated up to P8. However, PT treatment administered in the time-window P0–P15 was sufficient to obtain full rescue of cerebellar damage and motor impairment in FVB/NJ-Ugt1−/− mice. The possibility to modulate the severity of the phenotype by PT makes FVB/NJ-Ugt1−/− mice an excellent and versatile model to study bilirubin neurotoxicity, the role of modifier genes, alternative therapies and cerebellar development during high bilirubin conditions. PMID

  3. Regression approach to non-invasive determination of bilirubin in neonatal blood

    NASA Astrophysics Data System (ADS)

    Lysenko, S. A.; Kugeiko, M. M.

    2012-07-01

    A statistical ensemble of structural and biophysical parameters of neonatal skin was modeled based on experimental data. Diffuse scattering coefficients of the skin in the visible and infrared regions were calculated by applying a Monte-Carlo method to each realization of the ensemble. The potential accuracy of recovering the bilirubin concentration in dermis (which correlates closely with that in blood) was estimated from spatially resolved spectrometric measurements of diffuse scattering. The possibility to determine noninvasively the bilirubin concentration was shown by measurements of diffuse scattering at λ = 460, 500, and 660 nm at three source-detector separations under conditions of total variability of the skin biophysical parameters.

  4. Coupling in cytochrome c oxidase

    PubMed Central

    Kessler, R. J.; Blondin, G. A.; Zande, H. Vande; Haworth, R. A.; Green, D. E.

    1977-01-01

    Cytochrome c oxidase (ferrocytochrome c: oxygen oxidoreductase; EC 1.9.3.1) can be resolved into an electron transfer complex (ETC) and an ionophore transfer complex (ITC). Coupling requires an interaction between the moving electron in the ETC and a moving, positively charged ionophore-cation adduct in the ITC. The duplex character of cytochrome oxidase facilitates this interaction. The ITC mediates cyclical cation transport. It can be replaced as the coupling partner by the combination of valinomycin and nigericin in the presence of K+ when cytochrome oxidase is incorporated into liposomes containing acidic phospholipids or by the combination of lipid cytochrome c and bile acids in an ITC-resolved preparation of the ETC. Respiratory control can be induced by incorporating cytochrome oxidase into vesicles of unfractionated whole mitochondrial lipid. The activity of the ITC is suppressed by such incorporation and this suppression leads to the emergence of respiratory control. The ionophoroproteins of the ITC can be extracted into organic solvents; some 50% of the total protein of cytochrome oxidase is extractable. The release of free ionophore is achieved by tryptic digestion of the ionophoroprotein. Preliminary to this release the ionophoroprotein is degraded to an ionophoropeptide. Electrogenic ionophores, as well as uncoupler, are liberated by such proteolysis. The ITC contains a set of ionophoroproteins imbedded in a matrix of phospholipid. Images PMID:198794

  5. Association between bilirubin and risk of Non-Alcoholic Fatty Liver Disease based on a prospective cohort study.

    PubMed

    Tian, Jianbo; Zhong, Rong; Liu, Cheng; Tang, Yuhan; Gong, Jing; Chang, Jiang; Lou, Jiao; Ke, Juntao; Li, Jiaoyuan; Zhang, Yi; Yang, Yang; Zhu, Ying; Gong, Yajie; Xu, Yanyan; Liu, Peiyi; Yu, Xiao; Xiao, Lin; Du, Min; Yang, Ling; Yuan, Jing; Wang, Youjie; Chen, Weihong; Wei, Sheng; Liang, Yuan; Zhang, Xiaomin; He, Meian; Wu, Tangchun; Yao, Ping; Miao, Xiaoping

    2016-01-01

    The study aimed to assess the association between total, direct, and indirect bilirubin and nonalcoholic fatty live disease (NAFLD) risk given its high prevalence and serious clinical prognosis. Among 27,009 subjects who participated in a healthy screening program from the Dongfeng-Tongji cohort study in 2008, 8189 eligible subjects (aged 35-86 years; males, 43.95%) were ultimately enrolled. The incidence rates of NAFLD in 2013 were compared with respect to baseline bilirubin levels among subjects free of NAFLD, and the effect sizes were estimated by logistic regression analysis. During 5 years follow-up, we observed 1956 cases of newly developed NAFLD with the overall incidence of 23.88%. Direct bilirubin was presented to inversely associate with NAFLD risk. Compared with quartile 1 of direct bilirubin, the multivariable-adjusted ORs (95% CIs) for NAFLD of quartile 2 to 4 were 1.104 (0.867-1.187), 0.843 (0.719-0.989), and 0.768 (0.652-0.905), respectively, P for trend 0.002). Similarly, inverse effects of direct bilirubin on NAFLD incidence were also observed when stratified by sex and BMI. However, no significant associations were found between total, and indirect bilirubin and NAFLD risk. Direct bilirubin reduced NAFLD risk independent of possible confounders among middle-aged and elderly Chinese population, probably based on the endogenous antioxidation of bilirubin. PMID:27484402

  6. Association between bilirubin and risk of Non-Alcoholic Fatty Liver Disease based on a prospective cohort study

    PubMed Central

    Tian, Jianbo; Zhong, Rong; Liu, Cheng; Tang, Yuhan; Gong, Jing; Chang, Jiang; Lou, Jiao; Ke, Juntao; Li, Jiaoyuan; Zhang, Yi; Yang, Yang; Zhu, Ying; Gong, Yajie; Xu, Yanyan; Liu, Peiyi; Yu, Xiao; Xiao, Lin; Du, Min; Yang, Ling; Yuan, Jing; Wang, Youjie; Chen, Weihong; Wei, Sheng; Liang, Yuan; Zhang, Xiaomin; He, Meian; Wu, Tangchun; Yao, Ping; Miao, Xiaoping

    2016-01-01

    The study aimed to assess the association between total, direct, and indirect bilirubin and nonalcoholic fatty live disease (NAFLD) risk given its high prevalence and serious clinical prognosis. Among 27,009 subjects who participated in a healthy screening program from the Dongfeng-Tongji cohort study in 2008, 8189 eligible subjects (aged 35–86 years; males, 43.95%) were ultimately enrolled. The incidence rates of NAFLD in 2013 were compared with respect to baseline bilirubin levels among subjects free of NAFLD, and the effect sizes were estimated by logistic regression analysis. During 5 years follow-up, we observed 1956 cases of newly developed NAFLD with the overall incidence of 23.88%. Direct bilirubin was presented to inversely associate with NAFLD risk. Compared with quartile 1 of direct bilirubin, the multivariable-adjusted ORs (95% CIs) for NAFLD of quartile 2 to 4 were 1.104 (0.867–1.187), 0.843 (0.719–0.989), and 0.768 (0.652–0.905), respectively, P for trend 0.002). Similarly, inverse effects of direct bilirubin on NAFLD incidence were also observed when stratified by sex and BMI. However, no significant associations were found between total, and indirect bilirubin and NAFLD risk. Direct bilirubin reduced NAFLD risk independent of possible confounders among middle-aged and elderly Chinese population, probably based on the endogenous antioxidation of bilirubin. PMID:27484402

  7. Effect of sodium phenobarbital on bilirubin metabolism in an infant with congenital, nonhemolytic, unconjugated hyperbilirubinemia, and kernicterus

    PubMed Central

    Crigler, John F.; Gold, Norman I.

    1969-01-01

    Sodium phenobarbital and various hormones, compounds capable of hepatic enzyme induction, were given to an infant boy with congenital, nonhemolytic, unconjugated, hyperbilirubinemia and severe kernicterus for prolonged periods between the ages of 2 and 25 months to determine their effect on serum bilirubin concentrations. Phenobarbital, 5 mg/day orally, on two occasions decreased serum bilirubin concentrations approximately threefold over a period of 30 days. Withdrawal of phenobarbital after the first study resulted in a gradual (30 days) return of serum bilirubin to pretreatment levels. The lower serum bilirubin concentrations observed when phenobarbital therapy was reinstituted were maintained for 61 days on 2.5 mg/kg per day of the drug. Orally administered L-triiodothyronine, 0.05-0.1 mg/day for 71 days, intramuscular human growth hormone, 1 mg/day for 21 days, and testosterone propionate, 0.1 mg/day for 9 days, did not decrease serum bilirubin levels below lowest control values of 18 mg/100 ml. Bilirubin-3H was administered twice before and once with bilirubin-14C during phenobarbital therapy to study the kinetics of bilirubin metabolism. Results of the first and second control studies and of the bilirubin-3H and bilirubin-14C phenobarbital studies, respectively, were as follows: total body bilirubin pools, 200, 184, 73, and 72 mg; half-lives, 111, 84, 37, and 39 hr; and turnover, 30, 37, 33, and 31 mg/day. The data show that the approximate threefold decrease in serum bilirubin concentration and total body pool resulted from a comparable decrease in bilirubin half-life without a significant change in turnover. In vitro histological (electron microscopy) and enzymological studies of liver obtained by surgical biopsies before and during phenobaribtal administration showed that both the hepatocyte content of agranular endoplasmic reticulum (AER) and the ability of liver homogenate to conjugate p-nitrophenol were significantly increased during phenobarbital

  8. Expression of alternative oxidase in tomato

    SciTech Connect

    Kakefuda, M.; McIntosh, L. )

    1990-05-01

    Tomato fruit ripening is characterized by an increase in ethylene biosynthesis, a burst in respiration (i.e. the climacteric), fruit softening and pigmentation. As whole tomatoes ripened from mature green to red, there was an increase in the alternative oxidase capacity. Aging pink tomato slices for 24 and 48 hrs also showed an increase of alternative oxidase and cytochrome oxidase capacities. Monoclonal antibodies prepared to the Sauromatum guttatum alternative oxidase were used to follow the appearance of alternative oxidase in tomato fruits. There is a corresponding increase in a 36kDa protein with an increase in alternative oxidase capacity. Effects of ethylene and norbornadiene on alternative oxidase capacity were also studied. We are using an alternative oxidase cDNA clone from potato to study the expression of mRNA in ripening and wounded tomatoes to determine if the gene is transcriptionally regulated.

  9. Association between Serum Bilirubin and Acute Intraoperative Hyperglycemia Induced by Prolonged Intermittent Hepatic Inflow Occlusion in Living Liver Donors

    PubMed Central

    Han, Sangbin; Jin, Sang-Man; Ko, Justin Sangwook; Kim, Young Ri; Gwak, Mi Sook; Son, Hee Jeong; Joh, Jae-Won; Kim, Gaab Soo

    2016-01-01

    Background Intermittent hepatic inflow occlusion (IHIO) is associated with acute hyperglycemia during living donor hepatectomy when the ischemia is prolonged. Bilirubin is a potent antioxidant to play an important role for maintaining insulin sensitivity and preventing hyperglycemia. Thus, we aimed to test whether serum bilirubin level is associated with prolonged IHIO-induced intraoperative hyperglycemia. Methods Seventy-five living liver donors who underwent a prolonged IHIO with a >30 minute cumulative ischemia were included. The association between preoperative serum bilirubin concentrations and the risk of intraoperative hyperglycemia (blood glucose concentration >180 mg/dl) was analyzed using binary logistic regression with adjusting for potential confounders including age and steatosis. Results The number of donors who underwent 3, 4, 5, and 6 rounds of IHIO was 41, 22, 7, and 5, respectively. Twenty-nine (35%) donors developed intraoperative hyperglycemia. Total bilirubin concentration was inversely associated with hyperglycemia risk (odds ratio [OR] 0.033, 95% confidence interval [CI] 0.004–0.313, P = 0.003). There was an interaction between age and total bilirubin concentration: the effect of lower serum total bilirubin (≤0.7 mg/dl) on the development of hyperglycemia was greater in older donors (>40 years) than in younger donors (P = 0.0.028 versus P = 0.212). Both conjugated bilirubin (OR 0.001 95% CI 0.001–0.684) and unconjugated bilirubin (OR 0.011 95% CI 0.001–0.246) showed an independent association with hyperglycemia risk. Conclusions Lower preoperative serum bilirubin was associated with greater risk of prolonged IHIO-induced hyperglycemia during living donor hepatectomy particularly in older donors. Thus, more meticulous glycemic management is recommended when prolonged IHIO is necessary for surgical purposes in old living donors with lower serum bilirubin levels. PMID:27367602

  10. Bilirubin measured on a blood gas analyser: a suitable alternative for near-patient assessment of neonatal jaundice?

    PubMed

    Peake, M; Mazzachi, B; Fudge, A; Bais, R

    2001-09-01

    The reliability of a recently released total bilirubin assay for a blood gas analyser was assessed in two Australian hospital laboratories. The instrument computes total bilirubin concentration from multi-wavelength absorbance measurements of undiluted whole blood or plasma. Performance of the Radiometer ABL 735 blood gas analyser bilirubin method (software version 3.6) was compared with a proven Roche diazo method for Hitachi analysers, calibrated using primary standards prepared from NIST SRM 916a bilirubin. Acceptable bilirubin results were found over a wide concentration range for most neonatal samples of whole blood or plasma. For adult specimens, bilirubin results were approximately 10% lower on the blood gas analyser. Within-run imprecision (whole blood) was < 2.5%, between-day imprecision (synthetic controls) < 1.0%, and the bilirubin assay for both whole blood and plasma was linear to 1,000 micromol/L. Using sampling options from 35 microL to 195 microL, bilirubin results differed by less than 3%, with a 95 microL syringe option producing the highest results. We conclude that the Radiometer ABL 735 bilirubin assay is suitable for near-patient assessment of neonatal jaundice using whole blood, thus eliminating the need for sample centrifugation. Verification using laboratory methods can be used when required. A positive correction of approximately 10% is required for adult specimens to conform with Hitachi results (SRM 916a calibration), possibly due to the optical characteristics of the higher proportion of conjugated bilirubin and other substances present in most adult samples. PMID:11587132

  11. 21 CFR 862.1110 - Bilirubin (total or direct) test system.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Bilirubin (total or direct) test system. 862.1110 Section 862.1110 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

  12. 21 CFR 862.1110 - Bilirubin (total or direct) test system.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Bilirubin (total or direct) test system. 862.1110 Section 862.1110 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

  13. 21 CFR 862.1110 - Bilirubin (total or direct) test system.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Bilirubin (total or direct) test system. 862.1110 Section 862.1110 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

  14. 21 CFR 862.1110 - Bilirubin (total or direct) test system.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Bilirubin (total or direct) test system. 862.1110 Section 862.1110 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

  15. 21 CFR 862.1110 - Bilirubin (total or direct) test system.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Bilirubin (total or direct) test system. 862.1110 Section 862.1110 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES CLINICAL CHEMISTRY AND CLINICAL TOXICOLOGY DEVICES Clinical Chemistry...

  16. Bilirubin, platelet activation and heart disease: a missing link to cardiovascular protection in Gilbert's syndrome?

    PubMed

    Kundur, Avinash R; Singh, Indu; Bulmer, Andrew C

    2015-03-01

    Gilbert's syndrome (GS) is a relatively common condition, inducing a benign, non-hemolytic, unconjugated hyperbilirubinemia. Gilbert's Syndrome is associated with mutation in the Uridine Glucuronosyl Transferase 1A1 (UGT1A1) gene promoter, reducing UGT1A1 activity, which normally conjugates bilirubin allowing its elimination from the blood. Individuals with GS demonstrate mildly elevated plasma antioxidant capacity caused by elevated levels of unconjugated bilirubin (UCB), reduced thiols and glutathione. Interestingly, the development of, and risk of mortality from, cardiovascular disease is remarkably reduced in GS individuals. An explanation for this protection may be explained by bilirubin's ability to inhibit multiple processes that induce platelet hyper-reactivity and thrombosis, thus far under-appreciated in the literature. Reactive oxygen species are produced continuously via metabolic processes and have the potential to oxidatively modify proteins and lipids within cell membranes, which may encourage the development of thrombosis and CVDs. Oxidative stress induced platelet hyper-reactivity significantly increases the risk of thrombosis, which can potentially lead to tissue infarction. Here, we discuss the possible mechanisms by which increased antioxidant status might influence platelet function and link this to cardiovascular protection in GS. In summary, this is the first article to discuss the possible role of bilirubin as an anti-thrombotic agent, which inhibits platelet activation and potentially, organ infarction, which could contribute to the reduced mortality rate in mildly hyperbilirbinemic individuals. PMID:25576848

  17. Selective nonenzymatic bilirubin detection in blood samples using a Nafion/Mn-Cu sensor.

    PubMed

    Noh, Hui-Bog; Won, Mi-Sook; Shim, Yoon-Bo

    2014-11-15

    The specific detection of biological organics without the use of an enzyme is challenging, and it is crucial for analytical and clinical chemistry. We report specific nonenzymatic bilirubin detection through the catalytic oxidation of bilirubin molecule on the Nafion/Mn-Cu surface. The catalytic ability, true surface area, morphology, crystallinity, composition, and oxidation state of the sensor surface were assessed using voltammetry, coulometry, XPS, XRD, Brunauer-Emmett-Teller (BET), SEM, EDXS, and TOF-SIMS experiments. The results showed that the surface was composed of microporous Mn-Cu bimetallic crystal in flake shape with a large BET surface area (3.635 m(2)g(-1)), where the surface area and crystallinity mainly affected the sensor performance. Product analysis of the catalytic reaction on the sensor probe revealed a specific two-electron oxidation of dipyrromethane moiety to dipyrromethene in the bilirubin molecule. Experimental variables affecting the analysis of bilirubin were optimized in terms of probe composition, temperature, pH, and potential. At the optimized condition, the dynamic range was between 1.2 μM and 0.42 mM, which yielded the equation of ΔI (μA)=(1.03 ± 0.72)+(457.0 ± 4.03) [C] (mM) with 0.999 of correlation coefficient, and the detection limit was 25.0 ± 1.8 nM (n=5, k=3). The stability test, interference effects, and analysis of real clinical samples, human whole blood and certified serum samples were demonstrated to confirm the reliability of the proposed bilirubin sensor. PMID:24953842

  18. [Quantifying intestino-esophageal reflux with a fiberoptic bilirubin detection probe].

    PubMed

    Stein, H J; Kraemer, S J; Feussner, H; Siewert, J R

    1994-05-01

    Currently available methods to assess reflux of duodenal contents into the esophagus are cumbersome, unphysiologic, and inaccurate. The role of intestino-esophageal reflux has therefore been controversial. We assessed intestino-esophageal reflux using a new system which allows prolonged intraesophageal measurement of bilirubin, the major pigment of bile. Measurements were made with a newly developed fiber-optic sensor electrode connected to a portable data processing unit (BILITEC 2000, Synectics Medical Inc., Sweden). Light absorption was measured at the absorption peak of bilirubin and a reference point. Studies were performed in 9 subjects without esophagitis, 9 subjects with esophagitis and primary reflux disease and 7 subjects with erosive esophagitis after a total or subtotal gastrectomy. The fiberoptic electrode was placed 5 cm above the lower esophageal sphincter. In vitro studies showed linear correlations between absorbance measurements obtained with the BILITEC-unit and known bilirubin and bile acid concentrations, respectively (p < 0.01). Compared to both other groups, light absorption was markedly increased in the subjects who had esophagitis after a total or subtotal gastrectomy (p < 0.05) indicating severe biliary reflux. An increase in bilirubin absorption occurred particularly during the post-prandial and supine periodes (p < 0.01). A Roux-en-Y biliary diversion procedure completely abolished bile reflux in 2 of these patients. These data indicate that ambulatory 24-hour fiberoptic measurement of bilirubin in the esophagus is feasible and allows quantitation of intestino-esophageal reflux. Intestino-esophageal reflux occurs particularly during the postprandial period and the early morning hours in patients who had a previous subtotal or total gastrectomy. PMID:8073796

  19. The Role of Bilirubin in Diabetes, Metabolic Syndrome, and Cardiovascular Diseases

    PubMed Central

    Vítek, Libor

    2012-01-01

    Bilirubin belongs to a phylogenetically old superfamily of tetrapyrrolic compounds, which have multiple biological functions. Although for decades bilirubin was believed to be only a waste product of the heme catabolic pathway at best, and a potentially toxic compound at worst; recent data has convincingly demonstrated that mildly elevated serum bilirubin levels are strongly associated with a lower prevalence of oxidative stress-mediated diseases. Indeed, serum bilirubin has been consistently shown to be negatively correlated to cardiovascular diseases (CVD), as well as to CVD-related diseases and risk factors such as arterial hypertension, diabetes mellitus, metabolic syndrome, and obesity. In addition, the clinical data are strongly supported by evidence arising from both in vitro and in vivo experimental studies. This data not only shows the protective effects of bilirubin per se; but additionally, of other products of the heme catabolic pathway such as biliverdin and carbon monoxide, as well as its key enzymes (heme oxygenase and biliverdin reductase); thus, further underlining the biological impacts of this pathway. In this review, detailed information on the experimental and clinical evidence between the heme catabolic pathway and CVD, and those related diseases such as diabetes, metabolic syndrome, and obesity is provided. All of these pathological conditions represent an important threat to human civilization, being the major killers in developed countries, with a steadily increasing prevalence. Thus, it is extremely important to search for novel markers of these diseases, as well as for novel therapeutic modalities to reverse this unfavorable situation. The heme catabolic pathway seems to fulfill the criteria for both diagnostic purposes as well as for potential therapeutical interventions. PMID:22493581

  20. Protoporphyrinogen Oxidase-Inhibiting Herbicides

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Protoporphyrinogen oxidase-inhibiting herbicides (also referred to as Protox- or PPO-inhibiting herbicides) were commercialized in the 1960s and their market share reached approximately 10% (total herbicide active ingredient output) in the late 1990’s. The wide-spread adoption of glyphosate-resista...

  1. Chromate reduction by rabbit liver aldehyde oxidase

    SciTech Connect

    Banks, R.B.; Cooke, R.T. Jr.

    1986-05-29

    Chromate was reduced during the oxidation of 1-methylnicotinamide chlorine by partially purified rabbit liver aldehyde oxidase. In addition to l-methylnicotinamide, several other electron donor substrates for aldehyde oxidase were able to support the enzymatic chromate reduction. The reduction required the presence of both enzyme and the electron donor substrate. The rate of the chromate reduction was retarded by inhibitors or aldehyde oxidase but was not affected by substrates or inhibitors of xanthine oxidase. These results are consistent with the involvement of aldehyde oxidase in the reduction of chromate by rabbit liver cytosolic enzyme preparations.

  2. The effect of bilirubin on the excitability of mitral cells in the olfactory bulb of the rat

    PubMed Central

    Chen, Xiao-Juan; Zhou, Hui-Qun; Ye, Hai-bo; Li, Chun-Yan; Zhang, Wei-Tian

    2016-01-01

    Olfactory dysfunction is a common clinical phenomenon observed in various liver diseases. Previous studies have shown a correlation between smell disorders and bilirubin levels in patients with hepatic diseases. Bilirubin is a well-known neurotoxin; however, its effect on neurons in the main olfactory bulb (MOB), the first relay in the olfactory system, has not been examined. We investigated the effect of bilirubin (>3 μM) on mitral cells (MCs), the principal output neurons of the MOB. Bilirubin increased the frequency of spontaneous firing and the frequency but not the amplitude of spontaneous excitatory postsynaptic currents (sEPSCs). TTX completely blocked sEPSCs in almost all of the cells tested. Bilirubin activity was partially blocked by N-methyl-D-aspartate (NMDA) and α-amino-3-hydroxy-5-methyl-4-isoxazolepro pionic acid (AMPA) receptor antagonists. Furthermore, we found that bilirubin increased the frequency of intrinsic firing independent of synaptic transmission in MCs. Our findings suggest that bilirubin enhances glutamatergic transmission and strengthens intrinsic firing independent of synaptic transmission, all of which cause hyperexcitability in MCs. Our findings provide the basis for further investigation into the mechanisms underlying olfactory dysfunction that are often observed in patients with severe liver disease. PMID:27611599

  3. The effect of bilirubin on the excitability of mitral cells in the olfactory bulb of the rat.

    PubMed

    Chen, Xiao-Juan; Zhou, Hui-Qun; Ye, Hai-Bo; Li, Chun-Yan; Zhang, Wei-Tian

    2016-01-01

    Olfactory dysfunction is a common clinical phenomenon observed in various liver diseases. Previous studies have shown a correlation between smell disorders and bilirubin levels in patients with hepatic diseases. Bilirubin is a well-known neurotoxin; however, its effect on neurons in the main olfactory bulb (MOB), the first relay in the olfactory system, has not been examined. We investigated the effect of bilirubin (>3 μM) on mitral cells (MCs), the principal output neurons of the MOB. Bilirubin increased the frequency of spontaneous firing and the frequency but not the amplitude of spontaneous excitatory postsynaptic currents (sEPSCs). TTX completely blocked sEPSCs in almost all of the cells tested. Bilirubin activity was partially blocked by N-methyl-D-aspartate (NMDA) and α-amino-3-hydroxy-5-methyl-4-isoxazolepro pionic acid (AMPA) receptor antagonists. Furthermore, we found that bilirubin increased the frequency of intrinsic firing independent of synaptic transmission in MCs. Our findings suggest that bilirubin enhances glutamatergic transmission and strengthens intrinsic firing independent of synaptic transmission, all of which cause hyperexcitability in MCs. Our findings provide the basis for further investigation into the mechanisms underlying olfactory dysfunction that are often observed in patients with severe liver disease. PMID:27611599

  4. Role of brain cytochrome P450 mono-oxygenases in bilirubin oxidation-specific induction and activity.

    PubMed

    Gambaro, Sabrina E; Robert, Maria C; Tiribelli, Claudio; Gazzin, Silvia

    2016-02-01

    In the Crigler-Najjar type I syndrome, the genetic absence of efficient hepatic glucuronidation of unconjugated bilirubin (UCB) by the uridine 5'-diphospho-glucuronosyltransferase1A1 (UGT1A1) enzyme produces the rise of UCB level in blood. Its entry to central nervous system could generate toxicity and neurological damage, and even death. In the past years, a compensatory mechanism to liver glucuronidation has been indicated in the hepatic cytochromes P450 enzymes (Cyps) which are able to oxidize bilirubin. Cyps are expressed also in the central nervous system, the target of bilirubin toxicity, thus making them theoretically important to confer a protective activity toward bilirubin accumulation and neurotoxicity. We therefore investigated the functional induction (mRNA, EROD/MROD) and the ability to oxidize bilirubin of Cyp1A1, 1A2, and 2A3 in primary astrocytes cultures obtained from two rat brain region (cortex: Cx and cerebellum: Cll). We observed that Cyp1A1 was the Cyp isoform more easily induced by beta-naphtoflavone (βNF) in both Cx and Cll astrocytes, but oxidized bilirubin only after uncoupling by 3, 4,3',4'-tetrachlorobiphenyl (TCB). On the contrary, Cyp1A2 was the most active Cyp in bilirubin clearance without uncoupling, but its induction was confined only in Cx cells. Brain Cyp2A3 was not inducible. In conclusion, the exposure of astrocytes to βNF plus TCB significantly enhanced Cyp1A1 mediating bilirubin clearance, improving cell viability in both regions. These results may be a relevant groundwork for the manipulation of brain Cyps as a therapeutic approach in reducing bilirubin-induced neurological damage. PMID:25370011

  5. Bilirubin prevents acute DSS-induced colitis by inhibiting leukocyte infiltration and suppressing upregulation of inducible nitric oxide synthase.

    PubMed

    Zucker, Stephen D; Vogel, Megan E; Kindel, Tammy L; Smith, Darcey L H; Idelman, Gila; Avissar, Uri; Kakarlapudi, Ganesh; Masnovi, Michelle E

    2015-11-15

    Bilirubin is thought to exert anti-inflammatory effects by inhibiting vascular cell adhesion molecule-1 (VCAM-1)-dependent leukocyte migration and by suppressing the expression of inducible nitric oxide synthase (iNOS). As VCAM-1 and iNOS are important mediators of tissue injury in the dextran sodium sulfate (DSS) murine model of inflammatory colitis, we examined whether bilirubin prevents colonic injury in DSS-treated mice. Male C57BL/6 mice were administered 2.5% DSS in the drinking water for 7 days, while simultaneously receiving intraperitoneal injections of bilirubin (30 mg/kg) or potassium phosphate vehicle. Disease activity was monitored, peripheral blood counts and serum nitrate levels were determined, and intestinal specimens were analyzed for histological injury, leukocyte infiltration, and iNOS expression. The effect of bilirubin on IL-5 production by HSB-2 cells and on Jurkat cell transendothelial migration also was determined. DSS-treated mice that simultaneously received bilirubin lost less body weight, had lower serum nitrate levels, and exhibited reduced disease severity than vehicle-treated animals. Concordantly, histopathological analyses revealed that bilirubin-treated mice manifested significantly less colonic injury, including reduced infiltration of eosinophils, lymphocytes, and monocytes, and diminished iNOS expression. Bilirubin administration also was associated with decreased eosinophil and monocyte infiltration into the small intestine, with a corresponding increase in peripheral blood eosinophilia. Bilirubin prevented Jurkat migration but did not alter IL-5 production. In conclusion, bilirubin prevents DSS-induced colitis by inhibiting the migration of leukocytes across the vascular endothelium and by suppressing iNOS expression. PMID:26381705

  6. Photoaffinity labeling of protoporphyrinogen oxidase, the molecular target of diphenylether-type herbicides.

    PubMed

    Camadro, J M; Matringe, M; Thome, F; Brouillet, N; Mornet, R; Labbe, P

    1995-05-01

    Diphenylether-type herbicides are extremely potent inhibitors of protoporphyrinogen oxidase, a membrane-bound enzyme involved in the heme and chlorophyll biosynthesis pathways. Tritiated acifluorfen and a diazoketone derivative of tritiated acifluorfen were specifically bound to a single class of high-affinity binding sites on yeast mitochondrial membranes with apparent dissociation constants of 7 nM and 12.5 nM, respectively. The maximum density of specific binding sites, determined by Scatchard analysis, was 3 pmol.mg-1 protein. Protoporphyrinogen oxidase specific activity was estimated to be 2500 nmol protoporphyrinogen oxidized h-1.mol-1 enzyme. The diazoketone derivative of tritiated acifluorfen was used to specifically photolabel yeast protoporphyrinogen oxidase. The specifically labeled polypeptide in wild-type mitochondrial membranes had an apparent molecular mass of 55 kDa, identical to the molecular mass of the purified enzyme. This photolabeled polypeptide was not detected in a protoporphyrinogen-oxidase-deficient yeast strain, but the membranes contained an equivalent amount of inactive immunoreactive protoporphyrinogen oxidase protein. PMID:7758461

  7. Amyloid-β Peptide Binds to Cytochrome C Oxidase Subunit 1

    PubMed Central

    Hernandez-Zimbron, Luis Fernando; Luna-Muñoz, Jose; Mena, Raul; Vazquez-Ramirez, Ricardo; Kubli-Garfias, Carlos; Cribbs, David H.; Manoutcharian, Karen; Gevorkian, Goar

    2012-01-01

    Extracellular and intraneuronal accumulation of amyloid-beta aggregates has been demonstrated to be involved in the pathogenesis of Alzheimer's disease (AD). However, the precise mechanism of amyloid-beta neurotoxicity is not completely understood. Previous studies suggest that binding of amyloid-beta to a number of macromolecules has deleterious effects on cellular functions. Mitochondria were found to be the target for amyloid-beta, and mitochondrial dysfunction is well documented in AD. In the present study we have shown for the first time that Aβ 1–42 bound to a peptide comprising the amino-terminal region of cytochrome c oxidase subunit 1. Phage clone, selected after screening of a human brain cDNA library expressed on M13 phage and bearing a 61 amino acid fragment of cytochrome c oxidase subunit 1, bound to Aβ 1–42 in ELISA as well as to Aβ aggregates present in AD brain. Aβ 1–42 and cytochrome c oxidase subunit 1 co-immunoprecipitated from mitochondrial fraction of differentiated human neuroblastoma cells. Likewise, molecular dynamics simulation of the cytochrome c oxidase subunit 1 and the Aβ 1–42 peptide complex resulted in a reliable helix-helix interaction, supporting the experimental results. The interaction between Aβ 1–42 and cytochrome c oxidase subunit 1 may explain, in part, the diminished enzymatic activity of respiratory chain complex IV and subsequent neuronal metabolic dysfunction observed in AD. PMID:22927926

  8. Enhancements of the production of bilirubin and the expression of β-globin by carbon monoxide during erythroid differentiation.

    PubMed

    Mu, Anfeng; Li, Ming; Tanaka, Masakazu; Adachi, Yasushi; Tai, Tran Tien; Liem, Pham Hieu; Izawa, Shingo; Furuyama, Kazumichi; Taketani, Shigeru

    2016-05-01

    Heme is degraded by heme oxygenase to form iron, carbon monoxide (CO), and biliverdin. However, information about the catabolism of heme in erythroid cells is limited. In this study, we showed the production and export of bilirubin in murine erythroleukemia (MEL) cells. The production of bilirubin by MEL cells was enhanced when heme synthesis was induced. When mouse bone marrow cells were induced with erythropoietin to differentiate into erythroid cells, the synthesis of bilirubin increased. The expression of β-globin was enhanced by CO at the transcriptional level. These results indicate that constant production of CO from heme regulates erythropoiesis. PMID:27087140

  9. Examination of Cholesterol oxidase attachment to magnetic nanoparticles

    PubMed Central

    Kouassi, Gilles K; Irudayaraj, Joseph; McCarty, Gregory

    2005-01-01

    Magnetic nanoparticles (Fe3O4) were synthesized by thermal co-precipitation of ferric and ferrous chlorides. The sizes and structure of the particles were characterized using transmission electron microscopy (TEM). The size of the particles was in the range between 9.7 and 56.4 nm. Cholesterol oxidase (CHO) was successfully bound to the particles via carbodiimide activation. FTIR spectroscopy was used to confirm the binding of CHO to the particles. The binding efficiency was between 98 and 100% irrespective of the amount of particles used. Kinetic studies of the free and bound CHO revealed that the stability and activity of the enzyme were significantly improved upon binding to the nanoparticles. Furthermore, the bound enzyme exhibited a better tolerance to pH, temperature and substrate concentration. The activation energy for free and bound CHO was 13.6 and 9.3 kJ/mol, respectively. This indicated that the energy barrier of CHO activity was reduced upon binding onto Fe3O4 nanoparticles. The improvements observed in activity, stability, and functionality of CHO resulted from structural and conformational changes of the bound enzyme. The study indicates that the stability and activity of CHO could be enhanced via attachment to magnetic nanoparticles and subsequently will contribute to better uses of this enzyme in various biological and clinical applications. PMID:15661076

  10. Water-soluble adsorbent β-cyclodextrin-grafted polyethyleneimine for removing bilirubin from plasma.

    PubMed

    Wang, Zhi; Wei, Houliang; Jia, Lingyun; Xu, Li; Zou, Chunyi; Xie, Jian

    2012-10-01

    A water-soluble adsorbent was developed for removing bilirubin from the plasma of hyperbilirubinemia patient. The adsorbent was synthesized by grafting β-cyclodextrin (β-CD) to branched polyethyleneimine (PEI) matrix. The resulting β-CD-PEI polymer had an average molecular weight of 163.7 kD, and it contained 56.3 β-CD functional groups. In β-CD-PEI-spiked dialysis, 35.8% of plasma bilirubin was removed, which was higher than that removed by the same concentration of bovine serum albumin. β-CD-PEI also removed aromatic amino acids and bile acids. The results indicated that β-CD-PEI could be an effective adsorbent for blood purification application aiming at the removal of toxins. PMID:22836125

  11. 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) increases bilirubin formation but hampers quantitative hepatic conversion of biliverdin to bilirubin in rats with wild-type AH receptor.

    PubMed

    Niittynen, Marjo; Simanainen, Ulla; Pohjanvirta, Raimo; Sankari, Satu; Tuomisto, Jouni T

    2014-06-01

    In haem degradation, haem oxygenase-1 (HO-1) first cleaves haem to biliverdin, which is reduced to bilirubin by biliverdin IXα reductase (BVR-A). The environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) causes hepatic accumulation of biliverdin in moderately TCDD-resistant line B (Kuopio) rats. Using line B and two TCDD-sensitive rat strains, the present study set out to probe the dose-response and biochemical mechanisms of this accumulation. At 28 days after exposure to 3-300 μg/kg TCDD in line B rats, already the lowest dose of TCDD tested, 3 μg/kg, affected serum bilirubin conjugates, and after doses ≥100 μg/kg, the liver content of bilirubin, biliverdin and their conjugates (collectively 'bile pigments') as well as HO-1 was elevated. BVR-A activity and serum bile acids were increased only by the doses of 100 and 300 μg/kg TCDD, respectively. Biliverdin conjugates correlated best with biliverdin suggesting it to be their immediate precursor. TCDD (100 μg/kg, 10 days) increased hepatic bilirubin and biliverdin levels also in TCDD-sensitive Long-Evans (Turku/AB; L-E) rats. Hepatic bilirubin and bile acids, but not biliverdin, were increased in feed-restricted L-E control rats. In TCDD-sensitive line C (Kuopio) rats, 10 μg/kg of TCDD increased the body-weight-normalized biliary excretion of bilirubin. Altogether, the results suggest that at acutely toxic doses, TCDD induces the formation of bilirubin in rats. However, concurrently, TCDD seems to hamper the quantitative conversion of biliverdin to bilirubin in line B and L-E rats' liver. Biliverdin conjugates are most likely formed as secondary products of biliverdin. PMID:24418412

  12. Urate oxidase: primary structure and evolutionary implications.

    PubMed Central

    Wu, X W; Lee, C C; Muzny, D M; Caskey, C T

    1989-01-01

    Urate oxidase, or uricase (EC 1.7.3.3), is a peroxisomal enzyme that catalyzes the oxidation of uric acid to allantoin in most mammals. In humans and certain other primates, however, the enzyme has been lost by some unknown mechanism. To identify the molecular basis for this loss, urate oxidase cDNA clones were isolated from pig, mouse, and baboon, and their DNA sequences were determined. The mouse urate oxidase open reading frame encodes a 303-amino acid polypeptide, while the pig and baboon urate oxidase cDNAs encode a 304-amino acid polypeptide due to a single codon deletion/insertion event. The authenticity of this single additional codon was confirmed by sequencing the mouse and pig genomic copies of the gene. The urate oxidase sequence contains a domain similar to the type 2 copper binding motif found in other copper binding proteins, suggesting that the copper ion in urate oxidase is coordinated as a type 2 structure. Based upon a comparison of the NH2-terminal peptide and deduced sequences, we propose that the maturation of pig urate oxidase involves the posttranslational cleavage of a six-amino acid peptide. Two nonsense mutations were found in the human urate oxidase gene, which confirms, at the molecular level, that the urate oxidase gene in humans is nonfunctional. The sequence comparisons favor the hypothesis that the loss of urate oxidase in humans is due to a sudden mutational event rather than a progressive mutational process. Images PMID:2594778

  13. CAR and PXR agonists stimulate hepatic bile acid and bilirubin detoxification and elimination pathways in mice.

    PubMed

    Wagner, Martin; Halilbasic, Emina; Marschall, Hanns-Ulrich; Zollner, Gernot; Fickert, Peter; Langner, Cord; Zatloukal, Kurt; Denk, Helmut; Trauner, Michael

    2005-08-01

    Induction of hepatic phase I/II detoxification enzymes and alternative excretory pumps may limit hepatocellular accumulation of toxic biliary compounds in cholestasis. Because the nuclear xenobiotic receptors constitutive androstane receptor (CAR) and pregnane X receptor (PXR) regulate involved enzymes and transporters, we aimed to induce adaptive alternative pathways with different CAR and PXR agonists in vivo. Mice were treated with the CAR agonists phenobarbital and 1,4-bis-[2-(3,5-dichlorpyridyloxy)]benzene, as well as the PXR agonists atorvastatin and pregnenolone-16alpha-carbonitrile. Hepatic bile acid and bilirubin-metabolizing/detoxifying enzymes (Cyp2b10, Cyp3a11, Ugt1a1, Sult2a1), their regulatory nuclear receptors (CAR, PXR, farnesoid X receptor), and bile acid/organic anion and lipid transporters (Ntcp, Oatp1,2,4, Bsep, Mrp2-4, Mdr2, Abcg5/8, Asbt) in the liver and kidney were analyzed via reverse-transcriptase polymerase chain reaction and Western blotting. Potential functional relevance was tested in common bile duct ligation (CBDL). CAR agonists induced Mrp2-4 and Oatp2; PXR agonists induced only Mrp3 and Oatp2. Both PXR and CAR agonists profoundly stimulated bile acid-hydroxylating/detoxifying enzymes Cyp3a11 and Cyp2b10. In addition, CAR agonists upregulated bile acid-sulfating Sult2a1 and bilirubin-glucuronidating Ugt1a1. These changes were accompanied by reduced serum levels of bilirubin and bile acids in healthy and CBDL mice and by increased levels of polyhydroxylated bile acids in serum and urine of cholestatic mice. Atorvastatin significantly increased Oatp2, Mdr2, and Asbt, while other transporters and enzymes were moderately affected. In conclusion, administration of specific CAR or PXR ligands results in coordinated stimulation of major hepatic bile acid/bilirubin metabolizing and detoxifying enzymes and hepatic key alternative efflux systems, effects that are predicted to counteract cholestasis. PMID:15986414

  14. Direct Antioxidant Properties of Bilirubin and Biliverdin. Is there a Role for Biliverdin Reductase?

    PubMed Central

    Jansen, Thomas; Daiber, Andreas

    2012-01-01

    Reactive oxygen species (ROS) and signaling events are involved in the pathogenesis of endothelial dysfunction and represent a major contribution to vascular regulation. Molecular signaling is highly dependent on ROS. But depending on the amount of ROS production it might have toxic or protective effects. Despite a large number of negative outcomes in large clinical trials (e.g., HOPE, HOPE-TOO), antioxidant molecules and agents are important players to influence the critical balance between production and elimination of reactive oxygen and nitrogen species. However, chronic systemic antioxidant therapy lacks clinical efficacy, probably by interfering with important physiological redox signaling pathways. Therefore, it may be a much more promising attempt to induce intrinsic antioxidant pathways in order to increase the antioxidants not systemically but at the place of oxidative stress and complications. Among others, heme oxygenase (HO) has been shown to be important for attenuating the overall production of ROS in a broad range of disease states through its ability to degrade heme and to produce carbon monoxide and biliverdin/bilirubin. With the present review we would like to highlight the important antioxidant role of the HO system and especially discuss the contribution of the biliverdin, bilirubin, and biliverdin reductase (BVR) to these beneficial effects. The BVR was reported to confer an antioxidant redox amplification cycle by which low, physiological bilirubin concentrations confer potent antioxidant protection via recycling of biliverdin from oxidized bilirubin by the BVR, linking this sink for oxidants to the NADPH pool. To date the existence and role of this antioxidant redox cycle is still under debate and we present and discuss the pros and cons as well as our own findings on this topic. PMID:22438843

  15. Association between Serum Bilirubin and Estimated Glomerular Filtration Rate among Diabetic Patients

    PubMed Central

    Katoh, Takeaki; Kawamoto, Ryuichi; Kohara, Katsuhiko; Miki, Tetsuro

    2015-01-01

    The subjects comprised 230 men aged 77 ± 10 (range: 50–100) years and 279 women aged 81 ± 10 (50–101) years that visited the medical department. We examined the relationship between increased serum bilirubin and renal function evaluated by estimated glomerular filtration rate (eGFR) using CKD-EPI equations modified by a Japanese coefficient. Compared with the fourth quartile in serum bilirubin (1.01–1.97 mg/dL), the nonadjusted, age and gender-adjusted, and multivariate-adjusted odds ratios {95% confidence interval (CI)} of eGFR <60 mL/min/1.73 m2 for the first quartile in serum bilirubin (0.13–0.50 mg/dL) were 2.08 (1.25–3.44), 1.82 (1.07–3.09), and 1.53 (0.83–2.81), respectively. Moreover, compared with the fourth quartile, nonadjusted, age and gender-adjusted, and multivariate-adjusted odds ratios (95% CI) of eGFR <45 mL/min/1.73 m2 for the first quartile were 3.50 (1.95–6.23), 3.12 (1.72–5.65), and 3.53 (1.71–7.26), respectively. The data were further stratified by gender, age, medication (antihypertensive, antidyslipidemic, and antidiabetic agents), and prevalence of cardiovascular disease (CVD). The standardized coefficients for eGFR were significant in all the subgroups other than the prevalence of CVD, and there were significant interactions between the two groups regarding CVD. Our data demonstrated an independent positive association between serum bilirubin and eGFR among diabetic patients.

  16. Somatostatin inhibits the effect of secretin on bile flow and on hepatic bilirubin transport in the rat.

    PubMed Central

    Ricci, G L; Fevery, J

    1989-01-01

    Increasing amounts of porcine secretion (0.05 to 2.00 clinical units/h/100 g body wt) given to rats during a continuous infusion of bilirubin, increased bile flow and the apparent maximal biliary excretion of bilirubin ('Tm'). This increment was caused by an enhanced biliary output of bilirubin monoconjugates. The effect was dose dependent but maximal at a secretin infusion of 0.80 CU. Somatostatin 0.2 and 0.8 microgram/h/100 g body wt caused a dose related inhibition of the hepatic effects of secretin both on bile flow and on biliary output of bilirubin conjugates. As secretin elicits the release of somatostatin, a feed-back system could be envisaged whereby the somatostatin released stops the effects of secretin. PMID:2572517

  17. Bilirubin induces auditory neuropathy in neonatal guinea pigs via auditory nerve fiber damage.

    PubMed

    Ye, Hai-Bo; Shi, Hai-Bo; Wang, Jian; Ding, Da-Lian; Yu, Dong-Zhen; Chen, Zheng-Nong; Li, Chun-Yan; Zhang, Wei-Tian; Yin, Shan-Kai

    2012-11-01

    Bilirubin can cause temporary or permanent sensorineural deafness in newborn babies with hyperbilirubinemia. However, the underlying targets and physiological effects of bilirubin-induced damage in the peripheral auditory system are unclear. Using cochlear functional assays and electron microscopy imaging of the inner ear in neonatal guinea pigs, we show here that bilirubin exposure resulted in threshold elevation in both compound action potential (CAP) and auditory brainstem response (ABR), which was apparent at 1 hr and peaked 8 hr after drug administration. The threshold elevation was associated with delayed wave latencies and elongated interwave intervals in ABR and CAP. At 72 hr postinjection, these measures returned to control levels, except for the CAP amplitude. Cochlear microphonics remained unchanged during the experiment. Morphological abnormalities were consistent with the electrophysiological dysfunction, revealing fewer auditory nerve fibers (ANFs) in the basal turn, myelin sheath lesions of spiral ganglion neurons (SGNs) and ANFs, and loss of type 1 afferent endings beneath inner hair cells (IHCs) without loss of hair cells at 8 hr posttreatment. Similar to the electrophysiological findings, morphological changes were mostly reversed 10 days after treatment, except for the ANF reduction in the basal turn. These results suggest that hyperbilirubinemia in neonatal guinea pigs impaired auditory peripheral neuromechanisms that targeted mainly the IHC synapses and the myelin sheath of SGNs and their fibers. Our observations indicate a potential connection between hyperbilirubinemia and auditory neuropathy. PMID:22847875

  18. Interference of Bilirubin in the Determination of Magnesium with Methyl Thymol Blue

    PubMed Central

    Maksinovic, Rada; Ketin, Sonja; Biocanin, Rade

    2015-01-01

    Introduction: Jaundice is a disease named for the yellow color of the skin. This color is the result of elevated levels of bilirubin in the blood serum. In Roma from Krusevac region in the last few years have seen the emergence of jaundice. Material and methods: In 80 of them (40 suffering and 40 from control group) were performed tests of numerous parameters in the laboratories of the Health Center in Krusevac. Magnesium was determined by spectrophotometry with methyl thymol blue, titanium yellow and blue xylidene. Bilirubin was determined by Jandrešek Grofov’s method. Results: The results were within the expectations, in addition to magnesium which was determined with methyl thymol blue. In all patients suffering from jaundice concentration of magnesium (0.67 ±0.14 mmol/l) statistically was significantly lower than tested of the control group (0.91± 0.059 mmol/). There is no theoretical data to reduce the concentration of magnesium in serum as a result of jaundice. That’s why we determined magnesium both in the control group and in sufferings with two methods as the titanium yellow, and xylidene blew. With these two methods we obtained results that were examined were within normal limits. Conclusion: This has led us to conclude that the determination of bilirubin interferes with magnesium methyl thymol blue. PMID:26244045

  19. The urinary concentrating defect in the Gunn strain of rat. Role of bilirubin.

    PubMed

    Call, N B; Tisher, C C

    1975-02-01

    The role of high serum and tissue levels of unconjegated bilirubin in the pathogenesis of the impaired urinary concentrating ability was investigated in homozygous (jj) Gunn rats with the congenital absence of hepatic glucuronyl transferase. Continuous phototherapy with blue fluorescent lights at a wave length of 460 nm or oral cholestyramine feeding or both reduced serum levels of unconjugated hilirubin to levels consistently below 3.0 mg/100 ml for several weeks in both weanling and adult jj Gunn rats. The renal concentrating defect was already present in weanling jj Gunn rats by 21 days of age. In treated weanling jj animals, maximum concentrating ability and the concentration of urea and nonurea solutes in the papilla and medulla, determined after 24 h of fluid deprivation, were normal when compared to unaffected heterozygous (Jj) littermates. Solute-free water reabsorption which is reduced in jaundiced jj Gunn rats was restored to normal in treated weanling jj rats. The tissue concentration of unconjugated bilirubin was reduced throughout the papilla and inner and outer medulla in the treated jj rats in comparison with untreated jj littermates. The defect in urinary concentrating ability was only partially reversible and sometimes irreversible in adult jj rats, probably because of permanent renal parenchymal damage occurring secondary to massive crystalline deposits in the papilla and medulla. It is concluded that unconjugated bilirubin is directly involved in the pathogenesis of the concentrating defect in jaundiced jj Gunn rats. PMID:1127102

  20. Serum Bilirubin Is Inversely Associated with Increased Arterial Stiffness in Men with Pre-Hypertension but Not Normotension

    PubMed Central

    Huang, Yao-Hsien; Yang, Yi-Ching; Lu, Feng-Hwa; Sun, Zih-Jie; Wu, Jin-Shang; Chang, Chih-Jen

    2016-01-01

    Objective Serum bilirubin level has shown to be inversely associated with coronary atherosclerosis, and may serve as a protective biomarker of coronary artery disease. Serum bilirubin has also been shown to be negatively associated with brachial-ankle pulse wave velocity (baPWV) in men without a history of hypertension, and in men with hypertension. It is unknown whether such associations can be observed in the pre-hypertensive or normotensive population. This study thus aimed to investigate the relationship between serum bilirubin level and increased arterial stiffness in subjects with pre-hypertension and normotension for both genders. Methods A cross-sectional sample of 3,399 apparently healthy subjects undergoing a medical check-up at National Cheng Kung University Hospital was enrolled between October 2006 and August 2009, after excluding subjects with serum total bilirubin level greater than 20.52 μmol/L. Increased arterial stiffness was defined as baPWV of 1,400 cm/s or higher as the dichotomous variable and bilirubin as the continuous variable. Results Based on multiple linear regression analysis, serum bilirubin level was inversely associated with baPWV in non-hypertensive men (β = -0.066, p < 0.001) but not in non-hypertensive women. In addition, the inverse relationship between bilirubin level and baPWV was found statistically significant only in pre-hypertensive men (β = -0.110, p < 0.001). Multiple logistic regression analysis showed that serum bilirubin was inversely associated with increased arterial stiffness in men with pre-hypertension (odds ratio = 0.955, 95% confidence interval = 0.916–0.996, p < 0.05) but not normotension after adjustment for other confounding factors. However, the relationship between total bilirubin level and increased arterial stiffness did not reach statistical significance for female subjects with pre-hypertension and normotension. Conclusion Serum bilirubin is inversely associated with increased arterial stiffness in

  1. Crystal structures of intermediates in the nitroalkane oxidase reaction.

    PubMed

    Héroux, Annie; Bozinovski, Dragana M; Valley, Michael P; Fitzpatrick, Paul F; Orville, Allen M

    2009-04-21

    The flavoenzyme nitroalkane oxidase is a member of the acyl-CoA dehydrogenase superfamily. Nitroalkane oxidase catalyzes the oxidation of neutral nitroalkanes to nitrite and the corresponding aldehydes or ketones. Crystal structures to 2.2 A resolution or better of enzyme complexes with bound substrates and of a trapped substrate-flavin adduct are described. The D402N enzyme has no detectable activity with neutral nitroalkanes [Valley, M. P., and Fitzpatrick, P. F. (2003) J. Am. Chem. Soc. 125, 8738-8739]. The structure of the D402N enzyme crystallized in the presence of 1-nitrohexane or 1-nitrooctane shows the presence of the substrate in the binding site. The aliphatic chain of the substrate extends into a tunnel leading to the enzyme surface. The oxygens of the substrate nitro group interact both with amino acid residues and with the 2'-hydroxyl of the FAD. When nitroalkane oxidase oxidizes nitroalkanes in the presence of cyanide, an electrophilic flavin imine intermediate can be trapped [Valley, M. P., Tichy, S. E., and Fitzpatrick, P. F. (2005) J. Am. Chem. Soc. 127, 2062-2066]. The structure of the enzyme trapped with cyanide during oxidation of 1-nitrohexane shows the presence of the modified flavin. A continuous hydrogen bond network connects the nitrogen of the CN-hexyl-FAD through the FAD 2'-hydroxyl to a chain of water molecules extending to the protein surface. Together, our complementary approaches provide strong evidence that the flavin cofactor is in the appropriate oxidation state and correlates well with the putative intermediate state observed within each of the crystal structures. Consequently, these results provide important structural descriptions of several steps along the nitroalkane oxidase reaction cycle. PMID:19265437

  2. Crystal Structures of Intermediates in the Nitroalkane Oxidase Reaction

    SciTech Connect

    Heroux, A.; Bozinovski, D; Valley, M; Fitzpatrick, P; Orville, A

    2009-01-01

    The flavoenzyme nitroalkane oxidase is a member of the acyl-CoA dehydrogenase superfamily. Nitroalkane oxidase catalyzes the oxidation of neutral nitroalkanes to nitrite and the corresponding aldehydes or ketones. Crystal structures to 2.2 {angstrom} resolution or better of enzyme complexes with bound substrates and of a trapped substrate-flavin adduct are described. The D402N enzyme has no detectable activity with neutral nitroalkanes. The structure of the D402N enzyme crystallized in the presence of 1-nitrohexane or 1-nitrooctane shows the presence of the substrate in the binding site. The aliphatic chain of the substrate extends into a tunnel leading to the enzyme surface. The oxygens of the substrate nitro group interact both with amino acid residues and with the 2'-hydroxyl of the FAD. When nitroalkane oxidase oxidizes nitroalkanes in the presence of cyanide, an electrophilic flavin imine intermediate can be trapped (Valley, M. P., Tichy, S. E., and Fitzpatrick, P. F. (2005) J. Am. Chem. Soc. 127, 2062-2066). The structure of the enzyme trapped with cyanide during oxidation of 1-nitrohexane shows the presence of the modified flavin. A continuous hydrogen bond network connects the nitrogen of the CN-hexyl-FAD through the FAD 2'-hydroxyl to a chain of water molecules extending to the protein surface. Together, our complementary approaches provide strong evidence that the flavin cofactor is in the appropriate oxidation state and correlates well with the putative intermediate state observed within each of the crystal structures. Consequently, these results provide important structural descriptions of several steps along the nitroalkane oxidase reaction cycle.

  3. Significance and prognostic value of increased serum direct bilirubin level for lymph node metastasis in Chinese rectal cancer patients

    PubMed Central

    Gao, Chun; Fang, Long; Li, Jing-Tao; Zhao, Hong-Chuan

    2016-01-01

    AIM: To determine the significance of increased serum direct bilirubin level for lymph node metastasis (LNM) in Chinese rectal cancer patients, after those with known hepatobiliary and pancreatic diseases were excluded. METHODS: A cohort of 469 patients, who were treated at the China-Japan Friendship Hospital, Ministry of Health (Beijing, China), in the period from January 2003 to June 2011, and with a pathological diagnosis of rectal adenocarcinoma, were recruited. They included 231 patients with LNM (49.3%) and 238 patients without LNM. Follow-up for these patients was taken through to December 31, 2012. RESULTS: The baseline serum direct bilirubin concentration was (median/inter-quartile range) 2.30/1.60-3.42 μmol/L. Univariate analysis showed that compared with patients without LNM, the patients with LNM had an increased level of direct bilirubin (2.50/1.70-3.42 vs 2.10/1.40-3.42, P = 0.025). Multivariate analysis showed that direct bilirubin was independently associated with LNM (OR = 1.602; 95%CI: 1.098-2.338, P = 0.015). Moreover, we found that: (1) serum direct bilirubin differs between male and female patients; a higher concentration was associated with poor tumor classification; (2) as the baseline serum direct bilirubin concentration increased, the percentage of patients with LNM increased; and (3) serum direct bilirubin was associated with the prognosis of rectal cancer patients and higher values indicated poor prognosis. CONCLUSION: Higher serum direct bilirubin concentration was associated with the increased risk of LNM and poor prognosis in our rectal cancers. PMID:26937145

  4. Serum bilirubin value predicts hospital admission in carbon monoxide-poisoned patients. Active player or simple bystander?

    PubMed Central

    Cervellin, Gianfranco; Comelli, Ivan; Buonocore, Ruggero; Picanza, Alessandra; Rastelli, Gianni; Lippi, Giuseppe

    2015-01-01

    OBJECTIVES: Although carbon monoxide poisoning is a major medical emergency, the armamentarium of recognized prognostic biomarkers displays unsatisfactory diagnostic performance for predicting cumulative endpoints. METHODS: We performed a retrospective and observational study to identify all patients admitted for carbon monoxide poisoning during a 2-year period. Complete demographical and clinical information, along with the laboratory data regarding arterial carboxyhemoglobin, hemoglobin, blood lactate and total serum bilirubin, was retrieved. RESULTS: The study population consisted of 38 poisoned patients (23 females and 15 males; mean age 39±21 years). Compared with discharged subjects, hospitalized patients displayed significantly higher values for blood lactate and total serum bilirubin, whereas arterial carboxyhemoglobin and hemoglobin did not differ. In a univariate analysis, hospitalization was significantly associated with blood lactate and total serum bilirubin, but not with age, sex, hemoglobin or carboxyhemoglobin. The diagnostic performance obtained after combining the blood lactate and total serum bilirubin results (area under the curve, 0.90; 95% CI, 0.81-0.99; p<0.001) was better than that obtained for either parameter alone. CONCLUSION: Although it remains unclear whether total serum bilirubin acts as an active player or a bystander, we conclude that the systematic assessment of bilirubin may, alongside lactate levels, provide useful information for clinical decision making regarding carbon monoxide poisoning. PMID:26375565

  5. Unilobar Versus Bilobar Biliary Drainage: Effect on Quality of Life and Bilirubin Level Reduction

    PubMed Central

    Gamanagatti, Shivanand; Singh, Tejbir; Sharma, Raju; Srivastava, Deep N; Dash, Nihar Ranjan; Garg, Pramod Kumar

    2016-01-01

    Background: Percutaneous biliary drainage is an accepted palliative treatment for malignant biliary obstruction. Purpose: To assess the effect on quality of life (QOL) and bilirubin level reduction in patients with inoperable malignant biliary obstruction treated by unilobar or bilobar percutaneous transhepatic biliary drainage (PTBD). Materials and Methods: Over a period of 2 years, 49 patients (age range, 22–75 years) of inoperable malignant biliary obstruction were treated by PTBD. Technical and clinical success rates, QOL, patency rates, survival rates, and complications were recorded. Clinical success rates, QOL, and bilirubin reduction were compared in patients treated with complete (n = 21) versus partial (n = 28) liver parenchyma drainage. QOL before and 1 month after biliary drainage were analyzed retrospectively between these two groups. Results: Biliary drainage was successful in all 49 patients, with an overall significant reduction of the postintervention bilirubin levels (P < 0.001) resulting in overall clinical success rate of 89.97%. Clinical success rates were similar in patients treated with whole-liver drainage versus partial-liver drainage. Mean serum bilirubin level before PTBD was 19.85 mg/dl and after the procedure at 1 month was 6.02 mg/dl. The mean baseline functional score was 39.35, symptom scale score was 59.55, and global health score was 27.45. At 1 month, mean functional score was 61.25, symptom scale score was 36.0 4, and global health score was 56.33, with overall significant improvement in QOL (<0.001). There was a statistically significant difference in the improvement of the QOL scores (P = 0.002), among patients who achieved clinical success, compared with those patients who did not achieve clinical success at 1 month. We did not find any significant difference in the QOL scores in patients according to the amount of liver drained (unilateral or bilateral drainage), the type of internalization used (ring biliary or stent

  6. Arsenite Oxidase Also Functions as an Antimonite Oxidase

    PubMed Central

    Wang, Qian; Warelow, Thomas P.; Kang, Yoon-Suk; Romano, Christine; Osborne, Thomas H.; Lehr, Corinne R.; Bothner, Brian; McDermott, Timothy R.

    2015-01-01

    Arsenic and antimony are toxic metalloids and are considered priority environmental pollutants by the U.S. Environmental Protection Agency. Significant advances have been made in understanding microbe-arsenic interactions and how they influence arsenic redox speciation in the environment. However, even the most basic features of how and why a microorganism detects and reacts to antimony remain poorly understood. Previous work with Agrobacterium tumefaciens strain 5A concluded that oxidation of antimonite [Sb(III)] and arsenite [As(III)] required different biochemical pathways. Here, we show with in vivo experiments that a mutation in aioA [encoding the large subunit of As(III) oxidase] reduces the ability to oxidize Sb(III) by approximately one-third relative to the ability of the wild type. Further, in vitro studies with the purified As(III) oxidase from Rhizobium sp. strain NT-26 (AioA shares 94% amino acid sequence identity with AioA of A. tumefaciens) provide direct evidence of Sb(III) oxidation but also show a significantly decreased Vmax compared to that of As(III) oxidation. The aioBA genes encoding As(III) oxidase are induced by As(III) but not by Sb(III), whereas arsR gene expression is induced by both As(III) and Sb(III), suggesting that detection and transcriptional responses for As(III) and Sb(III) differ. While Sb(III) and As(III) are similar with respect to cellular extrusion (ArsB or Acr3) and interaction with ArsR, they differ in the regulatory mechanisms that control the expression of genes encoding the different Ars or Aio activities. In summary, this study documents an enzymatic basis for microbial Sb(III) oxidation, although additional Sb(III) oxidation activity also is apparent in this bacterium. PMID:25576601

  7. Bound states and the Bekenstein bound

    SciTech Connect

    Bousso, Raphael

    2003-10-16

    We explore the validity of the generalized Bekenstein bound, S<= pi M a. We define the entropy S as the logarithm of the number of states which have energy eigenvalue below M and are localized to a flat space region of width alpha. If boundary conditions that localize field modes are imposed by fiat, then the bound encounters well-known difficulties with negative Casimir energy and large species number, as well as novel problems arising only in the generalized form. In realistic systems, however, finite-size effects contribute additional energy. We study two different models for estimating such contributions. Our analysis suggests that the bound is both valid and nontrivial if interactions are properly included, so that the entropy S counts the bound states of interacting fields.

  8. Manganese(IV) Oxide Production by Acremonium sp. Strain KR21-2 and Extracellular Mn(II) Oxidase Activity

    PubMed Central

    Miyata, Naoyuki; Tani, Yukinori; Maruo, Kanako; Tsuno, Hiroshi; Sakata, Masahiro; Iwahori, Keisuke

    2006-01-01

    Ascomycetes that can deposit Mn(III, IV) oxides are widespread in aquatic and soil environments, yet the mechanism(s) involved in Mn oxide deposition remains unclear. A Mn(II)-oxidizing ascomycete, Acremonium sp. strain KR21-2, produced a Mn oxide phase with filamentous nanostructures. X-ray absorption near-edge structure (XANES) spectroscopy showed that the Mn phase was primarily Mn(IV). We purified to homogeneity a laccase-like enzyme with Mn(II) oxidase activity from cultures of strain KR21-2. The purified enzyme oxidized Mn(II) to yield suspended Mn particles; XANES spectra indicated that Mn(II) had been converted to Mn(IV). The pH optimum for Mn(II) oxidation was 7.0, and the apparent half-saturation constant was 0.20 mM. The enzyme oxidized ABTS [2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid)] (pH optimum, 5.5; Km, 1.2 mM) and contained two copper atoms per molecule. Moreover, the N-terminal amino acid sequence (residues 3 to 25) was 61% identical with the corresponding sequence of an Acremonium polyphenol oxidase and 57% identical with that of a Myrothecium bilirubin oxidase. These results provide the first evidence that a fungal multicopper oxidase can convert Mn(II) to Mn(IV) oxide. The present study reinforces the notion of the contribution of multicopper oxidase to microbially mediated precipitation of Mn oxides and suggests that Acremonium sp. strain KR21-2 is a good model for understanding the oxidation of Mn in diverse ascomycetes. PMID:17021194

  9. Crystal structure and site-directed mutagenesis of a nitroalkane oxidase from Streptomyces ansochromogenes.

    PubMed

    Li, Yanhua; Gao, Zengqiang; Hou, Haifeng; Li, Lei; Zhang, Jihui; Yang, Haihua; Dong, Yuhui; Tan, Huarong

    2011-02-18

    Nitroalkane oxidase (NAO) catalyzes neutral nitroalkanes to their corresponding aldehydes or ketones, hydrogen peroxide and nitrite. The crystal structure of NAO from Streptomyces ansochromogenes was determined; it consists of two domains, a TIM barrel domain bound to FMN and C-terminal domain with a novel folding pattern. Site-directed mutagenesis of His179, which is spatially adjacent to FMN, resulted in the loss of enzyme activity, demonstrating that this amino acid residue is important for catalysis. The crystal structure of mutant H179D-nitroethane was also analyzed. Interestingly, Sa-NAO shows the typical function as nitroalkane oxidase but its structure is similar to that of 2-nitropropane dioxygenase. Overall, these results suggest that Sa-NAO is a novel nitroalkane oxidase with TIM barrel structure. PMID:21147069

  10. Studies on the Mechanism of Aldehyde Oxidase and Xanthine Oxidase

    PubMed Central

    Alfaro, Joshua F.

    2009-01-01

    DFT calculations support a concerted mechanism for xanthine oxidase and aldehyde oxidase hydride displacement from the sp2 carbon of 6-substituted 4-quinazolinones. The variations in transition state structure show that C-O bond formation is nearly complete in the transition state and the transition state changes are anti-Hammond with the C-H and C-O bond lengths being more product-like for the faster reactions. The C-O bond length in the transition state is around 90% formed. However, the C-H bond is only about 80% broken. This leads to a very tetrahedral transition state with an O-C-N angle of 109 degrees. Thus, while the mechanism is concerted, the anti-bonding orbital of the C-H bond that is broken is not directly attacked by the nucleophile and instead hydride displacement occurs after almost complete tetrahedral transition state formation. In support of this the C=N bond is lengthened in the transition state indicating that attack on the electrophilic carbon occurs by addition to the C=N bond with negative charge increasing on the nitrogen. Differences in experimental reaction rates are accurately reproduced by these calculations, and tend to support this mechanism. PMID:18998731

  11. Studies on the mechanism of aldehyde oxidase and xanthine oxidase.

    PubMed

    Alfaro, Joshua F; Jones, Jeffrey P

    2008-12-01

    DFT calculations support a concerted mechanism for xanthine oxidase and aldehyde oxidase hydride displacement from the sp(2) carbon of 6-substituted 4-quinazolinones. The variations in transition state structure show that C-O bond formation is nearly complete in the transition state and the transition state changes are anti-Hammond with the C-H and C-O bond lengths being more product-like for the faster reactions. The C-O bond length in the transition state is around 90% formed. However, the C-H bond is only about 80% broken. This leads to a very tetrahedral transition state with an O-C-N angle of 109 degrees. Thus, while the mechanism is concerted, the antibonding orbital of the C-H bond that is broken is not directly attacked by the nucleophile and instead hydride displacement occurs after almost complete tetrahedral transition state formation. In support of this the C=N bond is lengthened in the transition state indicating that attack on the electrophilic carbon occurs by addition to the C=N bond with negative charge increasing on the nitrogen. Differences in experimental reaction rates are accurately reproduced by these calculations and tend to support this mechanism. PMID:18998731

  12. Monoamine Oxidase Inhibitors: Clinical Review

    PubMed Central

    Remick, Ronald A.; Froese, Colleen

    1990-01-01

    Monoamine oxidase inhibitors (MAOIs) are effective antidepressant agents. They are increasingly and effectively used in a number of other psychiatric and non-psychiatric medical syndromes. Their potential for serious toxicity (i.e., hypertensive reaction) is far less than original reports suggest, and newer reversible substrate-specific MAOIs may offer even less toxicity. The author reviews the pharmacology, mechanism of action, clinical indications, and dosing strategies of MAOIs. The common MAOI side-effects (hypotension, weight gain, sexual dysfunction, insomnia, daytime sedation, myoclonus, and hypertensive episodes) are described and management techniques suggested. Recent clinical developments involving MAOIs are outlined. PMID:21233984

  13. A matrix lower bound

    SciTech Connect

    Grcar, Joseph F.

    2002-02-04

    A matrix lower bound is defined that generalizes ideas apparently due to S. Banach and J. von Neumann. The matrix lower bound has a natural interpretation in functional analysis, and it satisfies many of the properties that von Neumann stated for it in a restricted case. Applications for the matrix lower bound are demonstrated in several areas. In linear algebra, the matrix lower bound of a full rank matrix equals the distance to the set of rank-deficient matrices. In numerical analysis, the ratio of the matrix norm to the matrix lower bound is a condition number for all consistent systems of linear equations. In optimization theory, the matrix lower bound suggests an identity for a class of min-max problems. In real analysis, a recursive construction that depends on the matrix lower bound shows that the level sets of continuously differential functions lie asymptotically near those of their tangents.

  14. Immunological comparison of sulfite oxidase

    SciTech Connect

    Pollock, V.; Barber, M.J. )

    1991-03-11

    Polyclonal antibodies (rabbit), elicited against FPLC-purified chicken and rat liver sulfite oxidase (SO), have been examined for inhibition and binding to purified chicken (C), rat (R), bovine (B), alligator (A) and shark (S) liver enzymes. Anti-CSO IgG cross-reacted with all five enzymes, with varying affinities, in the order CSO=ASO{gt}RSO{gt}BSO{gt}SSO. Anti-ROS IgG also cross-reacted with all five enzymes in the order RSO{gt}CSO=ASO{gt}BSO{gt}SSO. Anti-CSO IgG inhibited sulfite:cyt. c reductase (S:CR), sulfite:ferricyanide reductase (S:FR) and sulfite:dichlorophenolindophenol reductase (S:DR) activities of CSO to different extents (S:CR{gt}S:FR=S:DR). Similar differential inhibition was found for anti-ROS IgG and RSO S:CR, S:FR and S:DR activities. Anti-CSO IgG inhibited S:CR activities in the order CSO=ASO{much gt}SSO{gt}BSO. RSO was uninhibited. For anti-RSO IgG the inhibition order was RSO{gt}SSO{gt}BSO{gt}ASO. CSO was uninhibited. Anti-CSO and RSO IgGs partially inhibited Chlorella nitrate reductase (NR). Minor cross-reactivity was found for xanthine oxidase. Common antigenic determinants for all five SO's and NR are indicated.

  15. Mitochondrial Cytochrome c Oxidase Deficiency

    PubMed Central

    Rak, Malgorzata; Bénit, Paule; Chrétien, Dominique; Bouchereau, Juliette; Schiff, Manuel; El-Khoury, Riyad; Tzagoloff, Alexander; Rustin, Pierre

    2016-01-01

    As with other mitochondrial respiratory chain components, marked clinical and genetic heterogeneity is observed in patients with a cytochrome c oxidase deficiency. This constitutes a considerable diagnostic challenge and raises a number of puzzling questions. So far, pathological mutations have been reported in more than 30 genes, in both mitochondrial and nuclear DNA, affecting either structural subunits of the enzyme or proteins involved in its biogenesis. In this review, we discuss the possible causes of the discrepancy between the spectacular advances made in the identification of the molecular bases of cytochrome oxidase deficiency and the lack of any efficient treatment in diseases resulting from such deficiencies. This brings back many unsolved questions related to the frequent delay of clinical manifestation, variable course and severity, and tissue-involvement often associated with these diseases. In this context, we stress the importance to study different models of these diseases, but also discuss the limitations encountered in most available disease models. In the future, with the possible exception of replacement therapy using genes, cells or organs, a better understanding of underlying mechanism(s) of these mitochondrial diseases is presumably required to develop efficient therapy. PMID:26846578

  16. In vivo oxalate degradation by liposome encapsulated oxalate oxidase in rat model of hyperoxaluria

    PubMed Central

    Dahiya, Tulika; Pundir, C.S.

    2013-01-01

    Background & objectives: High level of urinary oxalate substantially increases the risk of hyperoxaluria, a significant risk factor for urolithiasis. The primary goal of this study was to reduce urinary oxalate excretion employing liposome encapsulated oxalate oxidase in animal model. Methods: A membrane bound oxalate oxidase was purified from Bougainvillea leaves. The enzyme in its native form was less effective at the physiological pH of the recipient animal. To increase its functional viability, the enzyme was immobilized on to ethylene maleic anhydride (EMA). Rats were injected with liposome encapsulated EMA- oxalate oxidase and the effect was observed on degradation of oxalic acid. Results: The enzyme was purified to apparent homogeneity with 60-fold purification and 31 per cent yield. The optimum pH of EMA-derivative enzyme was 6.0 and it showed 70 per cent of its optimal activity at pH 7.0. The EMA-bound enzyme encapsulated into liposome showed greater oxalate degradation in 15 per cent casein vitamin B6 deficient fed rats as compared with 30 per cent casein vitamin B6 deficient fed rats and control rats. 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. PMID:23481063

  17. Single mutations that redirect internal proton transfer in the ba3 oxidase from Thermus thermophilus

    PubMed Central

    Smirnova, Irina; Chang, Hsin-Yang; von Ballmoos, Christoph; Ädelroth, Pia; Gennis, Robert B.; Brzezinski, Peter

    2014-01-01

    The ba3-type cytochrome c oxidase from Thermus thermophilus is a membrane-bound proton pump. Results from earlier studies have shown that with the aa3-type oxidases proton uptake to the catalytic site and “pump site” occur simultaneously. However, with the ba3 oxidase the pump site is loaded before proton transfer to the catalytic site because the proton transfer to the latter is slower than with the aa3 oxidases. In addition, the timing of formation and decay of catalytic intermediates is different in the two types of oxidases. In the present study, we have investigated two mutant ba3 CytcOs in which residues of the proton pathway leading to the catalytic site as well as the pump site were exchanged, Thr312Val and Tyr244Phe. Even though the ba3 CytcO uses only a single proton pathway for transfer of the substrate and “pumped” protons, the amino-acid residue substitutions had distinctly different effects on the kinetics of proton transfer to the catalytic site and the pump site, respectively. The results indicate that the rates of these reactions can be modified independently by replacement of single residues within the proton pathway. Furthermore, the data suggest that the Thr312Val and Tyr244Phe mutations interfere with a structural rearrangement in the proton pathway that is rate limiting for proton transfer to the catalytic site. PMID:24004023

  18. Utility of bilirubins and bile acids as endogenous biomarkers for the inhibition of hepatic transporters.

    PubMed

    Watanabe, Tomoko; Miyake, Manami; Shimizu, Toshinobu; Kamezawa, Miho; Masutomi, Naoya; Shimura, Takesada; Ohashi, Rikiya

    2015-04-01

    It is useful to identify endogenous substrates for the evaluation of drug-drug interactions via transporters. In this study, we investigated the utility of bilirubins, substrates of OATPs and MRP2, and bile acids and substrates of NTCP and BSEP, as biomarkers for the inhibition of transporters. In rats administered 20 and 80 mg/kg rifampicin, the plasma levels of bilirubin glucuronides were elevated, gradually decreased, and almost returned to the baseline level at 24 hours after administration without an elevation of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). This result indicates the transient inhibition of rOatps and/or rMrp2. Although the correlation between free plasma concentrations and IC50 values of rOatps depended on the substrates used in the in vitro studies, the inhibition of rOatps by rifampicin was confirmed in the in vivo study using valsartan as a substrate of rOatps. In rats administered 10 and 30 mg/kg cyclosporin A, the plasma levels of bile acids were elevated and persisted for up to 24 hours after administration without an elevation of ALT and AST. This result indicates the continuous inhibition of rNtcp and/or rBsep, although there were differences between the free plasma or liver concentrations and IC50 values of rNtcp or rBsep, respectively. This study suggests that the monitoring of bilirubins and bile acids in plasma is useful in evaluating the inhibitory potential of their corresponding transporters. PMID:25581390

  19. Physical Uncertainty Bounds (PUB)

    SciTech Connect

    Vaughan, Diane Elizabeth; Preston, Dean L.

    2015-03-19

    This paper introduces and motivates the need for a new methodology for determining upper bounds on the uncertainties in simulations of engineered systems due to limited fidelity in the composite continuum-level physics models needed to simulate the systems. We show that traditional uncertainty quantification methods provide, at best, a lower bound on this uncertainty. We propose to obtain bounds on the simulation uncertainties by first determining bounds on the physical quantities or processes relevant to system performance. By bounding these physics processes, as opposed to carrying out statistical analyses of the parameter sets of specific physics models or simply switching out the available physics models, one can obtain upper bounds on the uncertainties in simulated quantities of interest.

  20. Asymptotic entropy bounds

    NASA Astrophysics Data System (ADS)

    Bousso, Raphael

    2016-07-01

    We show that known entropy bounds constrain the information carried off by radiation to null infinity. We consider distant, planar null hypersurfaces in asymptotically flat spacetime. Their focusing and area loss can be computed perturbatively on a Minkowski background, yielding entropy bounds in terms of the energy flux of the outgoing radiation. In the asymptotic limit, we obtain boundary versions of the quantum null energy condition, of the generalized Second Law, and of the quantum Bousso bound.

  1. Self-assembly of aqueous bilirubin ditaurate, a natural conjugated bile pigment, to contraposing enantiomeric dimers and M(-) and P(+) tetramers and their selective hydrophilic disaggregation by monomers and micelles of bile salts.

    PubMed

    Neubrand, Michael W; Carey, Martin C; Laue, Thomas M

    2015-02-24

    The solution behavior of bilirubin ditaurate (BDT), the first naturally occurring conjugated bile pigment to be physically and chemically characterized, was assessed in aqueous solution and in monomeric and micellar solutions of common taurine-conjugated bile salts (BS). Analytical ultracentrifugation revealed that BDT self-associates in monomer-dimer equilibria between 1 and 500 μM, forming limiting tetramers at low millimolar concentrations. Self-association was enthalpically driven with ΔG values of ≈5 kcal/mol, suggesting strong hydrophobic interactions. Added NaCl and decreases in temperature shifted the oligomerization to lower BDT concentrations. On the basis of circular dichroism spectra and the limiting size of the self-aggregates, we infer that the tetramers are composed of 2P(+) and 2M(-) enantiomeric BDT pairs in "ridge-tile" conformations interacting in a "double-bookend" structure. With added monomeric BS, blue shifts in the UV-vis spectra and tight isosbestic points revealed that BDT/BS heterodimers form, followed by BDT "decorating" BS micelles mostly via hydrophilic interactions. Conformational enantiomerism, fluorescence intensities, and anisotropy, as well as resistance of the hybrid particles to disaggregation in 6 M urea, suggested that two or three hydrogen-bonding sites bound BDT monomers to the hydroxyl groups of BS, possibly via pyrrole-π-orbital-OH interactions. BDT stabilized these interactions by enveloping the BS in its "ridge-tile" pincers with variable strain that maximized van der Waals interactions. Possibly because the BDT molecule becomes highly strained with BS subtending a 7β-hydroxyl group, BDT became totally resistant to oxidation in air. This work predicts that, because of BS dissolution of the BDT self-aggregates, BS/bilirubin hybrid particles, which are stabilized hydrophilically, are likely to be the dominant mode of transport for all conjugated bilirubins in bile. PMID:25671490

  2. RRM analysis of protoporphyrinogen oxidase.

    PubMed

    Sauren, M; Pirogova, E; Cosic, I

    2004-12-01

    Enzymes are crucial in accelerating metabolic reactions in living organisms. Protoporphyrinogen oxidase (PpOI) is an enzyme that catalyses the production of protoporphyrin IX (PpIX), a protein used in a cancer treatment known as photodynamic therapy (PDT). In this study, a structure-function analysis of PpOI was carried out using the Resonant Recognition Model (RRM), a physico-mathematical approach for analysis of proteins interactions. This method is based on the finding that the distribution of delocalised electron energies along the protein plays a crucial role in determining the protein's biological activity. Two digital signal processing (DSP) methods were used: Fourier Transform (FT) and Continuous Wavelet Transform (CWT). Here we have determined the characteristic frequencies and the "hot spot" amino acids, and predicted the location of proteins' active site(s). Several proteins that potentially belong to the PpOI functional group were also analysed to distinguish their viability in this role. PMID:15712584

  3. Impact of Rhesus disease on the global problem of bilirubin-induced neurologic dysfunction.

    PubMed

    Zipursky, Alvin; Bhutani, Vinod K

    2015-02-01

    Clinical experience with Rhesus (Rh) disease and its post-icteric sequelae is limited among high-income countries because of nearly over four decades of effective prevention care. We hypothesized that Rh disease is prevalent in other regions of the world because it is likely that protection is limited or non-existent. Following a worldwide study, it has been concluded that Rh hemolytic disease is a significant public health problem resulting in stillbirths and neonatal deaths, and is a major cause of severe hyperbilirubinemia with its sequelae, kernicterus and bilirubin-induced neurologic dysfunction. Knowing that effective Rh-disease prophylaxis depends on maternal blood-type screening, healthcare afforded to the high-risk mothers needs to be free of bottlenecks and coupled with unfettered access to effective Rh-immunoglobulin. Future studies that match the universal identification of Rh-negative status of women and targeted use of immunoprophylaxis to prevent childhood bilirubin neurotoxicity are within reach, based on vast prior experiences. PMID:25582277

  4. Method for Estimating Bilirubin Isomerization Efficiency in Phototherapy to Treat Neonatal Jaundice

    NASA Astrophysics Data System (ADS)

    Lisenko, S. A.; Kugeiko, M. M.

    2014-11-01

    We propose a method for quantitative assessment of the efficacy of phototherapy to treat neonatal jaundice using the diffuse reflectance spectrum for the newborn's skin, based on the analytical dependence of the measured spectrum on the structural and morphological parameters of the skin, affecting the optical conditions in the medium, and an algorithm for rapid calculation of the bilirubin photoisomerization rate in the skin tissues as a function of the structural and morphological parameters of the skin and the wavelength of the exciting radiation. From the results of a numerical simulation of the process of radiation transport in the skin, we assess the stability of our method to variations in the scattering properties of the skin and the concentrations of its optically active chromophores (melanin, oxyhemoglobin, deoxyhemoglobin). We show that in order to achieve the maximum efficacy of phototherapy, we should use light from the range 484-496 nm. In this case, the intensity of the exciting radiation should be selected individually for each newborn according to the bilirubin photoisomerization rate characteristic for it.

  5. Unconjugated Bilirubin exerts Pro-Apoptotic Effect on Platelets via p38-MAPK activation

    PubMed Central

    NaveenKumar, Somanathapura K.; Thushara, Ram M.; Sundaram, Mahalingam S.; Hemshekhar, Mahadevappa; Paul, Manoj; Thirunavukkarasu, Chinnasamy; Basappa; Nagaraju, Ganesh; Raghavan, Sathees C.; Girish, Kesturu S.; Kemparaju, Kempaiah; Rangappa, Kanchugarakoppal S.

    2015-01-01

    Thrombocytopenia is one of the most frequently observed secondary complications in many pathological conditions including liver diseases, where hyperbilirubinemia is very common. The present study sought to find the cause of thrombocytopenia in unconjugated hyperbilirubinemic conditions. Unconjugated bilirubin (UCB), an end-product of heme catabolism, is known to have pro-oxidative and cytotoxic effects at high serum concentration. We investigated the molecular mechanism underlying the pro-apoptotic effect of UCB on human platelets in vitro, and followed it up with studies in phenylhydrazine-induced hyperbilirubinemic rat model and hyperbilirubinemic human subjects. UCB is indeed found to significantly induce platelet apoptotic events including elevated endogenous reactive oxygen species generation, mitochondrial membrane depolarization, increased intracellular calcium levels, cardiolipin peroxidation and phosphatidylserine externalization (p < 0.001) as evident by FACS analysis. The immunoblots show the elevated levels of cytosolic cytochrome c and caspase activation in UCB-treated platelets. Further, UCB is found to induce mitochondrial ROS generation leading to p38 activation, followed by downstream activation of p53, ultimately resulting in altered expression of Bcl-2 and Bax proteins as evident from immunoblotting. All these parameters conclude that elevated unconjugated bilirubin causes thrombocytopenia by stimulating platelet apoptosis via mitochondrial ROS-induced p38 and p53 activation. PMID:26459859

  6. Effects of aluminum chloride on serum proteins, bilirubin, and hepatic trace elements in chickens.

    PubMed

    Wang, Ben; Zhu, Yanzhu; Zhang, Hongling; Liu, Liming; Li, Guojiang; Song, Yongli; Li, Yanfei

    2016-09-01

    The aim of this study was to reveal the effects of aluminum chloride (AlCl3) on the hepatic metabolism function and trace elements' distribution. Two hundred healthy male chickens (1 day old) were intraperitoneally administered with AlCl3 (0, 18.31, 27.47, and 36.62 mg kg(-1) day(-1) of Al(3+)) consecutively for 3 days. Then the chickens were allowed to rest for 1 day. The cycle lasted four days. The cycle was repeated 15 times (60 days). The contents of serum total protein (TP), albumin (ALB), total bilirubin (TBI), direct bilirubin (DBI), hepatic aluminum (Al), copper (Cu), iron (Fe), and zinc (Zn) were examined. The results showed that the contents of serum TP and ALB and hepatic Fe and Zn decreased and the contents of serum TBI and DBI and hepatic Al and Cu increased in the chickens with AlCl3 This indicates that chronic administration of AlCl3 impairs the hepatic metabolism function and disorders the hepatic trace elements' distribution. PMID:25896954

  7. Crosstalk between mitochondria and NADPH oxidases

    PubMed Central

    Dikalov, Sergey

    2011-01-01

    Reactive oxygen species (ROS) play an important role in physiological and pathological processes. In recent years, a feed-forward regulation of the ROS sources has been reported. The interaction between main cellular sources of ROS, such as mitochondria and NADPH oxidases, however, remain obscure. This work summarizes the latest findings on the role of crosstalk between mitochondria and NADPH oxidases in pathophysiological processes. Mitochondria have the highest levels of antioxidants in the cell and play an important role in the maintenance of cellular redox status, thereby acting as an ROS and redox sink and limiting NADPH oxidase activity. Mitochondria, however, are not only a target for ROS produced by NADPH oxidase but also a significant source of ROS, which under certain condition may stimulate NADPH oxidases. This crosstalk between mitochondria and NADPH oxidases, therefore, may represent a feed-forward vicious cycle of ROS production which can be pharmacologically targeted under conditions of oxidative stress. It has been demonstrated that mitochondria-targeted antioxidants break this vicious cycle, inhibiting ROS production by mitochondria and reducing NADPH oxidase activity. This may provide a novel strategy for treatment of many pathological conditions including aging, atherosclerosis, diabetes, hypertension and degenerative neurological disorders in which mitochondrial oxidative stress seems to play a role. It is conceivable that the use of mitochondria-targeted treatments would be effective in these conditions. PMID:21777669

  8. Fluorescence excitation spectrum of bilirubin in blood: a model for the action spectrum for phototherapy of neonatal jaundice.

    PubMed

    Lamola, Angelo A; Russo, Marie

    2014-01-01

    A recent report (Lamola et al. 2013 Pediatric Research, 74, 54-60) presents a semiempirical model for facile calculation of an action spectrum for bilirubin photochemistry in vivo using the most current knowledge of the optics of neonatal skin. The calculations indicate that competition for phototherapy light by hemoglobin in the skin is the predominant factor that defines the spectrum of light absorbed by bilirubin. If the latter is correct, a valid physical analog of the calculated spectrum is the excitation spectrum of bilirubin in blood. The fluorescence excitation spectrum was recorded and, indeed, found to be very similar to the calculated spectrum. Both spectra exhibit maxima near 476 nm and widths at half height of about 50 nm. This result supports the conclusion that light between 460 and 490 nm is most effective for phototherapy of neonatal jaundice. PMID:23998276

  9. Molecular basis of bilirubin UDP-glucuronosyltransferase induction in spontaneously diabetic rats, acetone-treated rats and starved rats.

    PubMed Central

    Braun, L; Coffey, M J; Puskás, F; Kardon, T; Nagy, G; Conley, A A; Burchell, B; Mandl, J

    1998-01-01

    The co-ordinated induction of several hepatic drug-metabolizing enzymes is a common feature in the regulation of drug biotransformation under normal and pathological conditions. In the present study the activity and expression of bilirubin UDP-glucuronosyltransferase (UGT1A1) were investigated in livers of BioBreeding/Worcester diabetic, fasted and acetone-treated rats. Bilirubin glucuronidation was stimulated by all three treatments; this was correlated with an increase in the UGT1A1 protein concentration in hepatic microsomes. Transcriptional induction of UGT1A1 was also observed in diabetes and starvation but not with acetone treatment, which apparently caused translational stabilization of the enzyme protein. The hormonal/metabolic alterations in diabetes and starvation might be a model for postnatal development. The sudden interruption of maternal glucose supply signals the enhanced expression of UGT1A1, giving a novel explanation for the physiological induction of bilirubin glucuronidation in newborn infants. PMID:9841869

  10. Predictive effects of bilirubin on response of colorectal cancer to irinotecan-based chemotherapy

    PubMed Central

    Yu, Qian-Qian; Qiu, Hong; Zhang, Ming-Sheng; Hu, Guang-Yuan; Liu, Bo; Huang, Liu; Liao, Xin; Li, Qian-Xia; Li, Zhi-Huan; Yuan, Xiang-Lin

    2016-01-01

    AIM: To examine the predictive effects of baseline serum bilirubin levels and UDP-glucuronosyltransferase (UGT) 1A1*28 polymorphism on response of colorectal cancer to irinotecan-based chemotherapy. METHODS: The present study was based on a prospective multicenter longitudinal trial of Chinese metastatic colorectal cancer (mCRC) patients treated with irinotecan-based chemotherapy (NCT01282658). Baseline serum bilirubin levels, including total bilirubin (TBil) and unconjugated bilirubin (UBil), were measured, and genotyping of UGT1A1*28 polymorphism was performed. Receiver operating characteristic curve (ROC) analysis was used to determine cutoff values of TBil and UBil. The TBil values were categorized into > 13.0 or ≤ 13.0 groups; the UBil values were categorized into > 4.1 or ≤ 4.1 groups. Combining the cutoff values of TBil and UBil, which was recorded as CoBil, patients were classified into three groups. The classifier’s performance of UGT1A1*28 and CoBil for predicting treatment response was evaluated by ROC analysis. Associations between response and CoBil or UGT1A1*28 polymorphism were estimated using simple and multiple logistic regression models. RESULTS: Among the 120 mCRC patients, the serum bilirubin level was significantly different between the UGT1A1*28 wild-type and mutant genotypes. Patients with the mutant genotype had an increased likelihood of a higher TBil (P = 0.018) and a higher UBil (P = 0.014) level compared with the wild-type genotype. Patients were stratified into three groups based on CoBil. Group 1 was patients with TBil > 13.0 and UBil > 4.1; Group 2 was patients with TBil ≤ 13.0 and UBil > 4.1; and Group 3 was patients with TBil ≤ 13.0 and UBil ≤ 4.1. Patients in Group 3 had more than a 10-fold higher likelihood of having a response in the simple (OR = 11.250; 95%CI: 2.286-55.367; P = 0.003) and multiple (OR = 16.001; 95%CI: 2.802 -91.371; P = 0.002) analyses compared with the Group 1 individuals. Patients carrying the UGT1

  11. Longitudinal Blood Pressure Control, Long-Term Mortality, and Predictive Utility of Serum Liver Enzymes and Bilirubin in Hypertensive Patients.

    PubMed

    McCallum, Linsay; Panniyammakal, Jeemon; Hastie, Claire E; Hewitt, Jonathan; Patel, Rajan; Jones, Gregory C; Muir, Scott; Walters, Matthew; Sattar, Naveed; Dominiczak, Anna F; Padmanabhan, Sandosh

    2015-07-01

    There is accruing evidence from general population studies that serum bilirubin and liver enzymes affect blood pressure (BP) and cardiovascular risk, but it is unclear whether these have an impact on hypertensive patients in terms of long-term survival or BP control. We analyzed 12 000 treated hypertensive individuals attending a tertiary care clinic followed up for 35 years for association between baseline liver function tests and cause-specific mortality after adjustment for conventional cardiovascular covariates. Generalized estimating equations were used to study the association of liver tests and follow-up BP. The total time at risk was 173 806 person years with median survival 32.3 years. Follow-up systolic BP over 5 years changed by -0.4 (alanine transaminase and bilirubin), +2.1(alkaline phosphatase), +0.9(γ-glutamyl transpeptidase) mm Hg for each standard deviation increase. Serum total bilirubin and alanine transaminase showed a significant negative association with all-cause and cardiovascular mortality, whereas alkaline phosphatase and γ-glutamyl transpeptidase showed a positive association and aspartate transaminase showed a U-shapedassociation. Serum bilirubin showed an incremental improvement of continuous net reclassification improvement by 8% to 26% for 25 year and 35 year cardiovascular mortality, whereas all liver markers together improved continuous net reclassification improvement by 19% to 47% compared with reference model. In hypertensive patients, serum liver enzymes and bilirubin within 4 standard deviations of the mean show independent effects on mortality and BP control. Our findings would support further studies to elucidate the mechanisms by which liver enzymes and bilirubin may exert an effect on BP and cardiovascular risk, but there is little support for using them in risk stratification. PMID:25941342

  12. A structure-based catalytic mechanism for the xanthine oxidase family of molybdenum enzymes.

    PubMed Central

    Huber, R; Hof, P; Duarte, R O; Moura, J J; Moura, I; Liu, M Y; LeGall, J; Hille, R; Archer, M; Romão, M J

    1996-01-01

    The crystal structure of the xanthine oxidase-related molybdenum-iron protein aldehyde oxido-reductase from the sulfate reducing anaerobic Gram-negative bacterium Desulfovibrio gigas (Mop) was analyzed in its desulfo-, sulfo-, oxidized, reduced, and alcohol-bound forms at 1.8-A resolution. In the sulfo-form the molybdenum molybdopterin cytosine dinucleotide cofactor has a dithiolene-bound fac-[Mo, = O, = S, ---(OH2)] substructure. Bound inhibitory isopropanol in the inner compartment of the substrate binding tunnel is a model for the Michaelis complex of the reaction with aldehydes (H-C = O,-R). The reaction is proposed to proceed by transfer of the molybdenum-bound water molecule as OH- after proton transfer to Glu-869 to the carbonyl carbon of the substrate in concert with hydride transfer to the sulfido group to generate [MoIV, = O, -SH, ---(O-C = O, -R)). Dissociation of the carboxylic acid product may be facilitated by transient binding of Glu-869 to the molybdenum. The metal-bound water is replenished from a chain of internal water molecules. A second alcohol binding site in the spacious outer compartment may cause the strong substrate inhibition observed. This compartment is the putative binding site of large inhibitors of xanthine oxidase. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 PMID:8799115

  13. Human lysyl oxidase-like 2.

    PubMed

    Moon, Hee-Jung; Finney, Joel; Ronnebaum, Trey; Mure, Minae

    2014-12-01

    Lysyl oxidase like-2 (LOXL2) belongs to the lysyl oxidase (LOX) family, which comprises Cu(2+)- and lysine tyrosylquinone (LTQ)-dependent amine oxidases. LOXL2 is proposed to function similarly to LOX in the extracellular matrix (ECM) by promoting crosslinking of collagen and elastin. LOXL2 has also been proposed to regulate extracellular and intracellular cell signaling pathways. Dysregulation of LOXL2 has been linked to many diseases, including cancer, pro-oncogenic angiogenesis, fibrosis and heart diseases. In this review, we will give an overview of the current understandings and hypotheses regarding the molecular functions of LOXL2. PMID:25146937

  14. NADPH Oxidases and Angiotensin II Receptor Signaling

    PubMed Central

    Garrido, Abel Martin; Griendling, Kathy K.

    2010-01-01

    Over the last decade many studies have demonstrated the importance of reactive oxygen species (ROS) production by NADPH oxidases in angiotensin II (Ang II) signaling, as well as a role for ROS in the development of different diseases in which Ang II is a central component. In this review, we summarize the mechanism of activation of NADPH oxidases by Ang II and describe the molecular targets of ROS in Ang II signaling in the vasculature, kidney and brain. We also discuss the effects of genetic manipulation of NADPH oxidase function on the physiology and pathophysiology of the renin angiotensin system. PMID:19059306

  15. The role of gamma-aminobutyric acid/glycinergic synaptic transmission in mediating bilirubin-induced hyperexcitation in developing auditory neurons.

    PubMed

    Yin, Xin-Lu; Liang, Min; Shi, Hai-Bo; Wang, Lu-Yang; Li, Chun-Yan; Yin, Shan-Kai

    2016-01-01

    Hyperbilirubinemia is a common clinical phenomenon observed in human newborns. A high level of bilirubin can result in severe jaundice and bilirubin encephalopathy. However, the cellular mechanisms underlying bilirubin excitotoxicity are unclear. Our previous studies showed the action of gamma-aminobutyric acid (GABA)/glycine switches from excitatory to inhibitory during development in the ventral cochlear nucleus (VCN), one of the most sensitive auditory nuclei to bilirubin toxicity. In the present study, we investigated the roles of GABAA/glycine receptors in the induction of bilirubin hyperexcitation in early developing neurons. Using the patch clamp technique, GABAA/glycine receptor-mediated spontaneous inhibitory synaptic currents (sIPSCs) were recorded from bushy and stellate cells in acute brainstem slices from young mice (postnatal day 2-6). Bilirubin significantly increased the frequency of sIPSCs, and this effect was prevented by pretreatments of slices with either fast or slow Ca(2+) chelators BAPTA-AM and EGTA-AM suggesting that bilirubin can increase the release of GABA/glycine via Ca(2+)-dependent mechanisms. Using cell-attached recording configuration, we found that antagonists of GABAA and glycine receptors strongly attenuated spontaneous spiking firings in P2-6 neurons but produced opposite effect in P15-19 neurons. Furthermore, these antagonists reversed bilirubin-evoked hyperexcitability in P2-6 neurons, indicating that excitatory action of GABA/glycinergic transmission specifically contribute to bilirubin-induced hyperexcitability in the early stage of development. Our results suggest that bilirubin-induced enhancement of presynaptic release GABA/Glycine via Ca(2+)-dependent mechanisms may play a critical role in mediating neuronal hyperexcitation associated with jaundice, implicating potential new strategies for predicting, preventing, and treating bilirubin neurotoxicity. PMID:26476400

  16. Crystal Structure of the Deglycating Enzyme Fructosamine Oxidase (Amadoriase II)

    SciTech Connect

    Collard, François; Zhang, Jianye; Nemet, Ina; Qanungo, Kaustubha R.; Monnier, Vincent M.; Yee, Vivien C.

    2009-01-12

    Fructosamine oxidases (FAOX) catalyze the oxidative deglycation of low molecular weight fructosamines (Amadori products). These proteins are of interest in developing an enzyme to deglycate proteins implicated in diabetic complications. We report here the crystal structures of FAOX-II from the fungi Aspergillus fumigatus, in free form and in complex with the inhibitor fructosyl-thioacetate, at 1.75 and 1.6{angstrom} resolution, respectively. FAOX-II is a two domain FAD-enzyme with an overall topology that is most similar to that of monomeric sarcosine oxidase. Active site residues Tyr-60, Arg-112 and Lys-368 bind the carboxylic portion of the fructosamine, whereas Glu-280 and Arg-411 bind the fructosyl portion. From structure-guided sequence comparison, Glu-280 was identified as a signature residue for FAOX activity. Two flexible surface loops become ordered upon binding of the inhibitor in a catalytic site that is about 12{angstrom} deep, providing an explanation for the very low activity of FAOX enzymes toward protein-bound fructosamines, which would have difficulty accessing the active site. Structure-based mutagenesis showed that substitution of Glu-280 and Arg-411 eliminates enzyme activity. In contrast, modification of other active site residues or of amino acids in the flexible active site loops has little effect, highlighting these regions as potential targets in designing an enzyme that will accept larger substrates.

  17. Crystal Structure of the Deglycating Enzyme Fructosamine Oxidase (Amadoriase ll)

    SciTech Connect

    Collard, F.; Zhang, J; Nemet, I; Qanungo, K; Monnier, V; Yee, V

    2008-01-01

    Fructosamine oxidases (FAOX) catalyze the oxidative deglycation of low molecular weight fructosamines (Amadori products). These proteins are of interest in developing an enzyme to deglycate proteins implicated in diabetic complications. We report here the crystal structures of FAOX-II from the fungi Aspergillus fumigatus, in free form and in complex with the inhibitor fructosyl-thioacetate, at 1.75 and 1.6{angstrom} resolution, respectively. FAOX-II is a two domain FAD-enzyme with an overall topology that is most similar to that of monomeric sarcosine oxidase. Active site residues Tyr-60, Arg-112 and Lys-368 bind the carboxylic portion of the fructosamine, whereas Glu-280 and Arg-411 bind the fructosyl portion. From structure-guided sequence comparison, Glu-280 was identified as a signature residue for FAOX activity. Two flexible surface loops become ordered upon binding of the inhibitor in a catalytic site that is about 12{angstrom} deep, providing an explanation for the very low activity of FAOX enzymes toward protein-bound fructosamines, which would have difficulty accessing the active site. Structure-based mutagenesis showed that substitution of Glu-280 and Arg-411 eliminates enzyme activity. In contrast, modification of other active site residues or of amino acids in the flexible active site loops has little effect, highlighting these regions as potential targets in designing an enzyme that will accept larger substrates.

  18. Crystal Structure of the Deglycating Enzyme Fructosamine Oxidase (Amadoriase II)*

    PubMed Central

    Collard, François; Zhang, Jianye; Nemet, Ina; Qanungo, Kaustubha R.; Monnier, Vincent M.; Yee, Vivien C.

    2008-01-01

    Fructosamine oxidases (FAOX) catalyze the oxidative deglycation of low molecular weight fructosamines (Amadori products). These proteins are of interest in developing an enzyme to deglycate proteins implicated in diabetic complications. We report here the crystal structures of FAOX-II from the fungi Aspergillus fumigatus, in free form and in complex with the inhibitor fructosyl-thioacetate, at 1.75 and 1.6Å resolution, respectively. FAOX-II is a two domain FAD-enzyme with an overall topology that is most similar to that of monomeric sarcosine oxidase. Active site residues Tyr-60, Arg-112 and Lys-368 bind the carboxylic portion of the fructosamine, whereas Glu-280 and Arg-411 bind the fructosyl portion. From structure-guided sequence comparison, Glu-280 was identified as a signature residue for FAOX activity. Two flexible surface loops become ordered upon binding of the inhibitor in a catalytic site that is about 12Å deep, providing an explanation for the very low activity of FAOX enzymes toward protein-bound fructosamines, which would have difficulty accessing the active site. Structure-based mutagenesis showed that substitution of Glu-280 and Arg-411 eliminates enzyme activity. In contrast, modification of other active site residues or of amino acids in the flexible active site loops has little effect, highlighting these regions as potential targets in designing an enzyme that will accept larger substrates. PMID:18667417

  19. Synthesis and characterization of a biospecific adsorbent containing bovine serum albumin as a ligand and its use for bilirubin retention.

    PubMed

    Alvarez, C; Strumia, M; Bertorello, H

    2001-10-30

    Epoxy-activated gels from cross-linked polybutadiene-hydroxyethylmethacrylate (PB-HEMA) copolymer and epichlorohydrin (ECH) were prepared and characterized. Albumin was covalently bonded to the matrix and used as support of affinity chromatography in bilirubin (BR) retention experiments. PB-HEMA-ECH with different amounts of immobilized albumin (between 5.20 and 6.80 mg/g dry gel) were obtained. Bilirubin retention of 3.10 mg/g of these beads was observed at 5 degrees C. PMID:11694308

  20. Bounding Species Distribution Models

    NASA Technical Reports Server (NTRS)

    Stohlgren, Thomas J.; Jarnevich, Cahterine S.; Morisette, Jeffrey T.; Esaias, Wayne E.

    2011-01-01

    Species distribution models are increasing in popularity for mapping suitable habitat for species of management concern. Many investigators now recognize that extrapolations of these models with geographic information systems (GIS) might be sensitive to the environmental bounds of the data used in their development, yet there is no recommended best practice for "clamping" model extrapolations. We relied on two commonly used modeling approaches: classification and regression tree (CART) and maximum entropy (Maxent) models, and we tested a simple alteration of the model extrapolations, bounding extrapolations to the maximum and minimum values of primary environmental predictors, to provide a more realistic map of suitable habitat of hybridized Africanized honey bees in the southwestern United States. Findings suggest that multiple models of bounding, and the most conservative bounding of species distribution models, like those presented here, should probably replace the unbounded or loosely bounded techniques currently used [Current Zoology 57 (5): 642-647, 2011].

  1. Bounding species distribution models

    USGS Publications Warehouse

    Stohlgren, T.J.; Jarnevich, C.S.; Esaias, W.E.; Morisette, J.T.

    2011-01-01

    Species distribution models are increasing in popularity for mapping suitable habitat for species of management concern. Many investigators now recognize that extrapolations of these models with geographic information systems (GIS) might be sensitive to the environmental bounds of the data used in their development, yet there is no recommended best practice for "clamping" model extrapolations. We relied on two commonly used modeling approaches: classification and regression tree (CART) and maximum entropy (Maxent) models, and we tested a simple alteration of the model extrapolations, bounding extrapolations to the maximum and minimum values of primary environmental predictors, to provide a more realistic map of suitable habitat of hybridized Africanized honey bees in the southwestern United States. Findings suggest that multiple models of bounding, and the most conservative bounding of species distribution models, like those presented here, should probably replace the unbounded or loosely bounded techniques currently used. ?? 2011 Current Zoology.

  2. Causality and Tsirelson's bounds

    SciTech Connect

    Buhrman, H.; Massar, S.

    2005-11-15

    We study the properties of no-signaling correlations that cannot be reproduced by local measurements on entangled quantum states. We say that such correlations violate Tsirelson bounds. We show that if these correlations are obtained by some reversible unitary quantum evolution U, then U cannot be written in the product form U{sub A}xU{sub B}. This implies that U can be used for signaling and for entanglement generation. This result is completely general and in fact can be viewed as a characterization of Tsirelson bounds. We then show how this result can be used as a tool to study Tsirelson bounds and we illustrate this by rederiving the Tsirelson bound of 2{radical}(2) for the Clauser-Horn-Shimony-Holt inequality, and by deriving a new Tsirelson bound for qutrits.

  3. Regulation of NADPH oxidases in skeletal muscle.

    PubMed

    Ferreira, Leonardo F; Laitano, Orlando

    2016-09-01

    The only known function of NAD(P)H oxidases is to produce reactive oxygen species (ROS). Skeletal muscles express three isoforms of NAD(P)H oxidases (Nox1, Nox2, and Nox4) that have been identified as critical modulators of redox homeostasis. Nox2 acts as the main source of skeletal muscle ROS during contractions, participates in insulin signaling and glucose transport, and mediates the myocyte response to osmotic stress. Nox2 and Nox4 contribute to skeletal muscle abnormalities elicited by angiotensin II, muscular dystrophy, heart failure, and high fat diet. Our review addresses the expression and regulation of NAD(P)H oxidases with emphasis on aspects that are relevant to skeletal muscle. We also summarize: i) the most widely used NAD(P)H oxidases activity assays and inhibitors, and ii) studies that have defined Nox enzymes as protagonists of skeletal muscle redox homeostasis in a variety of health and disease conditions. PMID:27184955

  4. Activation of Polyphenol Oxidase of Chloroplasts 1

    PubMed Central

    Tolbert, N. E.

    1973-01-01

    Polyphenol oxidase of leaves is located mainly in chloroplasts isolated by differential or sucrose density gradient centrifugation. This activity is part of the lamellar structure that is not lost on repeated washing of the plastids. The oxidase activity was stable during prolonged storage of the particles at 4 C or —18 C. The Km (dihydroxyphenylalanine) for spinach leaf polyphenol oxidase was 7 mm by a spectrophotometric assay and 2 mm by the manometric assay. Polyphenol oxidase activity in the leaf peroxisomal fraction, after isopycnic centrifugation on a linear sucrose gradient, did not coincide with the peroxisomal enzymes but was attributed to proplastids at nearly the same specific density. Plants were grouped by the latency properties for polyphenol oxidase in their isolated chloroplasts. In a group including spinach, Swiss chard, and beet leaves the plastids immediately after preparation from fresh leaves required a small amount of light for maximal rates of oxidation of dihydroxyphenylalanine. Polyphenol oxidase activity in the dark or light increased many fold during aging of these chloroplasts for 1 to 5 days. Soluble polyphenol oxidase of the cytoplasm was not so stimulated. Chloroplasts prepared from stored leaves were also much more active than from fresh leaves. Maximum rates of dihydroxyphenylalanine oxidation were 2 to 6 mmoles × mg−1 chlorophyll × hr−1. Equal stimulation of latent polyphenol oxidase in fresh or aged chloroplasts in this group was obtained by either light, an aged trypsin digest, 3-(4-chlorophenyl)-1, 1-dimethylurea, or antimycin A. A variety of other treatments did not activate or had little effect on the oxidase, including various peptides, salts, detergents, and other proteolytic enzymes. Activation of latent polyphenol oxidase in spinach chloroplasts by trypsin amounted to as much as 30-fold. The trypsin activation occurred even after the trypsin had been treated with 10% trichloroacetic acid, 1.0 n HCl or boiled for 30

  5. D-amino acid oxidase from the yeast Trigonopsis variabilis.

    PubMed

    Kubicek-Pranz, E M; Röhr, M

    1985-04-01

    D-Amino acid oxidase (EC 1.4.3.3) has been purified from the yeast Trigonopsis variabilis by the application of ion-exchange chromatography on DEAE-cellulose, salt precipitation, gel filtration, and hydroxyapatite adsorption. Alternatively the last two steps can be substituted by a single fast protein liquid chromatographic ion-exchange step (Mono Q). The enzyme appeared homogeneous in PAGE, but small amounts of impurities (not exceeding 5% of total protein) appeared in sodium dodecyl sulfate (SDS)-PAGE. Its Mr in SDS-PAGE is 39,000; it exhibits an isoelectric point of 4.8 and contains 7% (w/v) covalently bound carbohydrate. Its absorption spectrum is similar to hog kidney D-amino acid oxidase, indicating the presence of bound FAD, which, however, could not be separated from the enzyme under non-denaturing conditions. The enzyme is inhibited by SH-oxidizing agents, but not by metal-chelate formers and not by benzoate or toluene. It uses O2 exclusively as the only H acceptor. Km and Vmax values were determined for 15 D-amino acids, which, among 23 tested, were substrates of the enzyme. The enzyme has highest affinity for D-phenylalanine and D-leucine, but maximal activity is obtained with D-citrulline and D-isoleucine. The specific activity of the purified preparation is even higher than that of the commercially available hog kidney enzyme (21.7 vs 16 U/mg). The yeast enzyme may be a useful analytical and preparative tool in view of the difference between its substrate specificity and that of the hog enzyme. PMID:2865242

  6. Bilirubin as a Protective Factor for Rheumatoid Arthritis: An NHANES Study of 2003 - 2006 Data

    PubMed Central

    Fischman, Daniel; Valluri, Ashok; Gorrepati, Venkata Subhash; Murphy, Megan E.; Peters, Ian; Cheriyath, Pramil

    2010-01-01

    Background Rheumatoid arthritis(RA) is a chronic inflammatory, autoimmune polyarthritis, with a prevalence estimated at one percent of the United States(US) population. Serum bilirubin, because of its antioxidant nature, has been conjectured to exert an anti-inflammatory biologic effect. The objective of this study is to discern whether higher serum Total Bilirubin(TBili) levels are protective against RA. Methods This is a secondary analysis of National Health and Nutrition Examination Survey (NHANES) data collected between 2003-2006. Study participants completed a comprehensive questionnaire regarding their health history, underwent a physical examination, and had body fluids collected for laboratory studies. In NHANES, to assess for the presence of RA, the following questions were asked: "Doctor ever said you had arthritis?" If so, "Which type of arthritis". Statistical analysis was performed, using SAS version 9.1, proc survey methods. Participant data were adjusted for demographic characteristics as well as risk factors for RA. Results NHANES 2003-2006 included 20,470 individuals, chosen as a representative sampling of the entire US population. Exclusion criteria included age less than twenty years or liver dysfunction, defined as history of abnormal liver function tests or liver disease. 8,147 subjects did not have any exclusion criteria and were included in the data analysis. RA is inversely related to the serum level of TBili with an odds ratio of 0.679 (95% CI 0.533-0.865) and remained significant even after adjusting for age, gender, race, education, and tobacco history, with an odds ratio 0.749 (95% CI 0.575 - 0.976). Conclusions Our study supports the hypothesis that higher TBili levels are protective against RA. A plausible mechanism for this association would be that the anti-oxidant effects of TBili exert a physiologic anti-inflammatory effect, which provides protection against RA. This explanation is supported by prior studies which show that higher

  7. Influence of glutathione S-transferase B (ligandin) on the intermembrane transfer of bilirubin. Implications for the intracellular transport of nonsubstrate ligands in hepatocytes.

    PubMed Central

    Zucker, S D; Goessling, W; Ransil, B J; Gollan, J L

    1995-01-01

    To examine the hypothesis that glutathione S-transferases (GST) play an important role in the hepatocellular transport of hydrophobic organic anions, the kinetics of the spontaneous transfer of unconjugated bilirubin between membrane vesicles and rat liver glutathione S-transferase B (ligandin) was studied, using stopped-flow fluorometry. Bilirubin transfer from glutathione S-transferase B to phosphatidylcholine vesicles was best described by a single exponential function, with a rate constant of 8.0 +/- 0.7 s-1 (+/- SD) at 25 degrees C. The variations in transfer rate with respect to acceptor phospholipid concentration provide strong evidence for aqueous diffusion of free bilirubin. This finding was verified using rhodamine-labeled microsomal membranes as acceptors. Bilirubin transfer from phospholipid vesicles to GST also exhibited diffusional kinetics. Thermodynamic parameters for bilirubin dissociation from GST were similar to those for human serum albumin. The rate of bilirubin transfer from rat liver basolateral plasma membranes to acceptor vesicles in the presence of glutathione S-transferase B declined asymptotically with increasing GST concentration. These data suggest that glutathione S-transferase B does not function as an intracellular bilirubin transporter, although expression of this protein may serve to regulate the delivery of bilirubin, and other nonsubstrate ligands, to sites of metabolism within the cell. Images PMID:7560084

  8. Smoking Cessation Is Followed by Increases in Serum Bilirubin, an Endogenous Antioxidant Associated With Lower Risk of Lung Cancer and Cardiovascular Disease

    PubMed Central

    Wu, Ran; Mayne, Susan T.; Jatlow, Peter I.

    2014-01-01

    Introduction: Lower concentrations of serum bilirubin, an endogenous antioxidant, have been associated with risk of many smoking-related diseases, including lung cancer and cardiovascular disease, and current smokers are reported to have lower bilirubin levels than nonsmokers and past smokers. This study evaluates the effects of smoking cessation on bilirubin levels. Methods: In a secondary analysis of a 6-week placebo-controlled trial of naltrexone for smoking cessation, indirect and total bilirubin concentrations were evaluated at baseline and following smoking cessation. Individuals who were continuously abstinent for 6 weeks (n = 155) were compared to those who were not (n = 193). Participants reported smoking ≥20 cigarettes daily at baseline and received smoking cessation counseling, 21mg nicotine patch daily, and either placebo or 1 of 3 doses of naltrexone (25, 50, or 100mg) for 6 weeks. Change in indirect and total bilirubin following the quit date was measured at Weeks 1, 4, and 6 compared to baseline. Results: Individuals who were continuously abstinent from smoking, independent of naltrexone condition, showed a significantly greater mean increase in indirect (~unconjugated) bilirubin (0.06mg/dl, SD = 0.165) compared to those who did not (mean = 0.02, SD = 0.148, p = .015). Similar results were obtained for total bilirubin (p = .037). Conclusions: Smoking cessation is followed by increases in bilirubin concentration that have been associated with lower risk of lung cancer and cardiovascular disease. PMID:24812024

  9. Bound infragravity waves

    NASA Astrophysics Data System (ADS)

    Okihiro, Michele; Guza, R. T.; Seymour, R. J.

    1992-07-01

    Model predictions of bound (i.e., nonlinearly forced by and coupled to wave groups) infragravity wave energy are compared with about 2 years of observations in 8- to 13-m depths at Imperial Beach, California, and Barbers Point, Hawaii. Frequency-directional spectra of free waves at sea and swell frequencies, estimated with a small array of four pressure sensors, are used to predict the bound wave spectra below 0.04 Hz. The predicted total bound wave energy is always less than the observed infragravity energy, and the underprediction increases with increasing water depth and especially with decreasing swell energy. At most half, and usually much less, of the observed infragravity energy is bound. Bound wave spectra are also predicted with data from a single wave gage in 183-m depth at Point Conception, California, and the assumption of unidirectional sea and swell. Even with energetic swell, less than 10% of the total observed infragravity energy in 183-m depth is bound. Free waves, either leaky or edge waves, are more energetic than bound waves at both the shallow and deep sites. The low level of infragravity energy observed in 183-m depth compared with 8- to 13-m depths, with similarly moderate sea and swell energy, suggests that leaky (and very high-mode edge) waves contribute less than 10% of the infragravity energy in 8-13 m. Most of the free infragravity energy in shallow water is refractively trapped and does not reach deep water.

  10. Bilirubin conjugates of human bile. The excretion of bilirubin as the acyl glycosides of aldobiouronic acid, pseudoaldobiouronic acid and hexuronosylhexuronic acid, with a branched-chain hexuronic acid as one of the components of hexuronosylhexuronide

    PubMed Central

    Kuenzle, Clive C.

    1970-01-01

    Structure elucidations have been performed on the bilirubin conjugates isolated from human hepatic bile as the phenylazo derivatives. The major bilirubin conjugates are excreted, not as was formerly thought in the form of glucuronides, but as the acyl glycosides of aldobiouronic acid, pseudoaldobiouronic acid and hexuronosylhexuronic acid. The isolated aldobiouronides are proposed to have the structures of an acyl 6-O-hexopyranosyluronic acid-hexopyranoside, an acyl 4-O-hexofuranosyluronic acid-d-glucopyranoside, and an acyl 4-O-β-d-glucofuranosyluronic acid-d-glucopyranoside respectively, with the acyl radicals being those of the phenylazo derivative of bilirubin. The pseudoaldobiouronide is suggested to be the acyl 4-O-α-d-glucofuranosyl-β-d -glucopyranosiduronic acid, with the acyl radical being that of the phenylazo derivative of vinylneoxanthobilirubinic acid. The hexuronosylhexuronide presumably is the acyl 4-O-(3-C-hydroxymethylribofuranosyluronic acid)-β-d-glucopyranosiduronic acid, with the acyl radical being that of the phenylazo derivative of bilirubin. The 3-C-hydroxymethylriburonic acid, isolated as one of the components of the hexuronosylhexuronide, is the first natural branched-chain hexuronic acid to be detected, and the first branched-chain sugar ever detected in humans. PMID:5500303

  11. Reductive trapping of substrate to bovine plasma amine oxidase

    SciTech Connect

    Hartmann, C.; Klinman, J.P.

    1987-01-25

    Plasma amine oxidases catalyze the oxidative deamination of amines to aldehydes, followed by a 2e- reduction of O/sub 2/ to H/sub 2/O/sub 2/. Pyrroloquinoline quinone (PQQ), previously believed to be restricted to prokaryotes, has recently been proposed to be the cofactor undergoing reduction in the first half-reaction of bovine plasma amine oxidase (Ameyama, M., Hayashi, U., Matsushita, K., Shinagawa, E., and Adachi, O. (1984) Agric. Biol. Chem. 48, 561-565; Lobenstein-Verbeek, C. L., Jongejan, J. A., Frank, J., and Duine, J. A. (1984) FEBS Lett. 170, 305-309). This result is unexpected, since model studies with PQQ implicate Schiff's base formation between a reactive carbonyl and substrates, whereas experiments with bovine plasma amine oxidase have failed to provide evidence for a carbonyl cofactor. We have, therefore, re-examined putative adducts between substrate and enzyme-bound cofactor, employing a combination of (/sup 14/C)benzylamine and (/sup 3/H)NaCNBH/sub 3/. The use of the relatively weak reductant, NaCNBH/sub 3/, affords Schiff's base specificity and permits the study of enzyme below pH 7.0. As we show, enzyme can only be inactivated by NaCNBH/sub 3/ in the presence of substrate, leading to the incorporation of 1 mol of (/sup 14/C)benzylamine/mol of enzyme subunit at complete inactivation. By contrast, we are unable to detect any labeling with (/sup 3/H)NaCNBH/sub 3/, analogous to an earlier study with (/sup 3/H)NaCNBH/sub 4/ (Suva, R. H., and Abeles, R. H. (1978) Biochemistry 17, 3538-3545). We conclude, first, that our inability to obtain adducts containing both carbon 14 and tritium rules out the reductive trapping either of amine substrate with pyridoxal phosphate or of aldehyde product with a lysyl side chain and, second, that the observed pattern of labeling is fully consistent with the presence of PQQ at the active site of bovine plasma amine oxidase.

  12. Multiple Genetic Modifiers of Bilirubin Metabolism Involvement in Significant Neonatal Hyperbilirubinemia in Patients of Chinese Descent

    PubMed Central

    Yang, Hui; Zheng, Lei; Lin, Min; Zheng, Xiang-bin; Lin, Fen

    2015-01-01

    The potential for genetic variation to modulate neonatal hyperbilirubinemia risk is increasingly being recognized. A case-control study was designed to assess comprehensive contributions of the multiple genetic modifiers of bilirubin metabolism on significant neonatal hyperbilirubinemia in Chinese descendents. Eleven common mutations and polymorphisms across five bilirubin metabolism genes, namely those encoding UGT1A1, HMOX1, BLVRA, SLCO1B1 and SLCO1B3, were determined using the high resolution melt (HRM) assay or PCR-capillary electrophoresis analysis. A total of 129 hyperbilirubinemic infants and 108 control subjects were evaluated. Breastfeeding and the presence of the minor A allele of rs4148323 (UGTA*6) were correlated with an increased risk of hyperbilirubinemia (OR=2.17, P=0.02 for breastfeeding; OR=9.776, P=0.000 for UGTA*6 homozygote; OR=3.151, P=0.000 for UGTA*6 heterozygote); whereas, increasing gestational age and the presence of –TA7 repeat variant of UGT1A1 decreased the risk (OR=0.721, P=0.003 for gestational age; OR=0.313, P=0.002 for heterozygote TA6/TA7). In addition, the SLCO1B1 and SLCO1B3 polymorphisms also contributed to an increased risk of hyperbilirubinemia. This detailed analysis revealed the impact of multiple genetic modifiers on neonatal hyperbilirubinemia. This may support the use of genetic tests for clinical risk assessment. Furthermore, the established HRM assay can serve as an effective method for large-scale investigation. PMID:26146841

  13. Heme oxygenase-1-derived bilirubin counteracts HIV protease inhibitor-mediated endothelial cell dysfunction.

    PubMed

    Liu, Xiao-Ming; Durante, Zane E; Peyton, Kelly J; Durante, William

    2016-05-01

    The use of HIV protease inhibitors (PIs) has extended the duration and quality of life for HIV-positive individuals. However there is increasing concern that this antiviral therapy may promote premature cardiovascular disease by impairing endothelial cell (EC) function. In the present study, we investigated the effect of HIV PIs on EC function and determined if the enzyme heme oxygenase (HO-1) influences the biological action of these drugs. We found that three distinct PIs, including ritonavir, atazanavir, and lopinavir, stimulated the expression of HO-1 protein and mRNA. The induction of HO-1 was associated with an increase in NF-E2-related factor-2 (Nrf2) activity and reactive oxygen species (ROS). PIs also stimulated HO-1 promoter activity and this was prevented by mutating the antioxidant responsive element or by overexpressing dominant-negative Nrf2. In addition, the PI-mediated induction of HO-1 was abolished by N-acetyl-l-cysteine and rotenone. Furthermore, PIs blocked EC proliferation and migration and stimulated the expression of intercellular adhesion molecule-1 and the adhesion of monocytes on ECs. Inhibition of HO-1 activity or expression potentiated the anti-proliferative and inflammatory actions of PIs which was reversed by bilirubin but not carbon monoxide. Alternatively, adenovirus-mediated overexpression of HO-1 attenuated the growth-inhibitory and inflammatory effect of PIs. In contrast, blocking HO-1 activity failed to modify the anti-migratory effect of the PIs. Thus, induction of HO-1 via the ROS-Nrf2 pathway in human ECs counteracts the anti-proliferative and inflammatory actions of PIs by generating bilirubin. Therapeutic approaches targeting HO-1 may provide a novel approach in preventing EC dysfunction and vascular disease in HIV-infected patients undergoing antiretroviral therapy. PMID:26968795

  14. Forage polyphenol oxidase and ruminant livestock nutrition

    PubMed Central

    Lee, Michael R. F.

    2014-01-01

    Polyphenol oxidase (PPO) is predominately associated with the detrimental effect of browning fruit and vegetables, however, interest within PPO containing forage crops (crops to be fed to animals) has grown since the browning reaction was associated with reduced nitrogen (N) losses in silo and the rumen. The reduction in protein breakdown in silo of red clover (high PPO forage) increased the quality of protein, improving N-use efficiency [feed N into product N (e.g., Milk): NUE] when fed to ruminants. A further benefit of red clover silage feeding is a significant reduction in lipolysis (cleaving of glycerol-based lipid) in silo and an increase in the deposition of beneficial C18 polyunsaturated fatty acid (PUFA) in animal products, which has also been linked to PPO activity. PPOs protection of plant protein and glycerol based-PUFA in silo is related to the deactivation of plant proteases and lipases. This deactivation occurs through PPO catalyzing the conversion of diphenols to quinones which bind with cellular nucleophiles such as protein reforming a protein-bound phenol (PBP). If the protein is an enzyme (e.g., protease or lipase) the complexing denatures the enzyme. However, PPO is inactive in the anaerobic rumen and therefore any subsequent protection of plant protein and glycerol based-PUFA in the rumen must be as a result of events that occurred to the forage pre-ingestion. Reduced activity of plant proteases and lipases would have little effect on NUE and glycerol based-PUFA in the rumen due to the greater concentration of rumen microbial proteases and lipases. The mechanism for PPOs protection of plant protein in the rumen is a consequence of complexing plant protein, rather than protease deactivation per se. These complexed proteins reduce protein digestibility in the rumen and subsequently increase undegraded dietary protein flow to the small intestine. The mechanism for protecting glycerol-based PUFA has yet to be fully elucidated but may be associated

  15. BILIRUBIN CONCENTRATIONS IN CLINICALLY HEALTHY AND DISEASED CAPTIVE WATERBUCK (KOBUS ELLIPSIPRYMNUS) AT THE SAN DIEGO ZOO SAFARI PARK.

    PubMed

    Sadler, Ryan A; Lamberski, Nadine; Christopher, Mary M

    2016-06-01

    Captive waterbuck ( Kobus ellipsiprymnus ) that appear clinically healthy have been noted to have high serum bilirubin concentrations compared with other ruminants; however, questions remain about the physiologic factors affecting bilirubin concentration and its potential association with underlying disease and icteric serum or mucous membranes. Serum bilirubin concentrations of healthy and diseased waterbuck housed at the San Diego Zoo Safari Park from 1989 to 2012 were retrospectively analyzed to determine any link between icteric serum, total bilirubin concentration (tBili), and disease entities in this species. Total bilirubin and direct (dBili) bilirubin concentrations and the prevalence of icteric serum were compared by subspecies, age group, and health status; associations with complete blood count and biochemical results and clinical diagnosis were assessed. No significant differences were found in tBili or dBili between Ellipsen (n = 32) and Defassa (n = 29) subspecies or in juveniles (n = 22) versus adults (n = 39). Clinically healthy waterbuck (n = 40) had significantly higher tBili (mean ± 2SD, 7.9 ± 1.2 mg/dl; P < 0.001) and dBili (3.7 ± 1.0 mg/dl; P < 0.001) than did diseased waterbuck (n = 21; tBili: 4.9 ± 2.56 mg/dl; dBili: 2.2 ± 0.8 mg/dl). No waterbuck had icteric tissues on physical examination. Twelve (19.7%) waterbuck (six healthy, six diseased) had icteric serum. Few minor correlations were seen between tBili or dBili and clinical, laboratory, or necropsy evidence of disease, though an inverse correlation between dBili and blood glucose was noted. Of the 40 healthy animals, reference intervals were calculated for tBili (5.5-10.3 mg/dl), dBili (1.7-5.7 mg/dl), and indirect bilirubin (2.2-6.2 mg/dl). These results suggest healthy waterbuck have relatively high tBili and dBili compared with related species. Icteric serum may be seen in up to 15% of healthy animals in the absence of icteric tissues. PMID:27468025

  16. Serum Bilirubin and 6-min Walk Distance as Prognostic Predictors for Inoperable Chronic Thromboembolic Pulmonary Hypertension: A Prospective Cohort Study

    PubMed Central

    Gong, Juan-Ni; Zhai, Zhen-Guo; Yang, Yuan-Hua; Liu, Yan; Gu, Song; Kuang, Tu-Guang; Xie, Wan-Mu; Miao, Ran; Wang, Chen

    2015-01-01

    Background: Inoperable chronic thromboembolic pulmonary hypertension (CTEPH) is a severe clinical syndrome characterized by right cardiac failure and possibly subsequent liver dysfunction. However, whether serum markers of liver dysfunction can predict prognosis in inoperable CTEPH patients has not been determined. Our study aimed to evaluate the potential role of liver function markers (such as serum levels of transaminase, bilirubin, and gamma-glutamyl transpeptidase [GGT]) combined with 6-min walk test in the prediction of prognosis in patients with inoperable CTEPH. Methods: From June 2005 to May 2013, 77 consecutive patients with inoperable CTEPH without confounding co-morbidities were recruited for this prospective cohort study. Baseline clinical characteristics and 6-min walk distance (6MWD) results were collected. Serum biomarkers of liver function, including levels of aspartate aminotransferase, alanine aminotransferase, GGT, uric acid, and serum bilirubin, were also determined at enrollment. All-cause mortality was recorded during the follow-up period. Results: During the follow-up, 22 patients (29%) died. Cox regression analyses demonstrated that increased serum concentration of total bilirubin (hazard ratio [HR] = 7.755, P < 0.001), elevated N-terminal of the prohormone brain natriuretic peptide (HR = 1.001, P = 0.001), decreased 6MWD (HR = 0.990, P < 0.001), increased central venous pressure (HR = 1.074, P = 0.040), and higher pulmonary vascular resistance (HR = 1.001, P = 0.018) were associated with an increased risk of mortality. Serum concentrations of total bilirubin (HR = 4.755, P = 0.007) and 6MWD (HR = 0.994, P = 0.017) were independent prognostic predictors for CTEPH patients. Patients with hyperbilirubinemia (≥23.7 μmol/L) had markedly worse survival than those with normobilirubinemia. Conclusion: Elevated serum bilirubin and decreased 6MWD are potential predictors for poor prognosis in inoperable CTEPH. PMID:26612283

  17. Proline dehydrogenase (oxidase) in cancer.

    PubMed

    Liu, Wei; Phang, James M

    2012-01-01

    Proline dehydrogenase (oxidase, PRODH/POX), the first enzyme in the proline degradative pathway, plays a special role in tumorigenesis and tumor development. Proline metabolism catalyzed by PRODH/POX is closely linked with the tricarboxylic acid (TCA) cycle and urea cycle. The proline cycle formed by the interconversion of proline and Δ(1) -pyrroline-5-carboxylate (P5C) between mitochondria and cytosol interlocks with pentose phosphate pathway. Importantly, by catalyzing proline to P5C, PRODH/POX donates electrons into the electron transport chain to generate ROS or ATP. In earlier studies, we found that PRODH/POX functions as a tumor suppressor to initiate apoptosis, inhibit tumor growth, and block the cell cycle, all by ROS signaling. It also suppresses hypoxia inducible factor signaling by increasing α-ketoglutarate. During tumor progression, PRODH/POX is under the control of various tumor-associated factors, such as tumor suppressor p53, inflammatory factor peroxisome proliferator-activated receptor gamma (PPARγ), onco-miRNA miR-23b*, and oncogenic transcription factor c-MYC. Recent studies revealed the two-sided features of PRODH/POX-mediated regulation. Under metabolic stress such as oxygen and glucose deprivation, PRODH/POX can be induced to serve as a tumor survival factor through ATP production or ROS-induced autophagy. The paradoxical roles of PRODH/POX can be understood considering the temporal and spatial context of the tumor. Further studies will provide additional insights into this protein and on its metabolic effects in tumors, which may lead to new therapeutic strategies. PMID:22886911

  18. The composition of milk xanthine oxidase

    PubMed Central

    Hart, L. I.; McGartoll, Mary A.; Chapman, Helen R.; Bray, R. C.

    1970-01-01

    The composition of milk xanthine oxidase has been reinvestigated. When the enzyme is prepared by methods that include a selective denaturation step in the presence of sodium salicylate the product is obtained very conveniently and in high yield, and is homogeneous in the ultracentrifuge and in recycling gel filtration. It has specific activity higher than previously reported preparations of the enzyme and its composition approximates closely to 2mol of FAD, 2g-atoms of Mo and 8g-atoms of Fe/mol of protein (molecular weight about 275000). In contrast, when purely conventional preparative methods are used the product is also homogeneous by the above criteria but has a lower specific activity and is generally comparable to the crystallized enzyme described previously. Such samples also contain 2mol of FAD/mol of protein but they have lower contents of Mo (e.g. 1.2g-atom/mol). Amino acid compositions for the two types of preparation are indistinguishable. These results confirm the previous conclusion that conventional methods give mixtures of xanthine oxidase with an inactive modification of the enzyme now termed `de-molybdo-xanthine oxidase', and show that salicylate can selectively denature the latter. The origin of de-molybdo-xanthine oxidase was investigated. FAD/Mo ratios show that it is present not only in enzyme purified by conventional methods but also in `milk microsomes' (Bailie & Morton, 1958) and in enzyme samples prepared without proteolytic digestion. We conclude that it is secreted by cows together with the active enzyme and we discuss its occurrence in the preparations of other workers. Studies on the milks of individual cows show that nutritional rather than genetic factors determine the relative amounts of xanthine oxidase and de-molybdo-xanthine oxidase. A second inactive modification of the enzyme, now termed `inactivated xanthine oxidase', causes variability in activity relative to E450 or to Mo content and formation of it decreases these ratios

  19. Mediated electron transfer of cellobiose dehydrogenase and glucose oxidase at osmium polymer-modified nanoporous gold electrodes.

    PubMed

    Salaj-Kosla, Urszula; Scanlon, Micheál D; Baumeister, Tobias; Zahma, Kawah; Ludwig, Roland; Ó Conghaile, Peter; MacAodha, Domhnall; Leech, Dónal; Magner, Edmond

    2013-04-01

    Nanoporous and planar gold electrodes were utilised as supports for the redox enzymes Aspergillus niger glucose oxidase (GOx) and Corynascus thermophilus cellobiose dehydrogenase (CtCDH). Electrodes modified with hydrogels containing enzyme, Os-redox polymers and the cross-linking agent poly(ethylene glycol)diglycidyl ether were used as biosensors for the determination of glucose and lactose. Limits of detection of 6.0 (±0.4), 16.0 (±0.1) and 2.0 (±0.1) μM were obtained for CtCDH-modified lactose and glucose biosensors and GOx-modified glucose biosensors, respectively, at nanoporous gold electrodes. Biofuel cells composed of GOx- and CtCDH-modified gold electrodes were utilised as anodes, together with Myrothecium verrucaria bilirubin oxidase (MvBOD) or Melanocarpus albomyces laccase as cathodes, in biofuel cells. A maximum power density of 41 μW/cm(2) was obtained for a CtCDH/MvBOD biofuel cell in 5 mM lactose and O2-saturated buffer (pH 7.4, 0.1 M phosphate, 150 mM NaCl). PMID:23274559

  20. Starchy biomass-powered enzymatic biofuel cell based on amylases and glucose oxidase multi-immobilized bioanode.

    PubMed

    Yamamoto, Kazuhiro; Matsumoto, Takuya; Shimada, Shota; Tanaka, Tsutomu; Kondo, Akihiko

    2013-06-25

    The present study reports the design of a novel bioanode to directly utilize starch as a fuel in an enzymatic biofuel cell. The enzymatic fuel cell is based on three enzymes (alpha-amylase, glucoamylase and glucose oxidase). The carbon paste electrode containing these three enzymes and tetrathiafulvalene can both saccharize and oxidize starchy biomass. In cyclic voltammetry, catalytic currents were successfully observed with both glucose and starchy white rice used as a substrate. Finally, a membrane-less white rice/O2 biofuel cell was assembled and the electrochemical performance was evaluated. The three enzyme based electrode was used as a bioanode and an immobilized bilirubin oxidase (derived from Myrothecium verrucaria) electrode was used as a biocathode. The biofuel cell delivered an open circuit voltage of 0.522V and power density of up to 99.0 μWcm(-2). Our results show that a readily available fuel can be used for enzymatic fuel cells, and will lead to new designs. PMID:23624306

  1. Bounding the Bogoliubov coefficients

    SciTech Connect

    Boonserm, Petarpa; Visser, Matt

    2008-11-15

    While over the last century or more considerable effort has been put into the problem of finding approximate solutions for wave equations in general, and quantum mechanical problems in particular, it appears that as yet relatively little work seems to have been put into the complementary problem of establishing rigourous bounds on the exact solutions. We have in mind either bounds on parametric amplification and the related quantum phenomenon of particle production (as encoded in the Bogoliubov coefficients), or bounds on transmission and reflection coefficients. Modifying and streamlining an approach developed by one of the present authors [M. Visser, Phys. Rev. A 59 (1999) 427-438, (arXiv:quant-ph/9901030)], we investigate this question by developing a formal but exact solution for the appropriate second-order linear ODE in terms of a time-ordered exponential of 2x2 matrices, then relating the Bogoliubov coefficients to certain invariants of this matrix. By bounding the matrix in an appropriate manner, we can thereby bound the Bogoliubov coefficients.

  2. NADPH oxidases: new actors in thyroid cancer?

    PubMed

    Ameziane-El-Hassani, Rabii; Schlumberger, Martin; Dupuy, Corinne

    2016-08-01

    Hydrogen peroxide (H2O2) is a crucial substrate for thyroid peroxidase, a key enzyme involved in thyroid hormone synthesis. However, as a potent oxidant, H2O2 might also be responsible for the high level of oxidative DNA damage observed in thyroid tissues, such as DNA base lesions and strand breakages, which promote chromosomal instability and contribute to the development of tumours. Although the role of H2O2 in thyroid hormone synthesis is well established, its precise mechanisms of action in pathological processes are still under investigation. The NADPH oxidase/dual oxidase family are the only oxidoreductases whose primary function is to produce reactive oxygen species. As such, the function and expression of these enzymes are tightly regulated. Thyrocytes express dual oxidase 2, which produces most of the H2O2 for thyroid hormone synthesis. Thyrocytes also express dual oxidase 1 and NADPH oxidase 4, but the roles of these enzymes are still unknown. Here, we review the structure, expression, localization and function of these enzymes. We focus on their potential role in thyroid cancer, which is characterized by increased expression of these enzymes. PMID:27174022

  3. Flavin-dependent alcohol oxidase from the yeast Pichia pinus. Spatial localization of the coenzyme FAD in the protein structure: hot-tritium bombardment and ESR experiments.

    PubMed Central

    Averbakh, A Z; Pekel, N D; Seredenko, V I; Kulikov, A V; Gvozdev, R I; Rudakova, I P

    1995-01-01

    The spatial localization of the coenzyme FAD in the quaternary structure of the alcohol oxidase from the yeast Pichia pinus was studied by tritium planigraphy and ESR methods. In the present paper we measured the specific radioactivity of FAD labelled as a part of the alcohol oxidase complex. The specific-radioactivity ratio for two FAD portions (FMN and AMP) was calculated. ESR experiments show 4 A (0.4 nm) to be the depth of immersion of paramagnetic isoalloxazines into alcohol oxidase octamer molecules. It is suggested that FAD molecules are bound to the surface of the octamer, rather than to the subunit interfaces. The orientation of the prosthetic group FAD in the alcohol oxidase protein is discussed. PMID:7654201

  4. Validation of EMP bounds

    SciTech Connect

    Warne, L.K.; Merewether, K.O.; Chen, K.C.; Jorgenson, R.E.; Morris, M.E.; Solberg, J.E.; Lewis, J.G.; Derr, W.

    1996-07-01

    Test data on canonical weapon-like fixtures are used to validate previously developed analytical bounding results. The test fixtures were constructed to simulate (but be slightly worse than) weapon ports of entry but have known geometries (and electrical points of contact). The exterior of the test fixtures exhibited exterior resonant enhancement of the incident fields at the ports of entry with magnitudes equal to those of weapon geometries. The interior consisted of loaded transmission lines adjusted to maximize received energy or voltage but incorporating practical weapon geometrical constraints. New analytical results are also presented for bounding the energies associated with multiple bolt joints and for bounding the exterior resonant enhancement of the exciting fields.

  5. Body Fat Percentage Is a Major Determinant of Total Bilirubin Independently of UGT1A1*28 Polymorphism in Young Obese

    PubMed Central

    Kohlova, Michaela; Bronze-da-Rocha, Elsa; Fernandes, João; Costa, Elísio; Catarino, Cristina; Aires, Luísa; Mansilha, Helena Ferreira; Rocha-Pereira, Petronila; Quintanilha, Alexandre; Rêgo, Carla; Santos-Silva, Alice

    2014-01-01

    Objectives Bilirubin has potential antioxidant and anti-inflammatory properties. The UGT1A1*28 polymorphism (TA repeats in the promoter region) is a major determinant of bilirubin levels and recent evidence suggests that raised adiposity may also be a contributing factor. We aimed to study the interaction between UGT1A1 polymorphism, hematological and anthropometric variables with total bilirubin levels in young individuals. Methods 350 obese (mean age of 11.6 years; 52% females) and 79 controls (mean age of 10.5 years; 59% females) were included. Total bilirubin and C-reactive protein (CRP) plasma levels, hemogram, anthropometric data and UGT1A1 polymorphism were determined. In a subgroup of 74 obese and 40 controls body composition was analyzed by dual-energy X-ray absorptiometry. Results The UGT1A1 genotype frequencies were 49.9%, 42.7% and 7.5% for 6/6, 6/7 and 7/7 genotypes, respectively. Patients with 7/7 genotype presented the highest total bilirubin levels, followed by 6/7 and 6/6 genotypes. Compared to controls, obese patients presented higher erythrocyte count, hematocrit, hemoglobin and CRP levels, but no differences in bilirubin or in UGT1A1 genotype distribution. Body fat percentage was inversely correlated with bilirubin in obese patients but not in controls. This inverse association was observed either in 6/7 or 6/6 genotype obese patients. UGT1A1 polymorphism and body fat percentage were the main factors affecting bilirubin levels within obese patients (linear regression analysis). Conclusion In obese children and adolescents, body fat composition and UGT1A1 polymorphism are independent determinants of total bilirubin levels. Obese individuals with 6/6 UGT1A1 genotype and higher body fat mass may benefit from a closer clinical follow-up. PMID:24901842

  6. Distortion of Flavin Geometry Is Linked to Ligand Binding in Cholesterol Oxidase

    SciTech Connect

    Lyubimov, A.Y.; Heard, K.; Tang, H.; Sampson, N.S.; Vrielink, A.

    2009-06-03

    Two high-resolution structures of a double mutant of bacterial cholesterol oxidase in the presence or absence of a ligand, glycerol, are presented, showing the trajectory of glycerol as it binds in a Michaelis complex-like position in the active site. A group of three aromatic residues forces the oxidized isoalloxazine moiety to bend along the N5-N10 axis as a response to the binding of glycerol in the active site. Movement of these aromatic residues is only observed in the glycerol-bound structure, indicating that some tuning of the FAD redox potential is caused by the formation of the Michaelis complex during regular catalysis. This structural study suggests a possible mechanism of substrate-assisted flavin activation, improves our understanding of the interplay between the enzyme, its flavin cofactor and its substrate, and is of use to the future design of effective cholesterol oxidase inhibitors.

  7. CYTOCHROME OXIDASE IN NORMAL AND REGENERATING NEURONS

    PubMed Central

    Howe, Howard A.; Mellors, Robert C.

    1945-01-01

    Manometric determinations of cytochrome oxidase activity were carried out on grey matter from the thalamus and anterior horn of cats and monkeys under various experimental conditions. The thalamus of the cat was studied following the degeneration of virtually all the thalamic neurons secondary to decortication. In comparing the deneuronated thalamus with the normal one, it was found that approximately 34 per cent of the cytochrome oxidase activity was contributed by the neurons and the balance by neuroglia and mesodermal tissues which on the operated side remained comparable to that of the normal side. Total activity of the normal thalamus averaged 5.52 units per mg. of dry weight where I unit is defined as the amount of cytochrome oxidase required to produce a net oxygen consumption of 10 c.mm. per hour under the specified conditions of the experiment. The grey matter of the anterior horns of the spinal cord was isolated by a special technique and its cytochrome oxidase activity was compared with anterior horns in which motoneurons had been stimulated to regenerative activity by section of peripheral nerves. Each animal was studied in relation to an anterior horn which was normal and one in which only the functional state of the motoneurons had been changed. Average normal levels of 2.23 units were found for cat anterior horn and 0.69 units for the monkey. Reductions of cytochrome oxidase activity in the range of 22 to 23 per cent were observed for both cat and monkey following nerve section. In the latter the time sequence was carefully studied in relation to the cytological cycle known as chromatolysis and a virus refractory state previously described by us. It was found that maximal reduction of cytochrome oxidase activity coincided with maximal refractoriness of the cells to poliomyelitis virus (30 to 70 days following nerve section). Neither of these states could be correlated in time with maximal chromatolysis (10 to 15 days). PMID:19871471

  8. Research on prevention of bilirubin-induced brain injury and kernicterus: National Institute of Child Health and Human Development conference executive summary. 2003.

    PubMed

    Blackmon, Lillian R; Fanaroff, Avroy A; Raju, Tonse N K

    2004-07-01

    In July 2003, the National Institute of Child Health and Human Development convened a conference, "Research on Prevention of Bilirubin-Induced Brain Injury and Kernicterus: Bench-to-Bedside." This article will provide a summary of presentations and discussions from this conference. The summary will focus on the identified knowledge gaps in 5 areas related to bilirubin-induced brain injury and kernicterus: 1) neurobiology and neuroimaging; 2) epidemiology and issues of clinical management; 3) methodologies for assessing clinical jaundice and direct and noninvasive measurement of serum bilirubin and hemolysis; 4) therapies for management of neonatal hyperbilirubinemia; and 5) public health surveillance and systems-based approaches to prevention. PMID:15231933

  9. Computing Graphical Confidence Bounds

    NASA Technical Reports Server (NTRS)

    Mezzacappa, M. A.

    1983-01-01

    Approximation for graphical confidence bounds is simple enough to run on programmable calculator. Approximation is used in lieu of numerical tables not always available, and exact calculations, which often require rather sizable computer resources. Approximation verified for collection of up to 50 data points. Method used to analyze tile-strength data on Space Shuttle thermal-protection system.

  10. Isolation and characterization of ubiquinol oxidase complexes from Paracoccus denitrificans cells cultured under various limiting growth conditions in the chemostat.

    PubMed

    Bosma, G; Braster, M; Stouthamer, A H; van Verseveld, H W

    1987-06-15

    To obtain more information about the composition of the respiratory chain under different growth conditions and about the regulation of electron-transfer to several oxidases and reductases, ubiquinol oxidase complexes were partially purified from membranes of Paracoccus denitrificans cells grown in carbon-source-limited aerobic, nitrate-limited anaerobic and oxygen-limited chemostat cultures. The isolated enzymes consisted of cytochromes bc1, c552 and aa3. In comparison with the aerobic ubiquinol oxidase complex, the oxygen- and nitrate-limited ones contained, respectively, less and far less of the cytochrome aa3 subunits and the anaerobic complex also contained lower amounts of cytochrome c552. In addition, extra haem-containing polypeptides were present with apparent Mr of 14,000, 30,000 and 45,000, the former one only in the anaerobic and the latter two in both the anaerobic and oxygen-limited preparations. This is the first report describing four different membrane-bound c-type cytochromes. The potentiometric and spectral characteristics of the redox components in membrane particles and isolated ubiquinol oxidase fractions were determined by combined potentiometric analysis and spectrum deconvolution. Membranes of nitrate- and oxygen-limited cells contained extra high-potential cytochrome b in comparison with the membranes of aerobically grown cells. No difference was detected between the three isolated ubiquinol oxidase complexes. Aberrances with already published values of redox potentials are discussed. PMID:3036512

  11. Mutation of a single residue in the ba3 oxidase specifically impairs protonation of the pump site

    PubMed Central

    von Ballmoos, Christoph; Gonska, Nathalie; Lachmann, Peter; Gennis, Robert B.; Ädelroth, Pia; Brzezinski, Peter

    2015-01-01

    The ba3-type cytochrome c oxidase from Thermus thermophilus is a membrane-bound protein complex that couples electron transfer to O2 to proton translocation across the membrane. To elucidate the mechanism of the redox-driven proton pumping, we investigated the kinetics of electron and proton transfer in a structural variant of the ba3 oxidase where a putative “pump site” was modified by replacement of Asp372 by Ile. In this structural variant, proton pumping was uncoupled from internal electron transfer and O2 reduction. The results from our studies show that proton uptake to the pump site (time constant ∼65 μs in the wild-type cytochrome c oxidase) was impaired in the Asp372Ile variant. Furthermore, a reaction step that in the wild-type cytochrome c oxidase is linked to simultaneous proton uptake and release with a time constant of ∼1.2 ms was slowed to ∼8.4 ms, and in Asp372Ile was only associated with proton uptake to the catalytic site. These data identify reaction steps that are associated with protonation and deprotonation of the pump site, and point to the area around Asp372 as the location of this site in the ba3 cytochrome c oxidase. PMID:25733886

  12. Analysis of binding ability of two tetramethylpyridylporphyrins to albumin and its complex with bilirubin.

    PubMed

    Solomonov, Alexey V; Shipitsyna, Maria K; Vashurin, Arthur S; Rumyantsev, Evgeniy V; Timin, Alexander S; Ivanov, Sergey P

    2016-11-01

    An interaction between 5,10,15,20-tetrakis-(N-methyl-x-pyridyl)porphyrins, x=2; 4 (TMPyPs) with bovine serum albumin (BSA) and its bilirubin (BR) complex was investigated by UV-Viz and fluorescence spectroscopy under imitated physiological conditions involving molecular docking studies. The parameters of forming intermolecular complexes (binding constants, quenching rate constants, quenching sphere radius etc.) were determined. It was showed that the interaction between proteins and TMPyPs occurs via static quenching of protein fluorescence and has predominantly hydrophobic and electrostatic character. It was revealed that obtained complexes are relatively stable, but in the case of TMPyP4 binding with proteins occurs better than TMPyP2. Nevertheless, both TMPyPs have better binding ability with free protein compared to BRBSA at the same time. The influence of TMPyPs on the conformational changes in protein molecules was studied using synchronous fluorescence spectroscopy. It was found that there is no competition of BR with TMPyPs for binging sites on protein molecule and BR displacement does not occur. Molecular docking calculations have showed that TMPyPs can bind with albumin via tryptophan residue in the hydrophilic binding site of protein molecule but it is not one possible interaction way. PMID:27267279

  13. Characterization of erythrosine B binding to bovine serum albumin and bilirubin displacement.

    PubMed

    Mathavan, Vinodaran M K; Boh, Boon Kim; Tayyab, Saad

    2009-08-01

    The interaction of crythrosine B (ErB), a commonly used dye for coloring foods and drinks, with bovine scrum albumin (BSA) was investigated both in the absence and presence of bilirubin (BR) using absorption and absorption difference spectroscopy. ErB binding to BSA was reflected from a significant red shift of 11 nm in the absorption maximum of ErB (527 nm) with the change in absorbance at lamdamax. Analysis of absorption difference spectroscopic titration results of BSA with increasing concentrations of ErB3 using Benesi-Hildebrand equation gave the association constant, K as 6.9 x 10(4) M(-1). BR displacing action of ErB was revealed by a significant blue shift in the absorption maximum, accompanied by a decrease in absorbance difference at lamdamax in the difference spectrum of BR-BSA complex upon addition of increasing concentrations of ErB. This was further substantiated by fluorescence spectroscopy, as addition of increasing concentrations of ErB to BR-BSA complex caused a significant decrease in fluoresccnce at 510 nm. The results suggest that ErB binds to a site in the vicinity of BR binding site on BSA. Therefore, intake of ErB may increase the risk of hyperbilirubinemia in the healthy subjects. PMID:19788065

  14. Rethinking the immune properties of bilirubin in viral hepatitis: from bench to bedside

    PubMed Central

    Corral-Jara, Karla F; Trujillo-Ochoa, Jorge L; Realpe, Mauricio; Panduro, Arturo; Roman, Sonia; Fierro, Nora A

    2015-01-01

    Communication between the immune system and metabolic components can be exemplified by the process of heme catabolism. The immunomodulatory functions of the enzymes, substrates and active products related to catabolism of the heme group have been extensively studied. Bilirubin (BR), the final breakdown product of heme, is primarily considered to be a toxic waste product but has recently been considered to be an immunomodulatory metabolite. Through mechanisms that include intracellular signaling and transcriptional control, BR affects those immune cell functions that regulate cell proliferation, differentiation and apoptosis. During the pathogenesis of viral hepatitis, the heme degradation pathway is disrupted, resulting in changes to normal BR concentrations. These alterations have been previously studied mainly as a consequence of the infection. However, little is known about the potential immunomodulatory role played by BR in the development of infectious hepatocellular diseases. Differences in BR levels in the context of viral hepatitis are likely to provide important insights into the metabolite-mediated mechanisms controlling the immune responses underlying both the long-term persistence of hepatitis C virus (HCV) infection and the resolution of hepatitis A virus (HAV) infection during the acute phase. In this review, the cross-talk between heme catabolism and immune function is described in detail. Special emphasis is given to discoveries that hold promise for identifying immunologic features of metabolic products in the resolution of viral diseases. PMID:26719800

  15. Development of a diagnostic system for bilirubin detection in cerebral spinal fluid

    NASA Astrophysics Data System (ADS)

    Bhadri, Prashant R.; Salgaonkar, Vasant A.; Majumdar, Anindya; Morgan, Chad J.; Zuccarello, Mario; Pyne, Gail J.; Dulaney, Elizabeth; Caffery, James, Jr.; Shukla, Rakesh; Beyette, Fred R., Jr.

    2004-11-01

    A weakened portion of an artery in the brain leads to a medical condition known as a cerebral aneurysm. A subarachnoid hemorrhage (SAH) occurs when an aneurysm ruptures. For those individuals suspected of having a SAH, a computerized tomography (CT) scan of the brain usually demonstrates evidence of the bleeding. However, in a considerable portion of people, the CT scan is unable to detect the blood that has escaped from the blood vessel. Recent studies have indicated nearly 30% of patients with a SAH are initially misdiagnosed. For circumstances when a SAH is suspected despite a normal CT scan, physicians make the diagnosis of SAH by performing a spinal tap. A spinal tap uses a needle to sample the cerebrospinal fluid (CSF) collected from the patient"s lumbar spine. However, it is also possible for blood to be introduced into the CSF as a result of the spinal tap procedure. Therefore, an effective solution is required to help medical personnel differentiate between the blood that results from a tap and that from a ruptured aneurysm. In this paper, the development of a prototype is described which is sensitive and specific for measuring bilirubin in CSF, hemorrhagic-CSF and CSF-like solutions. To develop this instrument a combination of spectrophotometric analysis, custom data analysis software and other hardware interfaces are assembled that lay the foundation for the development of portable and user-friendly equipment suitable for assisting trained medical personnel with the diagnosis of a ruptured cerebral aneurysm.

  16. The First Mammalian Aldehyde Oxidase Crystal Structure

    PubMed Central

    Coelho, Catarina; Mahro, Martin; Trincão, José; Carvalho, Alexandra T. P.; Ramos, Maria João; Terao, Mineko; Garattini, Enrico; Leimkühler, Silke; Romão, Maria João

    2012-01-01

    Aldehyde oxidases (AOXs) are homodimeric proteins belonging to the xanthine oxidase family of molybdenum-containing enzymes. Each 150-kDa monomer contains a FAD redox cofactor, two spectroscopically distinct [2Fe-2S] clusters, and a molybdenum cofactor located within the protein active site. AOXs are characterized by broad range substrate specificity, oxidizing different aldehydes and aromatic N-heterocycles. Despite increasing recognition of its role in the metabolism of drugs and xenobiotics, the physiological function of the protein is still largely unknown. We have crystallized and solved the crystal structure of mouse liver aldehyde oxidase 3 to 2.9 Å. This is the first mammalian AOX whose structure has been solved. The structure provides important insights into the protein active center and further evidence on the catalytic differences characterizing AOX and xanthine oxidoreductase. The mouse liver aldehyde oxidase 3 three-dimensional structure combined with kinetic, mutagenesis data, molecular docking, and molecular dynamics studies make a decisive contribution to understand the molecular basis of its rather broad substrate specificity. PMID:23019336

  17. Oxidative stress, NADPH oxidases, and arteries.

    PubMed

    Sun, Qi-An; Runge, Marschall S; Madamanchi, Nageswara R

    2016-05-10

    Atherosclerosis and its major complications - myocardial infarction and stroke - remain major causes of death and disability in the United States and world-wide. Indeed, with dramatic increases in obesity and diabetes mellitus, the prevalence and public health impact of cardiovascular diseases (CVD) will likely remain high. Major advances have been made in development of new therapies to reduce the incidence of atherosclerosis and CVD, in particular for treatment of hypercholesterolemia and hypertension. Oxidative stress is the common mechanistic link for many CVD risk factors. However, only recently have the tools existed to study the interface between oxidative stress and CVD in animal models. The most important source of reactive oxygen species (and hence oxidative stress) in vascular cells are the multiple forms of enzymes nicotinamide adenine dinucleotide phosphate oxidase (NADPH oxidase). Recently published and emerging studies now clearly establish that: 1) NADPH oxidases are of critical importance in atherosclerosis and hypertension in animal models; 2) given the tissue-specific expression of key components of NADPH oxidase, it may be possible to target vascular oxidative stress for prevention of CVD. PMID:25649240

  18. The substrate tolerance of alcohol oxidases.

    PubMed

    Pickl, Mathias; Fuchs, Michael; Glueck, Silvia M; Faber, Kurt

    2015-08-01

    Alcohols are a rich source of compounds from renewable sources, but they have to be activated in order to allow the modification of their carbon backbone. The latter can be achieved via oxidation to the corresponding aldehydes or ketones. As an alternative to (thermodynamically disfavoured) nicotinamide-dependent alcohol dehydrogenases, alcohol oxidases make use of molecular oxygen but their application is under-represented in synthetic biotransformations. In this review, the mechanism of copper-containing and flavoprotein alcohol oxidases is discussed in view of their ability to accept electronically activated or non-activated alcohols and their propensity towards over-oxidation of aldehydes yielding carboxylic acids. In order to facilitate the selection of the optimal enzyme for a given biocatalytic application, the substrate tolerance of alcohol oxidases is compiled and discussed: Substrates are classified into groups (non-activated prim- and sec-alcohols; activated allylic, cinnamic and benzylic alcohols; hydroxy acids; sugar alcohols; nucleotide alcohols; sterols) together with suitable alcohol oxidases, their microbial source, relative activities and (stereo)selectivities. PMID:26153139

  19. Research Advances. Image Pinpoints All 5 Million Atoms in Viral Coat; Bilirubin, "Animals-Only" Pigment, Found in Plants; New Evidence Shows Humans Make Salicylic Acid

    NASA Astrophysics Data System (ADS)

    King, Angela G.

    2009-08-01

    Recent "firsts" in chemical research: image of a viral capsid pinpointing 5 million atoms; isolation and identification of an "animal" pigment, bilirubin, from a plant source; evidence that humans make salicylic acid.

  20. Inhibitory Effects of Palm Tocotrienol-Rich Fraction Supplementation on Bilirubin-Metabolizing Enzymes in Hyperbilirubinemic Adult Rats

    PubMed Central

    Kamisah, Yusof; Lim, Jing Jye; Lim, Chew-Lian; Asmadi, Ahmad Y.

    2014-01-01

    Background Phenylhydrazine, a hemolytic agent, is widely used as a model of experimental hyperbilirubinemia. Palm tocotrienol-rich fraction (TRF) was shown to exert beneficial effects in hyperbilirubinemic rat neonates. Aim To investigate the effects of palm TRF supplementation on hepatic bilirubin-metabolizing enzymes and ocidative stress status in rats administered phenylhydrazine. Methods Twenty-four male Wistar rats were divided into two groups; one group was intraperitoneally injected with palm TRF at the dose of 30 mg/kg/day, while another group was only given vehicle (control) (vitamin E-free palm oil) for 14 days. Twenty-four hours after the last dose, each group was further subdivided into another two groups. One group was administered phenylhydrazine (100 mg/kg, intraperitoneally) and another group was administered normal saline. Twenty-four hours later, blood and liver were collected for biochemical parameter measurements. Results Phenylhydrazine increased plasma total bilirubin level and oxidative stress in the erythrocytes as well as in the liver, which were reduced by the pretreatment of palm TRF. Palm TRF also prevented the increases in hepatic heme oxygenase, biliverdin reductase and UDP-glucuronyltransferase activities induced by phenylhydrazine. Conclusion Palm tocotrienol-rich fraction was able to afford protection against phenylhydrazine-induced hyperbilirubinemia, possibly by reducing oxidative stress and inhibiting bilirubin-metabolizing enzymes in the liver. PMID:24586630

  1. Bilirubin removal from human plasma by Cibacron Blue F3GA using immobilized microporous affinity membranous capillary method.

    PubMed

    Zhang, Lei; Jin, Gu

    2005-07-01

    A novel affinity sorbent system for direct bilirubin removal from human plasma was developed. These new adsorbents comprise Cibacron Blue F3GA as the specific ligand, and microporous membranous poly(tetrafluoroethylene) capillary (modified by coating with a hydrophilic layer of poly(vinyl alcohol) after activation) as the carrier matrix. The affinity adsorbents carrying 126.5 micromol Cibacron Blue F3GA/g polymer was then used to remove bilirubin in a flow-injection system. Non-specific adsorption on the poly(vinyl alcohol) coated capillary remains low, and higher affinity adsorption capacity, of up to 76.2 mg/g polymer was obtained after dye immobilization. The bilirubin adsorption capacity of the affinity capillary decreased with increase in the recirculation rate of plasma. The adsorption capacity increased with increase the temperature while decreased with increase the ionic strength. The maximum adsorption was only observed in neutral solution (pH 6-7). The adsorption isotherm fitted the Langmuir model well. These new adsorbents have higher velocity of mass transfer, better adsorption capacity, less fouling, longer service life and good reusability. The results of blood tests suggested the dye affinity capillary has good blood compatibility. PMID:15894520

  2. Dimethyl ester of bilirubin exhibits anti-inflammatory activity through inhibition of secretory phospholipase A2, lipoxygenase and cyclooxygenase.

    PubMed

    Joshi, Vikram; Umashankara, M; Ramakrishnan, Chandrasekaran; Nanjaraj Urs, Ankanahalli N; Suvilesh, Kanve Nagaraj; Velmurugan, Devadasan; Rangappa, Kanchugarakoppal S; Vishwanath, Bannikuppe Sannanaik

    2016-05-15

    Overproduction of arachidonic acid (AA) mediated by secretory phospholipase A2 group IIA (sPLA2IIA) is a hallmark of many inflammatory disorders. AA is subsequently converted into pro-inflammatory eicosanoids through 5-lipoxygenase (5-LOX) and cyclooxygenase-1/2 (COX-1/2) activities. Hence, inhibition of sPLA2IIA, 5-LOX and COX-1/2 activities is critical in regulating inflammation. We have previously reported unconjugated bilirubin (UCB), an endogenous antioxidant, as sPLA2IIA inhibitor. However, lipophilic UCB gets conjugated in liver with glucuronic acid into hydrophilic conjugated bilirubin (CB). Since hydrophobicity is pre-requisite for sPLA2IIA inhibition, conjugation reduces the efficacy of UCB. In this regard, UCB was chemically modified and derivatives were evaluated for sPLA2IIA, 5-LOX and COX-1/2 inhibition. Among the derivatives, BD1 (dimethyl ester of bilirubin) exhibited ∼ 3 fold greater inhibitory potency towards sPLA2IIA compared to UCB. Both UCB and BD1 inhibited human 5-LOX and COX-2 activities; however only BD1 inhibited AA induced platelet aggregation. Molecular docking studies demonstrated BD1 as better inhibitor of aforesaid enzymes than UCB and other endogenous antioxidants. These data suggest that BD1 exhibits strong anti-inflammatory activity through inhibition of AA cascade enzymes which is of great therapeutic importance. PMID:27060751

  3. Electron transfer reactions in the NADPH oxidase system of neutrophils--involvement of an NADPH-cytochrome c reductase in the oxidase system.

    PubMed

    Fujii, H; Kakinuma, K

    1991-11-12

    Membrane-bound NADPH oxidase of pig blood neutrophils was solubilized with heptylthioglucoside in a high yield. The solubilized preparation from myristate-stimulated cells (sample S) showed high O2- generating activity, and the preparation from resting cells (sample R) had no activity, but the two samples had equal amounts of flavins and cytochrome b-558 (cyt b-558). The electron transfer reactions to exogenous cytochrome c (cyt c) or cyt b-558 in samples S and R were examined. Under anaerobic conditions, NADPH-dependent cyt c reductase activity appeared higher in sample S than in sample R, and the addition of FMN and FAD greatly enhanced the reductase activity of sample S, but not that of sample R. No marked difference between the reductase activities of samples S and R was seen with NADH. Photoreduction of the NADPH oxidase system was examined in the absence of NADPH under anaerobic conditions by monitoring the reduction rates of exogenous cyt c using a flashlight with cut-off filters between 400 and 500 nm. Cyt c reduction was much higher in sample S than in sample R on photoexcitation at about 450 nm. Photoreduction was carried out with a band-pass filter for selective irradiation at 450 nm. Marked reduction of exogenous cyt c was observed only in sample S: the small reduction of cyt c by sample R was independent of the light wavelength and was equal to the blank level. In contrast, no difference in the reduction of cyt b-558 by the two samples was found by either NADPH or photoreduction. Under aerobic conditions, no direct reduction of either cyt c or cyt b-558 was observed. These results suggest that an NADPH-cyt c reductase (a membrane-bound flavoprotein) is involved in the NADPH oxidase system of stimulated neutrophils. PMID:1659905

  4. Petawatt laser absorption bounded

    PubMed Central

    Levy, Matthew C.; Wilks, Scott C.; Tabak, Max; Libby, Stephen B.; Baring, Matthew G.

    2014-01-01

    The interaction of petawatt (1015 W) lasers with solid matter forms the basis for advanced scientific applications such as table-top particle accelerators, ultrafast imaging systems and laser fusion. Key metrics for these applications relate to absorption, yet conditions in this regime are so nonlinear that it is often impossible to know the fraction of absorbed light f, and even the range of f is unknown. Here using a relativistic Rankine-Hugoniot-like analysis, we show for the first time that f exhibits a theoretical maximum and minimum. These bounds constrain nonlinear absorption mechanisms across the petawatt regime, forbidding high absorption values at low laser power and low absorption values at high laser power. For applications needing to circumvent the absorption bounds, these results will accelerate a shift from solid targets, towards structured and multilayer targets, and lead the development of new materials. PMID:24938656

  5. Petawatt laser absorption bounded

    NASA Astrophysics Data System (ADS)

    Levy, Matthew C.; Wilks, Scott C.; Tabak, Max; Libby, Stephen B.; Baring, Matthew G.

    2014-06-01

    The interaction of petawatt (1015 W) lasers with solid matter forms the basis for advanced scientific applications such as table-top particle accelerators, ultrafast imaging systems and laser fusion. Key metrics for these applications relate to absorption, yet conditions in this regime are so nonlinear that it is often impossible to know the fraction of absorbed light f, and even the range of f is unknown. Here using a relativistic Rankine-Hugoniot-like analysis, we show for the first time that f exhibits a theoretical maximum and minimum. These bounds constrain nonlinear absorption mechanisms across the petawatt regime, forbidding high absorption values at low laser power and low absorption values at high laser power. For applications needing to circumvent the absorption bounds, these results will accelerate a shift from solid targets, towards structured and multilayer targets, and lead the development of new materials.

  6. Structure-function characterization reveals new catalytic diversity in the galactose oxidase and glyoxal oxidase family.

    PubMed

    Yin, DeLu Tyler; Urresti, Saioa; Lafond, Mickael; Johnston, Esther M; Derikvand, Fatemeh; Ciano, Luisa; Berrin, Jean-Guy; Henrissat, Bernard; Walton, Paul H; Davies, Gideon J; Brumer, Harry

    2015-01-01

    Alcohol oxidases, including carbohydrate oxidases, have a long history of research that has generated fundamental biological understanding and biotechnological applications. Despite a long history of study, the galactose 6-oxidase/glyoxal oxidase family of mononuclear copper-radical oxidases, Auxiliary Activity Family 5 (AA5), is currently represented by only very few characterized members. Here we report the recombinant production and detailed structure-function analyses of two homologues from the phytopathogenic fungi Colletotrichum graminicola and C. gloeosporioides, CgrAlcOx and CglAlcOx, respectively, to explore the wider biocatalytic potential in AA5. EPR spectroscopy and crystallographic analysis confirm a common active-site structure vis-à-vis the archetypal galactose 6-oxidase from Fusarium graminearum. Strikingly, however, CgrAlcOx and CglAlcOx are essentially incapable of oxidizing galactose and galactosides, but instead efficiently catalyse the oxidation of diverse aliphatic alcohols. The results highlight the significant potential of prospecting the evolutionary diversity of AA5 to reveal novel enzyme specificities, thereby informing both biology and applications. PMID:26680532

  7. Structure–function characterization reveals new catalytic diversity in the galactose oxidase and glyoxal oxidase family

    PubMed Central

    Yin, DeLu (Tyler); Urresti, Saioa; Lafond, Mickael; Johnston, Esther M.; Derikvand, Fatemeh; Ciano, Luisa; Berrin, Jean-Guy; Henrissat, Bernard; Walton, Paul H.; Davies, Gideon J.; Brumer, Harry

    2015-01-01

    Alcohol oxidases, including carbohydrate oxidases, have a long history of research that has generated fundamental biological understanding and biotechnological applications. Despite a long history of study, the galactose 6-oxidase/glyoxal oxidase family of mononuclear copper-radical oxidases, Auxiliary Activity Family 5 (AA5), is currently represented by only very few characterized members. Here we report the recombinant production and detailed structure–function analyses of two homologues from the phytopathogenic fungi Colletotrichum graminicola and C. gloeosporioides, CgrAlcOx and CglAlcOx, respectively, to explore the wider biocatalytic potential in AA5. EPR spectroscopy and crystallographic analysis confirm a common active-site structure vis-à-vis the archetypal galactose 6-oxidase from Fusarium graminearum. Strikingly, however, CgrAlcOx and CglAlcOx are essentially incapable of oxidizing galactose and galactosides, but instead efficiently catalyse the oxidation of diverse aliphatic alcohols. The results highlight the significant potential of prospecting the evolutionary diversity of AA5 to reveal novel enzyme specificities, thereby informing both biology and applications. PMID:26680532

  8. Application of high-performance liquid chromatography combined with ultra-sensitive thermal lens spectrometric detection for simultaneous biliverdin and bilirubin assessment at trace levels in human serum.

    PubMed

    Martelanc, Mitja; Žiberna, Lovro; Passamonti, Sabina; Franko, Mladen

    2016-07-01

    We present the applicability of a new ultra-sensitive analytical method for the simultaneous determination of biliverdin and bilirubin in human serum. The method comprises isocratic reversed-phase (RP) C18 high-performance liquid chromatography (HPLC) and thermal lens spectrometric detection (TLS) based on excitation by a krypton laser emission line at 407nm. This method enables the separation of IX-α biliverdin and IX-α bilirubin in 11min with limit of detection (LOD) and limit of quantitation (LOQ) for biliverdin of 1.2nM and 3nM, and 1nM and 2.8nM for bilirubin, respectively. In addition, a step-gradient elution was set up, by changing the mobile phase composition, in order to further enhance the sensitivity for bilirubin determination with LOD and LOQ of 0.5nM and 1.5nM, respectively. In parallel, an isocratic HPLC-DAD method was developed for benchmarking against HPLC-TLS methods. The LOD and LOQ for biliverdin were 6nM and 18nM, and 2.5nM and 8nM for bilirubin, respectively. Additionally, both isocratic methods were applied for measuring biliverdin and free bilirubin in human serum samples (from 2 male and 2 female healthy donors). Combining isocratic HPLC method with TLS detector was crucial for first ever biliverdin determination in serum together with simultaneous free bilirubin determination. We showed for the first time the concentration ratio of free bilirubin versus unbound biliverdin in human serum samples. PMID:27154653

  9. Serum Total Bilirubin Levels Provide Additive Risk Information over the Framingham Risk Score for Identifying Asymptomatic Diabetic Patients at Higher Risk for Coronary Artery Stenosis

    PubMed Central

    Leem, Jaechan; Koh, Eun Hee; Jang, Jung Eun; Woo, Chang-Yun; Oh, Jin Sun; Lee, Min Jung; Kang, Joon-Won; Lim, Tae-Hwan; Jung, Chang Hee; Lee, Woo Je; Park, Joong-Yeol

    2015-01-01

    Background The diagnosis of coronary artery disease (CAD) is often delayed in patients with type 2 diabetes. Serum total bilirubin levels are inversely associated with CAD. However, no studies have examined whether this can be used as a biochemical marker for identifying asymptomatic diabetic patients at higher risk for having obstructive CAD. Methods We performed a cross-sectional study of 460 consecutive asymptomatic patients with type 2 diabetes. All patients underwent coronary computed tomographic angiography, and their serum total bilirubin levels were measured. Obstructive CAD was defined as ≥50% diameter stenosis in at least one coronary artery. Results Serum total bilirubin tertiles showed an inverse association with the prevalence of obstructive CAD. In multivariate logistic regression analysis, the odds ratio for the highest versus the lowest tertile of total bilirubin was 0.227 (95% confidence interval [CI], 0.130 to 0.398), and an increment of 1 µmol/L in serum total bilirubin level was associated with a 14.6% decrease in obstructive CAD after adjustment for confounding variables. Receiver operating characteristic curve analysis showed that the area under the curve for the Framingham Risk Score (FRS) plus serum total bilirubin level was 0.712 (95% CI, 0.668 to 0.753), which is significantly greater than that of the FRS alone (P=0.0028). Conclusion Serum total bilirubin level is inversely associated with obstructive CAD and provides additive risk information over the FRS. Serum total bilirubin may be helpful for identifying asymptomatic patients with type 2 diabetes who are at higher risk for obstructive CAD. PMID:26566499

  10. Apoptosis in murine hepatoma hepa 1c1c7 wild-type, C12, and C4 cells mediated by bilirubin.

    PubMed

    Seubert, John M; Darmon, Alison J; El-Kadi, Ayman O S; D'Souza, Sudhir J A; Bend, John R

    2002-08-01

    Elevated serum and tissue bilirubin concentrations that occur in pathological conditions such as cholestasis, jaundice, and other liver diseases are known to stimulate cytotoxic responses. In preliminary studies, we noted that bilirubin seemed to cause apoptosis in murine hepatoma Hepa 1c1c7 wild-type (WT) cells. Consequently, we investigated apoptosis caused by bilirubin in WT, mutant C12 [aryl hydrocarbon receptor (AHR)-deficient], and C4 (AHR nuclear translocator-deficient) Hepa 1c1c7 cells. Three independent measures of apoptosis were used to quantify the effects of exogenous bilirubin (0, 1, 10, 25, 50, or 100 microM). Caspase-3 activity and cytochrome c release from mitochondria increased at 3 h post-treatment, before increased caspase-8 activity at 6 h, and nuclear condensation by 24 h after treatment with bilirubin. No differences in whole-cell lipid peroxidation were observed between the cell types; however, intracellular reactive oxygen species (ROS) production was greater in WT cells than C12 or C4 cells 3 h after bilirubin exposure. Pretreatment of cells for 1 h with 1 or 10 microM alpha-naphthoflavone, an AHR antagonist, before bilirubin exposure resulted in decreased caspase-3 activity at 6 h and nuclear condensation at 24 h in WT cells. These results indicate that bilirubin, a potential AHR ligand, causes apoptosis in murine Hepa 1c1c7 WT cells by a mechanism(s) partially involving the AHR, disruption of membrane integrity, and increased intracellular ROS production. PMID:12130676

  11. Human serum albumin-stabilized gold nanoclusters act as an electron transfer bridge supporting specific electrocatalysis of bilirubin useful for biosensing applications.

    PubMed

    Santhosh, Mallesh; Chinnadayyala, Somasekhar R; Singh, Naveen K; Goswami, Pranab

    2016-10-01

    Human serum albumin (HSA)-stabilized Au18 nanoclusters (AuNCs) were synthesized and chemically immobilized on an Indium tin oxide (ITO) plate. The assembly process was characterized by advanced electrochemical and spectroscopic techniques. The bare ITO electrode generated three irreversible oxidation peaks, whereas the HSA-AuNC-modified electrode produced a pair of redox peaks for bilirubin at a formal potential of 0.27V (vs. Ag/AgCl). However, the native HSA protein immobilized on the ITO electrode failed to produce any redox peak for bilirubin. The results indicate that the AuNCs present in HSA act as electron transfer bridge between bilirubin and the ITO plate. Docking studies of AuNC with HSA revealed that the best docked structure of the nanocluster is located around the vicinity of the bilirubin binding site, with an orientation that allows specific oxidation. When the HSA-AuNC-modified electrode was employed for the detection of bilirubin using chronoamperometry at 0.3V (vs. Ag/AgCl), a steady-state current response against bilirubin in the range of 0.2μM to 7μM, with a sensitivity of 0.34μAμM(-1) and limit of detection of 86.32nM at S/N 3, was obtained. The bioelectrode was successfully applied to measure the bilirubin content in spiked serum samples. The results indicate the feasibility of using HSA-AuNC as a biorecognition element for the detection of serum bilirubin levels using an electrochemical technique. PMID:27126550

  12. Impairment of enzymatic antioxidant defenses is associated with bilirubin-induced neuronal cell death in the cerebellum of Ugt1 KO mice

    PubMed Central

    Bortolussi, G; Codarin, E; Antoniali, G; Vascotto, C; Vodret, S; Arena, S; Cesaratto, L; Scaloni, A; Tell, G; Muro, A F

    2015-01-01

    Severe hyperbilirubinemia is toxic during central nervous system development. Prolonged and uncontrolled high levels of unconjugated bilirubin lead to bilirubin-induced encephalopathy and eventually death by kernicterus. Despite extensive studies, the molecular and cellular mechanisms of bilirubin toxicity are still poorly defined. To fill this gap, we investigated the molecular processes underlying neuronal injury in a mouse model of severe neonatal jaundice, which develops hyperbilirubinemia as a consequence of a null mutation in the Ugt1 gene. These mutant mice show cerebellar abnormalities and hypoplasia, neuronal cell death and die shortly after birth because of bilirubin neurotoxicity. To identify protein changes associated with bilirubin-induced cell death, we performed proteomic analysis of cerebella from Ugt1 mutant and wild-type mice. Proteomic data pointed-out to oxidoreductase activities or antioxidant processes as important intracellular mechanisms altered during bilirubin-induced neurotoxicity. In particular, they revealed that down-representation of DJ-1, superoxide dismutase, peroxiredoxins 2 and 6 was associated with hyperbilirubinemia in the cerebellum of mutant mice. Interestingly, the reduction in protein levels seems to result from post-translational mechanisms because we did not detect significant quantitative differences in the corresponding mRNAs. We also observed an increase in neuro-specific enolase 2 both in the cerebellum and in the serum of mutant mice, supporting its potential use as a biomarker of bilirubin-induced neurological damage. In conclusion, our data show that different protective mechanisms fail to contrast oxidative burst in bilirubin-affected brain regions, ultimately leading to neurodegeneration. PMID:25950469

  13. Diazonium Functionalisation of Carbon Nanotubes for Specific Orientation of Multicopper Oxidases: Controlling Electron Entry Points and Oxygen Diffusion to the Enzyme.

    PubMed

    Lalaoui, Noémie; Holzinger, Michael; Le Goff, Alan; Cosnier, Serge

    2016-07-18

    We report the controlled orientation of bilirubin oxidases (BOD) from Myrothecium verrucaria on multiwalled carbon nanotubes (MWCNTs) functionalised by electrografting of 6-carboxynaphthalenediazonium and 4-(2-aminoethyl)benzenediazonium salts. On negatively charged naphthoate-modified MWCNTs, a high-potential (0.44 V vs. SCE) oxygen reduction electrocatalysis is observed, occurring via the T1 copper centre. On positively charged ammonium-modified MWCNTs, a low-potential (0.15 V) oxygen reduction electrocatalysis is observed, occurring through a partially oxidised state of the T2/T3 trinuclear copper cluster. Finally, chemically modified naphthoate MWCNTs exhibit high bioelectrocatalytic current densities of 3.9 mA cm(-2) under air at gas-diffusion electrode. PMID:27328033

  14. Biliverdin reductase/bilirubin mediates the anti-apoptotic effect of hypoxia in pulmonary arterial smooth muscle cells through ERK1/2 pathway

    SciTech Connect

    Song, Shasha; Wang, Shuang; Ma, Jun; Yao, Lan; Xing, Hao; Zhang, Lei; Liao, Lin; Zhu, Daling

    2013-08-01

    Inhibition of pulmonary arterial smooth muscle cell (PASMC) apoptosis induced by hypoxia plays an important role in pulmonary arterial remodeling leading to aggravate hypoxic pulmonary arterial hypertension. However, the mechanisms of hypoxia acting on PASMC apoptosis remain exclusive. Biliverdin reductase (BVR) has many essential biologic roles in physiological and pathological processes. Nevertheless, it is unclear whether the hypoxia-induced inhibition on PASMC apoptosis is mediated by BVR. In the present work, we found BVR majorly localized in PASMCs and was up-regulated in levels of protein and mRNA by hypoxia. Then we studied the contribution of BVR to anti-apoptotic response of hypoxia in PASMCs. Our results showed that siBVR, blocking generation of bilirubin, reversed the effect of hypoxia on enhancing cell survival and apoptotic protein (Bcl-2, procasepase-9, procasepase-3) expression, preventing nuclear shrinkage, DNA fragmentation and mitochondrial depolarization in starved PASMCs, which were recovered by exogenous bilirubin. Moreover, the inhibitory effect of bilirubin on PASMC apoptosis under hypoxic condition was blocked by the inhibitor of ERK1/2 pathway. Taken together, our data indicate that BVR contributes to the inhibitory process of hypoxia on PASMC apoptosis, which is mediated by bilirubin through ERK1/2 pathway. Highlights: • BVR expresses in PASMC and is up-regulated by hypoxia in protein and mRNA levels. • BVR/bilirubin contribute to the inhibitive process of hypoxia on PASMC apoptosis. • Bilirubin protects PASMC from apoptosis under hypoxia via ERK1/2 pathway.

  15. The effect of the initial reactant molar ratio and doping with Fe3+ on the formation of calcium bilirubinate in water-oil microemulsions

    NASA Astrophysics Data System (ADS)

    Liu, Siyun; Zhu, Jinmiao; Shen, Yuhua; Xie, Anjian; Zhang, Chunyan; Qiu, Lingguang

    2007-07-01

    A series of calcium bilirubinate nanoparticles were synthesized in microemulsions consisting of polyoxyethylene octylphenol ether (Triton X-100), n-hexyl alcohol (n-C6H13OH), cyclohexane (c-C6H12), and an aqueous solution. The particles were characterized by Fourier transform infrared (FT-IR) spectra, powder X-ray diffraction (XRD), inductively coupled plasma atomic emission spectrometry (ICP-AES), transmission electron microscopy (TEM), and zeta potential measurements. The results showed that the initial CaCl2/Na2BR molar ratio and the molar ratio of water to surfactant (ω0) influenced the morphology and microstructure of calcium bilirubinate nanoparticles and the sites of Ca2+ coordination to BR2-. The composition of synthesized calcium bilirubinate was nonstoichiometric. The stability of calcium bilirubinate nanoparticles dispersed in water changes as the Ca2+/BR2- molar ratio and solution pH varied. Dopant Fe2+ ions played a certain role in the simulated mineralization of calcium bilirubinate. A possible mechanism of calcium bilirubinate formation in inverse microemulsions is discussed.

  16. Early predictors of transplant-related mortality (TRM) after allogeneic bone marrow transplants (BMT): blood urea nitrogen (BUN) and bilirubin.

    PubMed

    Bacigalupo, A; Oneto, R; Bruno, B; Soracco, M; Lamparelli, T; Gualandi, F; Occhini, D; Raiola, A; Mordini, N; Berisso, G; Bregante, S; Dini, G; Lombardi, A; Lint, M V; Brand, R

    1999-09-01

    Transplant-related mortality (TRM) following allo- geneic bone marrow transplantation (BMT) remains a major concern and early identification of patients at risk may be clinically relevant. In this study we describe a predictive score based on bilirubin and blood urea nitrogen (BUN) levels on day +7 after BMT. The patient population consisted of 309 consecutive patients who underwent BMT from sibling (n = 263) or unrelated donors (n = 46) for hematologic disorders between December 1990 and December 1996. Of 27 laboratory tests taken on day +7 after BMT, serum bilirubin (P = 0.02) and BUN (P = 0.007) were found to be independent predictors of TRM in multivariate analysis. The median levels of bilirubin (0.9 mg/dl) and of BUN (21 mg/dl) were then used as a cut-off and a score of 1 was given for values equal/greater than the median. There were 216 patients with scores 0-1 (low risk) on day +7 (bilirubin <0.9 and/or BUN <21) and 93 patients with score 2 (high risk) (bilirubin >/=0.9 and BUN >/=21): the latter had more grade III-IV acute graft-versus-host disease (P = 0.03), slower neutrophil (P = 0.02) and slower platelet engraftment (P = 0.002). The actuarial 5 year TRM is 22% for low risk vs44% for high risk patients (P = 0.0003). For HLA-identical siblings TRM is 20% vs35% (P = 0.01), for unrelated donors it is 20% vs 65% (P = 0.01). Day +7 score was highly predictive of TRM on multivariate analysis (hazard ratio 1.9, P < 0.01), after adjustment for year of transplant (P < 0.00001), unrelated vs sibling donors (P = 0.001), patient age (P = 0.01) and diagnosis (P = 0.01). These results were validated on an independent group of 82 allogeneic BMT recipients in a pediatric Unit who showed an actuarial TRM of 16% for low risk vs 46% for high risk patients (P = 0.002). This study suggests that it may be possible to identify patients with different risks of TRM on day +7 after BMT: high risk patients could be eligible for programs designed to intensify prophylaxis of post

  17. Ameliorative Effect of Vanillic Acid on Serum Bilirubin, Chronotropic and Dromotropic Properties in the Cholestasis-Induced Model Rats

    PubMed Central

    Atefipour, Narges; Dianat, Mahin; Badavi, Mohammad; Sarkaki, Alireza

    2016-01-01

    Introduction The liver modulates several important roles, such as metabolism and liver cirrhosis, which have several cardiovascular problems. Due to preservative role of antioxidant agents in cardiovascular disease, consequently, many of them are applied as medicinal plants in traditional medicine. Vanillic acid (VA), as an antioxidant agent, has a principal preservative role on some diseases. In this study, the effect of vanillic acid was examined on heart rate (as chronotropic property), P-R interval (as dromotropic property), and serum bilirubin in cholestasis-induced model rats. Methods In this study, 32 male Sprague-Dawley rats weighing 200–250 g were allocated into four groups, and each group contained eight rats as follows: Control (normal saline, 1 ml/kg, gavage, daily for 4 weeks), cirrhotic (normal saline, 1 ml/kg, gavage, daily for 4 weeks), vanillic acid (10 mg/kg, gavage, daily for 4 weeks), cirrhotic treated with vanillic acid (10 mg/kg, gavage, daily for 4 weeks). Chronic biliary cirrhosis was induced in cirrhotic groups by four weeks Bile Duct Ligation (BDL). At the first day and four weeks after surgery, the animals were anesthetized, electrocardiograms were recorded (lead II), and chronotropic and dromotropic properties (HR and PR interval) were investigated. At the end of experimental duration, the animals were anesthetized, and blood samples were taken to measure serum bilirubin. The results were analyzed using t-test and one-way ANOVA by SPSS software, version 22. Results After induced of BDL, the results presented that laboratory parameter (bilirubin) in the cirrhotic group significantly increased compared to the control group. The P-R interval was reduced in the cirrhotic group compared to the control group, and there was no significant difference between heart rate in all groups. Bilirubin were reduced in cirrhotic groups treated with vanillic acid (VA) compared to cirrhotic group and also administration of VA in the cirrhotic treated with

  18. Monoclonal antibodies to the alternative oxidase of higher plant mitochondria

    SciTech Connect

    Elthon, T.E.; Nickels, R.L.; McIntosh, L. )

    1989-04-01

    The higher plant mitochondrial electron transport chain contains, in addition to the cytochrome chain which terminates with cytochrome oxidase, an alternative pathway that terminates with an alternative oxidase. The alternative oxidase of Sauromatum guttatum Schott has recently been identified as a cluster of proteins with apparent M{sub r} of 37, 36, and 35 kilodaltons (kD). Monoclonal antibodies have now been prepared to these proteins and designated as AOA (binding all three proteins of the alternative oxidase cluster), AOU (binding the upper or 37 kD protein), and AOL (binding the lower or 36 and 35 kD proteins). All three antibodies bind to their respective alternative oxidase proteins whether the proteins are in their native or denatured states. AOA and AOU inhibit alternative oxidase activity around 49%, whereas AOL inhibits activity only 14%. When coupled individually to Sepharose 4B, all three monoclonal resins were capable of retaining the entire cluster of alternative oxidase proteins, suggesting that these proteins are physically associated in some manner. The monoclonals were capable of binding similar mitochondrial proteins in a number of thermogenic and nonthermogenic species, indicating that they will be useful in characterizing and purifying the alternative oxidase of different systems. The ability of the monoclonal-Sepharose 4B resins to retain the cluster of previously identified alternative oxidase proteins, along with the inhibition of alternative oxidase activity by these monoclonals, supports the role of these proteins in constituting the alternative oxidase.

  19. Heme Degradation by Heme Oxygenase Protects Mitochondria but Induces ER Stress via Formed Bilirubin

    PubMed Central

    Müllebner, Andrea; Moldzio, Rudolf; Redl, Heinz; Kozlov, Andrey V.; Duvigneau, J. Catharina

    2015-01-01

    Heme oxygenase (HO), in conjunction with biliverdin reductase, degrades heme to carbon monoxide, ferrous iron and bilirubin (BR); the latter is a potent antioxidant. The induced isoform HO-1 has evoked intense research interest, especially because it manifests anti-inflammatory and anti-apoptotic effects relieving acute cell stress. The mechanisms by which HO mediates the described effects are not completely clear. However, the degradation of heme, a strong pro-oxidant, and the generation of BR are considered to play key roles. The aim of this study was to determine the effects of BR on vital functions of hepatocytes focusing on mitochondria and the endoplasmic reticulum (ER). The affinity of BR to proteins is a known challenge for its exact quantification. We consider two major consequences of this affinity, namely possible analytical errors in the determination of HO activity, and biological effects of BR due to direct interaction with protein function. In order to overcome analytical bias we applied a polynomial correction accounting for the loss of BR due to its adsorption to proteins. To identify potential intracellular targets of BR we used an in vitro approach involving hepatocytes and isolated mitochondria. After verification that the hepatocytes possess HO activity at a similar level as liver tissue by using our improved post-extraction spectroscopic assay, we elucidated the effects of increased HO activity and the formed BR on mitochondrial function and the ER stress response. Our data show that BR may compromise cellular metabolism and proliferation via induction of ER stress. ER and mitochondria respond differently to elevated levels of BR and HO-activity. Mitochondria are susceptible to hemin, but active HO protects them against hemin-induced toxicity. BR at slightly elevated levels induces a stress response at the ER, resulting in a decreased proliferative and metabolic activity of hepatocytes. However, the proteins that are targeted by BR still have

  20. Heme oxygenase-1 induction prevents neuronal damage triggered during mitochondrial inhibition: role of CO and bilirubin.

    PubMed

    Orozco-Ibarra, Marisol; Estrada-Sánchez, Ana María; Massieu, Lourdes; Pedraza-Chaverrí, José

    2009-06-01

    Heme oxygenase (HO) catalyzes the breakdown of heme to iron, carbon monoxide (CO), and biliverdin, the latter being further reduced to bilirubin (BR). A protective role of the inducible isoform, HO-1, has been described in pathological conditions associated with reactive oxygen species (ROS) and oxidative damage. The aim of this study was to investigate the role of HO-1 in the neurotoxicity induced by the mitochondrial toxin 3-nitropropionic acid (3-NP) in primary cultures of cerebellar granule neurons (CGNs). Toxicity of 3-NP is associated with ROS production, and this metabolic toxin has been used to mimic pathological conditions such as Huntington's disease. We found that cell death caused by 3-NP exposure was exacerbated by inhibition of HO with tin mesoporphyrin (SnMP). In addition, HO-1 up-regulation induced by the exposure to cobalt protoporphyrin (CoPP) before the incubation with 3-NP, prevented the cell death and the increase in ROS induced by 3-NP. Interestingly, addition of SnMP to CoPP-pretreated CGNs exposed to 3-NP, abolished the protective effect of CoPP suggesting that HO activity was responsible for this protective effect. This was additionally supported by the fact that CORM-2, a CO-releasing molecule, and BR, were able to protect against cell death and the increase in ROS induced by 3-NP. Our data clearly show that HO-1 elicits in CGNs a neuroprotective action against the neurotoxicity of 3-NP and that CO and BR may be involved, at least in part, in this protective effect. The present results increase our knowledge about the role of HO-1 in neuropathological conditions. PMID:19063990

  1. The Negative Relationship between Bilirubin Level and Diabetic Retinopathy: A Meta-Analysis

    PubMed Central

    Wu, Xiaomei; Ning, Kang; Jiang, Feng; Zhang, Lu

    2016-01-01

    Objectives Findings on the relationship between total bilirubin level (TBL) and diabetic retinopathy (DR) are inconsistent. Thus, we carried out a meta-analysis to investigate the relationship between TBL and the risk of DR. Methods Relevant studies were selected from six databases up to 31 May 2016 using a search strategy. The relevant data were extracted from the included studies according to the inclusion and exclusion criteria, and the mean value with standard errors or odds ratio (OR) with 95% confidence intervals (CIs) were calculated. We compared TBL in patients with DR with that in patients with diabetes but without retinopathy (NDR), and analyzed the dose-response relationship between TBL and the risk of DR. Results Twenty-four studies were selected in this meta-analysis. Twenty studies were included to calculate the pooled SMD, and the results showed that TBL in the DR group was lower than that in the NDR group (SMD: –0.52, 95% CI: –0.67, –0.38). Nine studies were included to calculate the pooled ORs, and the results showed that there was a significant negative relationship between TBL and the risk of DR (OR: 0.19, 95% CI: 0.14, 0.25). Six studies were included to investigate the dose-response relationship between TBL and the risk of DR, and we found a nonlinear relationship between TBL and the risk of DR. The results of our meta-analysis were found to be reliable using subgroup and sensitivity analyses. Conclusions The results of our meta-analysis indicate that higher TBL may be protective against DR in subjects with diabetes, and TBL could be used as a biomarker to predict the risk of DR. PMID:27571522

  2. Universal bounds on current fluctuations

    NASA Astrophysics Data System (ADS)

    Pietzonka, Patrick; Barato, Andre C.; Seifert, Udo

    2016-05-01

    For current fluctuations in nonequilibrium steady states of Markovian processes, we derive four different universal bounds valid beyond the Gaussian regime. Different variants of these bounds apply to either the entropy change or any individual current, e.g., the rate of substrate consumption in a chemical reaction or the electron current in an electronic device. The bounds vary with respect to their degree of universality and tightness. A universal parabolic bound on the generating function of an arbitrary current depends solely on the average entropy production. A second, stronger bound requires knowledge both of the thermodynamic forces that drive the system and of the topology of the network of states. These two bounds are conjectures based on extensive numerics. An exponential bound that depends only on the average entropy production and the average number of transitions per time is rigorously proved. This bound has no obvious relation to the parabolic bound but it is typically tighter further away from equilibrium. An asymptotic bound that depends on the specific transition rates and becomes tight for large fluctuations is also derived. This bound allows for the prediction of the asymptotic growth of the generating function. Even though our results are restricted to networks with a finite number of states, we show that the parabolic bound is also valid for three paradigmatic examples of driven diffusive systems for which the generating function can be calculated using the additivity principle. Our bounds provide a general class of constraints for nonequilibrium systems.

  3. Bound charges and currents

    NASA Astrophysics Data System (ADS)

    Herczyński, Andrzej

    2013-03-01

    Bound charges and currents are among the conceptually challenging topics in advanced courses on electricity and magnetism. It may be tempting for students to believe that they are merely computational tools for calculating electric and magnetic fields in matter, particularly because they are usually introduced through abstract manipulation of integral identities, with the physical interpretation provided a posteriori. Yet these charges and currents are no less real than free charges and currents and can be measured experimentally. A simpler and more direct approach to introducing this topic, suggested by the ideas in the classic book by Purcell and emphasizing the physical origin of these phenomena, is proposed.

  4. Metal content and subunit composition of bean and bovine cytochrome C oxidases

    SciTech Connect

    Sowa, S.; Yewey, G.L.; Roos, E.E.; Caughey, W.S.

    1986-05-01

    Cytochrome c oxidase (CcO) isolated from imbibed whole beans (Phaseolus vulgaris) and bovine hearts have been compared in terms of metal content, subunit composition, and dioxygen reduction site. Both preparations contain Zn and Mg as well as Cu and Fe, with similar Fe/Zn ratios (2.5). Both enzymes also contain multiple subunits of comparable molecular weights and exhibit sharp CO-IR specta for heme iron bound carbonyl species. Alkaline detergent treatment of the bovine enzyme (i.e. incubation with 1% Triton X-100, pH 9.5, followed by ion exchange chromatography) has been reported by others to remove subunit III with loss of H+ pumping activity in reconstituted proteoliposomes without loss of oxidase activity. However, the authors find the treatment does not alter subunit distribution as visualized by SDS-Urea PAGE or HPLC analysis and markedly reduces activity (90%). Metal atom ratios of the treated enzyme indicate a loss of Fe and Mg (Cu/Fe 2.33 +/- 0.24, Fe/Zn 1.64 +/- 0.34, Mg/Zn 0.76 +/- 0.22) relative to native CcO (Cu/Fe 1.23 +/-0.06, Fe/Zn 2.47 +/- 0.14, Mg/Zn 0.94 +/- 0.05). These data provide evidence for similarity between plant and mammalian oxidases and lend further support for vital roles of Zn, Mg, Fe, and Cu in CcO.

  5. Mechanism-based inactivators of plant copper/quinone containing amine oxidases.

    PubMed

    Longu, Silvia; Mura, Anna; Padiglia, Alessandra; Medda, Rosaria; Floris, Giovanni

    2005-08-01

    Copper/quinone amine oxidases contain Cu(II) and the quinone of 2,4,5-trihydroxyphenylalanine (topaquinone; TPQ) as cofactors. TPQ is derived by post-translational modification of a conserved tyrosine residue in the protein chain. Major advances have been made during the last decade toward understanding the structure/function relationships of the active site in Cu/TPQ amine oxidases using specific inhibitors. Mechanism-based inactivators are substrate analogues that bind to the active site of an enzyme being accepted and processed by the normal catalytic mechanism of the enzyme. During the reaction a covalent modification of the enzyme occurs leading to irreversible inactivation. In this review mechanism-based inactivators of plant Cu/TPQ amine oxidases from the pulses lentil (Lens esculenta), pea (Pisum sativum), grass pea (Lathyrus sativus) and sainfoin (Onobrychis viciifolia,) are described. Substrates forming, in aerobiotic and in anaerobiotic conditions, killer products that covalently bound to the quinone cofactor or to a specific amino acid residue of the target enzyme are all reviewed. PMID:16054177

  6. Inactivation of monoamine oxidase by allylamine does not result in flavin attachment

    SciTech Connect

    Silverman, R.B.; Hiebert, C.K.; Vazquez, M.L.

    1985-11-25

    (1-TH)Allylamine was synthesized by sodium boro(TH)hydride reduction of acrolein followed by direct conversion of the (1-TH)allyl alcohol to N-allylphthalimide with triphenylphosphine, diethylazodicarboxylate, and phthalimide. The protecting group was removed with hydrazine. Inactivation of beef liver mitochondrial monoamine oxidase with (1-TH)allylamine led to incorporation of 1-6 eq of inactivator/active site depending upon the length of incubation time. Inactivation and radioactivity incorporation coincided; however, after 1 eq of tritium was incorporated and 5% enzyme activity remained, additional radioactivity continued to become incorporated into the enzyme. The optical spectrum of the FAD coenzyme changed during inactivation from that of oxidized to reduced flavin. Following dialysis of the inactivated enzyme, the spectrum remained reduced, but denaturation in urea rapidly resulted in reoxidation of the flavin. Under these same denaturing conditions, 96% of the radioactivity associated with the enzyme remained bound, therefore indicating that allylamine attachment is not to the flavin coenzyme but rather to an active site amino acid residue. The adduct also was stable to base and, to a lesser degree, acid treatment. Although allylamine and N-cyclopropylbenzylamine appear to be oxidized by monoamine oxidase to give 3-(amino acid residue) propanal adducts, two different amino acids seem to be involved because of a difference in stability of the adducts. The mechanisms for inactivation of monoamine oxidase by allylamine and reactivation by benzylamine are discussed in relation to previously reported results.

  7. NADPH oxidases in the arbuscular mycorrhizal symbiosis.

    PubMed

    Belmondo, Simone; Calcagno, Cristina; Genre, Andrea; Puppo, Alain; Pauly, Nicolas; Lanfranco, Luisa

    2016-04-01

    Plant NADPH oxidases are the major source of reactive oxygen species (ROS) that plays key roles as both signal and stressor in several plant processes, including defense responses against pathogens. ROS accumulation in root cells during arbuscular mycorrhiza (AM) development has raised the interest in understanding how ROS-mediated defense programs are modulated during the establishment of this mutualistic interaction. We have recently analyzed the expression pattern of 5 NADPH oxidase (also called RBOH) encoding genes in Medicago truncatula, showing that only one of them (MtRbohE) is specifically upregulated in arbuscule-containing cells. In line with this result, RNAi silencing of MtRbohE generated a strong alteration in root colonization, with a significant reduction in the number of arbusculated cells. On this basis, we propose that MtRBOHE-mediated ROS production plays a crucial role in the intracellular accommodation of arbuscules. PMID:27018627

  8. Human copper-dependent amine oxidases.

    PubMed

    Finney, Joel; Moon, Hee-Jung; Ronnebaum, Trey; Lantz, Mason; Mure, Minae

    2014-03-15

    Copper amine oxidases (CAOs) are a class of enzymes that contain Cu(2+) and a tyrosine-derived quinone cofactor, catalyze the conversion of a primary amine functional group to an aldehyde, and generate hydrogen peroxide and ammonia as byproducts. These enzymes can be classified into two non-homologous families: 2,4,5-trihydroxyphenylalanine quinone (TPQ)-dependent CAOs and the lysine tyrosylquinone (LTQ)-dependent lysyl oxidase (LOX) family of proteins. In this review, we will focus on recent developments in the field of research concerning human CAOs and the LOX family of proteins. The aberrant expression of these enzymes is linked to inflammation, fibrosis, tumor metastasis/invasion and other diseases. Consequently, there is a critical need to understand the functions of these proteins at the molecular level, so that strategies targeting these enzymes can be developed to combat human diseases. PMID:24407025

  9. Imaging Monoamine Oxidase in the Human Brain

    SciTech Connect

    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.

  10. INCREASED XANTHINE OXIDASE IN THE SKIN OF PREECLAMPTIC WOMEN

    PubMed Central

    Bainbridge, Shannon A.; Deng, Jau-Shyong; Roberts, James M.

    2010-01-01

    Xanthine oxioreductase is the holoenzyme responsible for terminal purine catabolism. Under conditions of metabolic stress or heightened pro-inflammatory cytokine production this enzyme is preferentially in it’s oxidized form, xanthine oxidase, with catalytic action that generates uric acid and the free radical superoxide. As preeclampsia is characterized by heightened inflammation, oxidative stress and hyperuricemia it has been proposed that xanthine oxidase plays a pivotal role in this hypertensive disorder of pregnancy. We sought to determine whether xanthine oxidase protein content was higher in maternal tissue of preeclamptic mothers, compared to healthy pregnant controls, using immunohistochemical analysis of skin biopsies. We further compared xanthine oxidase immunoreactivity in skin biopsies from preeclamptic women and patients with several inflammatory conditions. In preeclamptic women, intense xanthine oxidase immunoreactivity was present within the epidermis. By contrast, only very faint xanthine oxidase staining was observed in skin biopsies from healthy pregnant controls. Further, a role for inflammation in the increase of xanthine oxidase was suggested by similar findings of heightened xanthine oxidase immunoreactivity in the skin biopsies from non-pregnant individuals diagnosed with conditions of systemic inflammation. The finding of increased xanthine oxidase in maternal tissue, most likely as the result of heightened maternal inflammation, suggest maternal xanthine oxidase as a source of free radical and uric acid generation in preeclampsia. PMID:19196876

  11. BEHAVIORAL OUTCOMES OF MONOAMINE OXIDASE DEFICIENCY: PRECLINICAL AND CLINICAL EVIDENCE

    PubMed Central

    Bortolato, Marco; Shih, Jean C.

    2012-01-01

    Monoamine oxidase (MAO) isoenzymes A and B are mitochondrial-bound proteins, catalyzing the oxidative deamination of monoamine neurotransmitters as well as xenobiotic amines. Although they derive from a common ancestral progenitor gene, are located at X-chromosome and display 70% structural identity, their substrate preference, regional distribution, and physiological role are divergent. In fact, while MAO-A has high affinity for serotonin and norepinephrine, MAO-B primarily serves the catabolism of 2-phenylethylamine (PEA) and contributes to the degradation of other trace amines and dopamine. Convergent lines of preclinical and clinical evidence indicate that variations in MAO enzymatic activity—due to either genetic or environmental factors—can exert a profound influence on behavioral regulation and play a role in the pathophysiology of a large spectrum of mental and neurodegenerative disorders, ranging from antisocial personality disorder to Parkinson’s disease. Over the past few years, numerous advances have been made in our understanding of the phenotypical variations associated with genetic polymorphisms and mutations of the genes encoding for both isoenzymes. In particular, novel findings on the phenotypes of MAO-deficient mice are highlighting novel potential implications of both isoenzymes in a broad spectrum of mental disorders, ranging from autism and anxiety to impulse-control disorders and ADHD. These studies will lay the foundation for future research on the neurobiological and neurochemical bases of these pathological conditions, as well as the role of gene × environment interactions in the vulnerability to several mental disorders. PMID:21971001

  12. Reexamining Michaelis-Menten enzyme kinetics for xanthine oxidase.

    PubMed

    Bassingthwaighte, James B; Chinn, Tamara M

    2013-03-01

    Abbreviated expressions for enzyme kinetic expressions, such as the Michaelis-Menten (M-M) equations, are based on the premise that enzyme concentrations are low compared with those of the substrate and product. When one does progress experiments, where the solute is consumed during conversion to form a series of products, the idealized conditions are violated. Here, we analyzed data of xanthine oxidase in vitro from Escribano et al. (Biochem J 254: 829, 1988) on two conversions in series, hypoxanthine to xanthine to uric acid. Analyses were done using four models: standard irreversible M-M reactions (model 1), Escribano et al.'s M-M forward reaction expressions with product inhibition (model 2), fully reversible M-M equations (model 3), and standard differential equations allowing forward and backward reactions with mass balance accounting for binding (model 4). The results showed that the need for invoking product inhibition vanishes with more complete analyses. The reactions were not quite irreversible, so the backward reaction had a small effect. Even though the enzyme concentration was only 1-2% of the initial substrate concentrations, accounting for the fraction of solutes bound to the enzyme did influence the parameter estimates, but in this case, the M-M model overestimated Michaelis constant values by only about one-third. This article also presents the research and models in a reproducible and publicly available form. PMID:23471247

  13. Reexamining Michaelis-Menten enzyme kinetics for xanthine oxidase

    PubMed Central

    Chinn, Tamara M.

    2013-01-01

    Abbreviated expressions for enzyme kinetic expressions, such as the Michaelis-Menten (M-M) equations, are based on the premise that enzyme concentrations are low compared with those of the substrate and product. When one does progress experiments, where the solute is consumed during conversion to form a series of products, the idealized conditions are violated. Here, we analyzed data of xanthine oxidase in vitro from Escribano et al. (Biochem J 254: 829, 1988) on two conversions in series, hypoxanthine to xanthine to uric acid. Analyses were done using four models: standard irreversible M-M reactions (model 1), Escribano et al.'s M-M forward reaction expressions with product inhibition (model 2), fully reversible M-M equations (model 3), and standard differential equations allowing forward and backward reactions with mass balance accounting for binding (model 4). The results showed that the need for invoking product inhibition vanishes with more complete analyses. The reactions were not quite irreversible, so the backward reaction had a small effect. Even though the enzyme concentration was only 1–2% of the initial substrate concentrations, accounting for the fraction of solutes bound to the enzyme did influence the parameter estimates, but in this case, the M-M model overestimated Michaelis constant values by only about one-third. This article also presents the research and models in a reproducible and publicly available form. PMID:23471247

  14. Purification and characterization of polyphenol oxidase from fresh ginseng

    PubMed Central

    Kim, Jae-Joon; Kim, Woo-Yeon

    2013-01-01

    Polyphenol oxidase (PPO) was purified from fresh ginseng roots using acetone precipitation, carboxymethyl (CM)-Sepharose chromatography, and phenyl-Sepharose chromatography. Two isoenzymes (PPO 1 and PPO 2) were separated using an ion-exchange column with CM-Sepharose. PPO 1 was purified up to 13.2-fold with a 22.6% yield. PPO 2 bound to CM-Sepharose, eluted with NaCl, and was purified up to 22.5-fold with a 17.4% yield. PPO 2 was further chromatographed on phenyl-Sepharose. The molecular weight of the purified PPO 2 from fresh ginseng was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and was about 40 kDa. The optimum temperature and pH were 20℃ and 7.0, respectively, using catechol as a substrate. Pyrogallol showed the highest substrate specificity. The effect of a PPO inhibitor showed that its activity increased slightly in the presence of a low concentration of citric acid. High concentrations of acidic compounds and sulfite agents significantly inhibited purified ginseng PPO 2. PMID:23717165

  15. Secretory production of an FAD cofactor-containing cytosolic enzyme (sorbitol-xylitol oxidase from Streptomyces coelicolor) using the twin-arginine translocation (Tat) pathway of Corynebacterium glutamicum.

    PubMed

    Scheele, Sandra; Oertel, Dan; Bongaerts, Johannes; Evers, Stefan; Hellmuth, Hendrik; Maurer, Karl-Heinz; Bott, Michael; Freudl, Roland

    2013-03-01

    Carbohydrate oxidases are biotechnologically interesting enzymes that require a tightly or covalently bound cofactor for activity. Using the industrial workhorse Corynebacterium glutamicum as the expression host, successful secretion of a normally cytosolic FAD cofactor-containing sorbitol-xylitol oxidase from Streptomyces coelicolor was achieved by using the twin-arginine translocation (Tat) protein export machinery for protein translocation across the cytoplasmic membrane. Our results demonstrate for the first time that, also for cofactor-containing proteins, a secretory production strategy is a feasible and promising alternative to conventional intracellular expression strategies. PMID:23163932

  16. Secretory production of an FAD cofactor-containing cytosolic enzyme (sorbitol–xylitol oxidase from Streptomyces coelicolor) using the twin-arginine translocation (Tat) pathway of Corynebacterium glutamicum

    PubMed Central

    Scheele, Sandra; Oertel, Dan; Bongaerts, Johannes; Evers, Stefan; Hellmuth, Hendrik; Maurer, Karl-Heinz; Bott, Michael; Freudl, Roland

    2013-01-01

    Carbohydrate oxidases are biotechnologically interesting enzymes that require a tightly or covalently bound cofactor for activity. Using the industrial workhorse Corynebacterium glutamicum as the expression host, successful secretion of a normally cytosolic FAD cofactor-containing sorbitol–xylitol oxidase from Streptomyces coelicolor was achieved by using the twin-arginine translocation (Tat) protein export machinery for protein translocation across the cytoplasmic membrane. Our results demonstrate for the first time that, also for cofactor-containing proteins, a secretory production strategy is a feasible and promising alternative to conventional intracellular expression strategies. PMID:23163932

  17. Higher Serum Direct Bilirubin Levels Were Associated with a Lower Risk of Incident Chronic Kidney Disease in Middle Aged Korean Men

    PubMed Central

    Ryu, Seungho; Chang, Yoosoo; Zhang, Yiyi; Woo, Hee-Yeon; Kwon, Min-Jung; Park, Hyosoon; Lee, Kyu-Beck; Son, Hee Jung; Cho, Juhee; Guallar, Eliseo

    2014-01-01

    Background The association between serum bilirubin levels and incident chronic kidney disease (CKD) in the general population is unknown. We aimed to examine the association between serum bilirubin concentration (total, direct, and indirect) and the risk of incident CKD. Methods and Findings Longitudinal cohort study of 12,823 Korean male workers 30 to 59 years old without CKD or proteinuria at baseline participating in medical health checkup program in a large worksite. Study participants were followed for incident CKD from 2002 through 2011. Estimated glomerular filtration rate (eGFR) was estimated by using the CKD-EPI equation. CKD was defined as eGFR <60 mL/min per 1.73 m2. Parametric Cox models and pooled logistic regression models were used to estimate adjusted hazard ratios for incident CKD. We observed 238 incident cases of CKD during 70,515.8 person-years of follow-up. In age-adjusted models, the hazard ratios for CKD comparing quartiles 2–4 vs. quartile 1 of serum direct bilirubin were 0.93 (95% CI 0.67–1.28), 0.88 (0.60–1.27) and 0.60 (0.42–0.88), respectively. In multivariable models, the adjusted hazard ratio for CKD comparing the highest to the lowest quartile of serum direct bilirubin levels was 0.60 (95% CI 0.41–0.87; P trend = 0.01). Neither serum total nor indirect bilirubin levels were significantly associated with the incidence of CKD. Conclusions Higher serum direct bilirubin levels were significantly associated with a lower risk of developing CKD, even adjusting for a variety of cardiometabolic parameters. Further research is needed to elucidate the mechanisms underlying this association and to establish the role of serum direct bilirubin as a marker for CKD risk. PMID:24586219

  18. The caa3 terminal oxidase of Bacillus stearothermophilus. Transient spectroscopy of electron transfer and ligand binding.

    PubMed

    Giuffrè, A; D'Itri, E; Giannini, S; Brunori, M; Ubbink-Kok, T; Konings, W N; Antonini, G

    1996-06-14

    The thermophilic bacterium Bacillus stearothermophilus possesses a caa3-type terminal oxidase, which was previously purified (De Vrij, W., Heyne, R. I. R., and Konings, W. N. (1989) Eur. J. Biochem. 178, 763-770). We have carried out extensive kinetic experiments on the purified enzyme by stopped-flow time-resolved optical spectroscopy combined with singular value decomposition analysis. The results indicate a striking similarity of behavior between this enzyme and the electrostatic complex between mammalian cytochrome c and cytochrome c oxidase. CO binding to fully reduced caa3 occurs with a second order rate constant (k = 7.8 x 10(4)M-1 s-1) and an activation energy (E* = 6.1 kcal mol-1) similar to those reported for beef heart cytochrome c oxidase. Dithionite reduces cytochrome a with bimolecular kinetics, while cytochrome a3 (and CuB) is reduced via intramolecular electron transfer. When the fully reduced enzyme is mixed with O2, cytochrome a3, and cytochrome c are rapidly oxidized, whereas cytochrome a remains largely reduced in the first few milliseconds. When cyanide-bound caa3 is mixed with ascorbate plus TMPD, cytochrome c and cytochrome a are synchronously reduced; the value of the second order rate constant (k = 3 x 10(5) M-1 s-1 at 30 degrees C) suggests that cytochrome c is the electron entry site. Steady-state experiments indicate that cytochrome a has a redox potential higher than cytochrome c. The data from the reaction with O2 reveal a remarkable similarity in the kinetic, equilibrium, and optical properties of caa3 and the electrostatic complex cytochrome c/cytochrome c oxidase. PMID:8662862

  19. Cytochrome P450 2A5 and bilirubin: Mechanisms of gene regulation and cytoprotection

    SciTech Connect

    Kim, Sangsoo Daniel; Antenos, Monica; Squires, E. James; Kirby, Gordon M.

    2013-07-15

    Bilirubin (BR) has recently been identified as the first endogenous substrate for cytochrome P450 2A5 (CYP2A5) and it has been suggested that CYP2A5 plays a major role in BR clearance as an alternative mechanism to BR conjugation by uridine-diphosphate glucuronyltransferase 1A1. This study investigated the mechanisms of Cyp2a5 gene regulation by BR and the cytoprotective role of CYP2A5 in BR hepatotoxicity. BR induced CYP2A5 expression at the mRNA and protein levels in a dose-dependent manner in primary mouse hepatocytes. BR treatment also caused nuclear translocation of Nuclear factor-E2 p45-related factor 2 (Nrf2) in hepatocytes. In reporter assays, BR treatment of primary hepatocytes transfected with a Cyp2a5 promoter-luciferase reporter construct resulted in a 2-fold induction of Cyp2a5 reporter activity. Furthermore, cotransfection of the hepatocytes with a Nrf2 expression vector without BR treatment resulted in an increase in Cyp2a5 reporter activity of approximately 2-fold and BR treatment of Nrf2 cotransfectants further increased reporter activity by 4-fold. In addition, site-directed mutation of the ARE in the reporter construct completely abolished both the BR- and Nrf2-mediated increases in reporter activity. The cytoprotective role of CYP2A5 against BR-mediated apoptosis was also examined in Hepa 1–6 cells that lack endogenous CYP2A5. Transient overexpression of CYP2A5 partially blocked BR-induced caspase-3 cleavage in Hepa 1–6 cells. Furthermore, in vitro degradation of BR was increased by microsomes from Hepa 1–6 cells overexpressing CYP2A5 compared to control cells transfected with an empty vector. Collectively, these results suggest that Nrf2-mediated CYP2A5 transactivation in response to BR may provide an additional mechanism for adaptive cytoprotection against BR hepatotoxicity. - Highlights: • The mechanism of Cyp2a5 gene regulation by BR was investigated. • The cytoprotective role of CYP2A5 in BR hepatotoxicity was determined. • BR

  20. Comparison of kinetic properties of amine oxidases from sainfoin and lentil and immunochemical characterization of copper/quinoprotein amine oxidases.

    PubMed

    Zajoncová, L; Frébort, I; Luhová, L; Sebela, M; Galuszka, P; Pec, P

    1999-01-01

    Kinetic properties of novel amine oxidase isolated from sainfoin (Onobrychis viciifolia) were compared to those of typical plant amine oxidase (EC 1.4.3.6) from lentil (Lens culinaris). The amine oxidase from sainfoin was active toward substrates, such as 1,5-diaminopentane (cadaverine) with K(m) of 0.09 mM and 1,4-diaminobutane (putrescine) with K(m) of 0.24 mM. The maximum rate of oxidation for cadaverine at saturating concentration was 2.7 fold higher than that of putrescine. The amine oxidase from lentil had the maximum rate for putrescine comparable to the rate of sainfoin amine oxidase with the same substrate. Both amine oxidases, like other plant Cu-amine oxidases, were inhibited by substrate analogs (1,5-diamino-3-pentanone, 1,4-diamino-2-butanone and aminoguanidine), Cu2+ chelating agents (diethyltriamine, 1,10-phenanthroline, 8-hydroxyquinoline, 2,2'-bipyridyl, imidazole, sodium cyanide and sodium azide), some alkaloids (L-lobeline and cinchonine), some lathyrogens (beta-aminopropionitrile and aminoacetonitrile) and other inhibitors (benzamide oxime, acetone oxime, hydroxylamine and pargyline). Tested by Ouchterlony's double diffusion in agarose gel, polyclonal antibodies against the amine oxidase from sainfoin, pea and grass pea cross-reacted with amine oxidases from several other Fabaceae and from barley (Hordeum vulgare) of Poaceae, while amine oxidase from the filamentous fungus Aspergillus niger did not cross-react at all. However, using Western blotting after SDS-PAGE with rabbit polyclonal antibodies against the amine oxidase from Aspergillus niger, some degree of similarity of plant amine oxidases from sainfoin, pea, field pea, grass pea, fenugreek, common melilot, white sweetclover and Vicia panonica with the A. niger amine oxidase was confirmed. PMID:10092944

  1. Pathological changes in platelet histamine oxidases in atopic eczema

    PubMed Central

    Ionescu, Gruia

    1993-01-01

    Increased plasma histamine levels were associated with significantly lowered diamine and type B monoamine oxidase activities in platelet-rich plasma of atopic eczema (AE) patients. The diamine oxidase has almost normal cofactor levels (pyridoxal phosphate and Cu2+) but the cofactor levels for type B monoamine oxidase (flavin adenine dinucleotide and Fe2+) are lowered. The biogenic amines putrescine, cadaverine, spermidine, spermine, tyramine and serotonin in the sera, as well as dopamine and epinephrine in EDTA-plasma were found to be normal. It is unlikely, therefore, that these amines are responsible for the decreased activities of monoamine and diamine oxidase in these patients. The most likely causative factors for the inhibition of the diamine oxidase are nicotine, alcohol, food additives and other environmental chemicals, or perhaps a genetic defect of the diamine oxidase. PMID:18475554

  2. Reconstitution of Cu2+-depleted bovine serum amine oxidase with Co2+.

    PubMed Central

    Agostinelli, E; De Matteis, G; Mondovì, B; Morpurgo, L

    1998-01-01

    Two different Cu2+-depleted derivatives of bovine serum amine oxidase (BSAO) have recently been prepared, which contain about 0.5 mol/dimer of phenylhydrazine-reactive topa quinone (TPQ) cofactor and, depending on the reagents used, about 0.2 or 0.7 residual Cu2+/dimer [Agostinelli, De Matteis, Sinibaldi, Mondovi and Morpurgo (1997) Biochem. J. 324, 497-501]. The benzylamine oxidase activity of both derivatives was <5% and increased up to approximately 20% on incorporation of Co2+, irrespective of the residual Cu2+ content, which was unaffected by the treatment according to atomic absorption and ESR spectroscopy. The residual Cu2+ ions appeared to be distributed one per dimer and to be bound to inactive subunits, whereas Co2+ was bound to active subunits. The change in the active site had an appreciable influence on the kinetic behaviour. With several amines, the kinetic parameters, Km and kc, measured for Co2+-BSAO were different from those for native BSAO. This excludes the possibility that the catalytic activity was due to residual Cu2+. Furthermore, Co2+ restored to nearly native level the intensity of the TPQ 480 nm band and the reactions with phenylhydrazine or benzylhydrazine, which had been slowed down or abolished, respectively, in Cu2+-depleted samples. The CD spectrum, measured for the derivative with low Cu2+ content, was compatible with Co2+ binding to the copper site. The amine oxidase activity of the Co2+ derivative, which cannot form a semiquinone radical as an intermediate of the catalytic reaction, strongly suggests that the Cu+-semiquinone is not an obligatory intermediate of BSAO catalytic pathway. PMID:9461534

  3. Reduction of carbon monoxide to formaldehyde by the terminal oxidase of the marine bacterium Pseudomonas nautica strain 617.

    PubMed

    Arnaud, S; Malatesta, F; Denis, M

    1992-01-27

    When exposed to CO, the aerobic respiratory system of the marine bacterium Pseudomonas nautica strain 617, previously reduced with dithionite, undergoes reoxidation. When dealing with the purified oxidase (dithionite reduced) exposure of the enzyme to CO induces its reoxidation (collapse of its alpha band). Under our experimental conditions, this form of the oxidase could not be reduced again by dithionite. Addition of formaldehyde to the native oxidized enzyme resulted in full inhibition of the oxidase reduction by dithionite, presumably due to complex formation. We hypothesized a reduction of CO into formaldehyde and a locking of the active site by the reaction product. By using flash photolysis, it was possible to turn over the enzyme, accumulate the reaction product and identify it as formaldehyde. When using the membrane-bound enzyme, formaldehyde accumulated without the help of flash photolysis. This unusual reduction of CO to formaldehyde could be related to the previously reported uncommon features of the P. nautica oxidase, in particular O2 reduction into H2O2 as end product [(1989) FEBS Lett. 247, 475-479]. PMID:1537399

  4. Enzymatic and energetic properties of the aerobic respiratory chain-linked NADH oxidase system in the marine bacterium Pseudomonas nautica.

    PubMed

    Cho, K H; Kim, Y J

    2000-08-31

    Membranes of Pseudomonas nautica, grown aerobically on a complex medium, oxidized both NADH and deamino-NADH as substrates. The activity of membrane-bound NADH oxidase was activated by monovalent cations including Na+, Li+, and K+. The activation by Na+ was higher than that by Li+ and K+. The maximum activity of NADH oxidase was obtained at about pH 9.0 in the presence of 0.08 M NaCl. The NADH oxidase activity was completely inhibited by 60 microM 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO), while the NADH:quinone oxidoreductase activity was about 37% inhibited by 60 microM HQNO. The activities of NADH oxidase and NADH:quinone oxidoreductase were about 40% inhibited by 60 microM rotenone. The fluorescence quenching technique revealed that electron transfer from NADH to ubiquinone-1 (Q-1) or oxygen generated a membrane potential (deltapsi) which was larger and more stable in the presence of Na+ than in the absence of Na+. However, the All was highly sensitive to a protonophore, carbonyl-cyanide m-chlorophenylhydrazone (CCCP) even at alkaline pH. PMID:10987141

  5. Gravity Responsive NADH Oxidase of the Plasma Membrane

    NASA Technical Reports Server (NTRS)

    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.

  6. Xanthine oxidase status in ethanol-intoxicated rat liver.

    PubMed

    Abbondanza, A; Battelli, M G; Soffritti, M; Cessi, C

    1989-12-01

    The status of xanthine oxidase in ethanol-induced liver injury has been investigated in the rat, by acute and chronic ethanol treatments. A 38% increase of the enzyme O-form was observed after repeated ethanol administration. Chronic intoxication caused a significant decrease of total xanthine oxidase activity after both prolonged ethanol feeding and life span ethanol ingestion. The intermediate D/O-form of xanthine oxidase (that can act either as an oxidase or as a dehydrogenase, being able to react with O2 as well as with NAD+ as electron acceptor) increased 5.5-fold after prolonged ethanol feeding. PMID:2690670

  7. FcγR-stimulated activation of the NADPH oxidase: phosphoinositide-binding protein p40phox regulates NADPH oxidase activity after enzyme assembly on the phagosome

    PubMed Central

    Tian, Wei; Li, Xing Jun; Stull, Natalie D.; Ming, Wenyu; Suh, Chang-Il; Bissonnette, Sarah A.; Yaffe, Michael B.; Grinstein, Sergio; Atkinson, Simon J.

    2008-01-01

    The phagocyte NADPH oxidase generates superoxide for microbial killing, and includes a membrane-bound flavocytochrome b558 and cytosolic p67phox, p47phox, and p40phox subunits that undergo membrane translocation upon cellular activation. The function of p40phox, which binds p67phox in resting cells, is incompletely understood. Recent studies showed that phagocytosis-induced superoxide production is stimulated by p40phox and its binding to phosphatidylinositol-3-phosphate (PI3P), a phosphoinositide enriched in membranes of internalized phagosomes. To better define the role of p40phox in FcγR-induced oxidase activation, we used immunofluorescence and real-time imaging of FcγR-induced phagocytosis. YFP-tagged p67phox and p40phox translocated to granulocyte phagosomes before phagosome internalization and accumulation of a probe for PI3P. p67phox and p47phox accumulation on nascent and internalized phagosomes did not require p40phox or PI3 kinase activity, although superoxide production before and after phagosome sealing was decreased by mutation of the p40phox PI3P-binding domain or wortmannin. Translocation of p40phox to nascent phagosomes required binding to p67phox but not PI3P, although the loss of PI3P binding reduced p40phox retention after phagosome internalization. We conclude that p40phox functions primarily to regulate FcγR-induced NADPH oxidase activity rather than assembly, and stimulates superoxide production via a PI3P signal that increases after phagosome internalization. PMID:18711001

  8. Decolorization and removal of textile and non-textile dyes from polluted wastewater and dyeing effluent by using potato (Solanum tuberosum) soluble and immobilized polyphenol oxidase.

    PubMed

    Khan, Amjad Ali; Husain, Qayyum

    2007-03-01

    Celite bound potato polyphenol oxidase preparation was employed for the treatment of wastewater/dye effluent contaminated with reactive textile and non-textile dyes, Reactive Blue 4 and Reactive Orange 86. The maximum decolorization was found at pH 3.0 and 4.0 in case of Reactive Blue 4 and Reactive Orange 86, respectively. Immobilized potato polyphenol oxidase was significantly more effective in decolorizing the individual dye and complex mixtures of dyes as compared to soluble enzyme. The absorption spectra of the treated and untreated dye mixture and dyeing effluent exhibited a marked difference in the absorption value at various wavelengths. The polluted water contaminated with an individual dye or mixtures of dyes treated with soluble and immobilized potato polyphenol oxidase resulted in the remarkable loss in total organic carbon. PMID:16765044

  9. Petawatt laser absorption bounded

    NASA Astrophysics Data System (ADS)

    Levy, Matthew; Wilks, Scott; Tabak, Max; Libby, Stephen; Baring, Matthew

    2014-10-01

    The interaction of petawatt (1015 W) lasers with solid matter forms the basis for advanced scientific applications such as table-top relativistic particle accelerators, ultrafast charged particle imaging systems and fast ignition inertial confinement fusion. Key metrics for these applications relate to absorption, yet conditions in this regime are so nonlinear that it is often impossible to know the fraction of absorbed light f, and even the range of f is unknown. In this presentation, using a relativistic Rankine-Hugoniot-like analysis, we show how to derive the theoretical maximum and minimum of f. These boundaries constrain nonlinear absorption mechanisms across the petawatt regime, forbidding high absorption values at low laser power and low absorption values at high laser power. Close agreement is shown with several dozens of published experimental data points and simulation results, helping to confirm the theory. For applications needing to circumvent the absorption bounds, these results will accelerate a shift from solid targets, towards structured and multilayer targets, and lead the development of new materials.

  10. Nox NADPH Oxidases and the Endoplasmic Reticulum

    PubMed Central

    Araujo, Thaís L.S.; Abrahão, Thalita B.

    2014-01-01

    Abstract Significance: Understanding isoform- and context-specific subcellular Nox reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase compartmentalization allows relevant functional inferences. This review addresses the interplay between Nox NADPH oxidases and the endoplasmic reticulum (ER), an increasingly evident player in redox pathophysiology given its role in redox protein folding and stress responses. Recent Advances: Catalytic/regulatory transmembrane subunits are synthesized in the ER and their processing includes folding, N-glycosylation, heme insertion, p22phox heterodimerization, as shown for phagocyte Nox2. Dual oxidase (Duox) maturation also involves the regulation by ER-resident Duoxa2. The ER is the activation site for some isoforms, typically Nox4, but potentially other isoforms. Such location influences redox/Nox-mediated calcium signaling regulation via ER targets, such as sarcoendoplasmic reticulum calcium ATPase (SERCA). Growing evidence suggests that Noxes are integral signaling elements of the unfolded protein response during ER stress, with Nox4 playing a dual prosurvival/proapoptotic role in this setting, whereas Nox2 enhances proapoptotic signaling. ER chaperones such as protein disulfide isomerase (PDI) closely interact with Noxes. PDI supports growth factor-dependent Nox1 activation and mRNA expression, as well as migration in smooth muscle cells, and PDI overexpression induces acute spontaneous Nox activation. Critical Issues: Mechanisms of PDI effects include possible support of complex formation and RhoGTPase activation. In phagocytes, PDI supports phagocytosis, Nox activation, and redox-dependent interactions with p47phox. Together, the results implicate PDI as possible Nox organizer. Future Directions: We propose that convergence between Noxes and ER may have evolutive roots given ER-related functional contexts, which paved Nox evolution, namely calcium signaling and pathogen killing. Overall, the interplay between

  11. NADPH oxidase promotes neutrophil extracellular trap formation in pulmonary aspergillosis.

    PubMed

    Röhm, Marc; Grimm, Melissa J; D'Auria, Anthony C; Almyroudis, Nikolaos G; Segal, Brahm H; Urban, Constantin F

    2014-05-01

    NADPH oxidase is a crucial enzyme in antimicrobial host defense and in regulating inflammation. Chronic granulomatous disease (CGD) is an inherited disorder of NADPH oxidase in which phagocytes are defective in generation of reactive oxidant intermediates. Aspergillus species are ubiquitous, filamentous fungi, which can cause invasive aspergillosis, a major cause of morbidity and mortality in CGD, reflecting the critical role for NADPH oxidase in antifungal host defense. Activation of NADPH oxidase in neutrophils can be coupled to the release of proteins and chromatin that comingle in neutrophil extracellular traps (NETs), which can augment extracellular antimicrobial host defense. NETosis can be driven by NADPH oxidase-dependent and -independent pathways. We therefore undertook an analysis of whether NADPH oxidase was required for NETosis in Aspergillus fumigatus pneumonia. Oropharyngeal instillation of live Aspergillus hyphae induced neutrophilic pneumonitis in both wild-type and NADPH oxidase-deficient (p47(phox-/-)) mice which had resolved in wild-type mice by day 5 but progressed in p47(phox-/-) mice. NETs, identified by immunostaining, were observed in lungs of wild-type mice but were absent in p47(phox-/-) mice. Using bona fide NETs and nuclear chromatin decondensation as an early NETosis marker, we found that NETosis required a functional NADPH oxidase in vivo and ex vivo. In addition, NADPH oxidase increased the proportion of apoptotic neutrophils. Together, our results show that NADPH oxidase is required for pulmonary clearance of Aspergillus hyphae and generation of NETs in vivo. We speculate that dual modulation of NETosis and apoptosis by NADPH oxidase enhances antifungal host defense and promotes resolution of inflammation upon infection clearance. PMID:24549323

  12. Multicopper oxidase-1 orthologs from diverse insect species have ascorbate oxidase activity

    PubMed Central

    Peng, Zeyu; Dittmer, Neal T.; Lang, Minglin; Brummett, Lisa M.; Braun, Caroline L.; Davis, Lawrence C.; Kanost, Michael R.; Gorman, Maureen J.

    2015-01-01

    Members of the multicopper oxidase (MCO) family of enzymes can be classified by their substrate specificity; for example, ferroxidases oxidize ferrous iron, ascorbate oxidases oxidize ascorbate, and laccases oxidize aromatic substrates such as diphenols. Our previous work on an insect multicopper oxidase, MCO1, suggested that it may function as a ferroxidase. This hypothesis was based on three lines of evidence: RNAi-mediated knock down of Drosophila melanogaster MCO1 (DmMCO1) affects iron homeostasis, DmMCO1 has ferroxidase activity, and DmMCO1 has predicted iron binding residues. In our current study, we expanded our focus to include MCO1 from Anopheles gambiae, Tribolium castaneum, and Manduca sexta. We verified that MCO1 orthologs have similar expression profiles, and that the MCO1 protein is located on the basal surface of cells where it is positioned to oxidize substrates in the hemolymph. In addition, we determined that RNAi-mediated knock down of MCO1 in A. gambiae affects iron homeostasis. To further characterize the enzymatic activity of MCO1 orthologs, we purified recombinant MCO1 from all four insect species and performed kinetic analyses using ferrous iron, ascorbate and two diphenols as substrates. We found that all of the MCO1 orthologs are much better at oxidizing ascorbate than they are at oxidizing ferrous iron or diphenols. This result is surpring because ascorbate oxidases are thought to be specific to plants and fungi. An analysis of three predicted iron binding residues in DmMCO1 revealed that they are not required for ferroxidase or laccase activity, but two of the residues (His374 and Asp380) influence oxidation of ascorbate. These two residues are conserved in MCO1 orthologs from insects and crustaceans; therefore, they are likely to be important for MCO1 function. The results of this study suggest that MCO1 orthologs function as ascorbate oxidases and influence iron homeostasis through an unknown mechanism. PMID:25701385

  13. Thymoquinone, an active constituent of Nigella sativa seeds, binds with bilirubin and protects mice from hyperbilirubinemia and cyclophosphamide-induced hepatotoxicity.

    PubMed

    Laskar, Amaj A; Khan, Masood A; Rahmani, Arshad H; Fatima, Sana; Younus, Hina

    2016-08-01

    Some reports indicate that thymoquinone (TQ), the main constituent of Nigella sativa seeds, is hepatoprotective. The aim of this study was to determine whether TQ is able to bind directly to bilirubin, and whether TQ or liposomal formulation of TQ (Lip-TQ) can reduce cyclophosphamide (CYP)-induced liver toxicity, serum bilirubin level in mice. The binding of TQ with bilirubin was studied by UV-VIS, fluorescence and Near-UV CD spectroscopy. Inhibition of binding of bilirubin to erythrocytes by TQ was also examined. To increase the in vivo efficacy, Lip-TQ was prepared and used against CYP-induced toxicity. The protective role of TQ or Lip-TQ against CYP-induced toxicity was assessed by determining the liver function parameters, the levels of superoxide dismutase (SOD) and catalase (CAT), and histological studies. It was found that TQ binds to bilirubin and significantly inhibits the binding of bilirubin to erythrocytes. Lip-TQ (10 mg/kg) significantly reduced the levels of aspartate transaminase (AST) from 254 ± 48 to 66 ± 18 IU/L (P < 0.001), alanine transaminase (ALT) from 142 ± 28 to 47.8 ± 16 IU/L (P < 0.05) and serum bilirubin from 2.8 ± 0.50 to 1.24 ± 0.30 mg/dl (P < 0.05). Treatment with Lip-TQ reduced the CYP-induced inflammation and hemorrhage in liver tissues. Moreover, treatment with free or Lip-TQ protected the activity of SOD and CAT in CYP-injected mice. Therefore, TQ can reduce the level of bilirubin in systemic circulation in disease conditions that lead to hyperbilirubinemia and liver toxicity and hence may be used as a supplement in the treatment of liver ailments. PMID:27265787

  14. Effect of contraceptive steroids on monoamine oxidase activity

    PubMed Central

    Southgate, Jennifer; Collins, G. G. S.; Pryse-Davies, J.; Sandler, M.

    1969-01-01

    Cyclical variations in monoamine oxidase activity during the human menstrual cycle, specific to the endometrium and modified in women undergoing contraceptive steroid treatment, may reflect changes in hormonal environment. Treatment of rats with individual constituents of the contraceptive pill causes analogous changes: oestrogens inhibit and progestogens potentiate uterine monoamine oxidase activity. ImagesFig. 2Fig. 3

  15. Xanthine oxidase inhibitors from Garcinia esculenta twigs.

    PubMed

    Zhu, Lun-Lun; Fu, Wen-Wei; Watanabe, Shimpei; Shao, Yi-Nuo; Tan, Hong-Sheng; Zhang, Hong; Tan, Chang-Heng; Xiu, Yan-Feng; Norimoto, Hisayoshi; Xu, Hong-Xi

    2014-12-01

    The EtOAc-soluble portion of the 80 % (v/v) EtOH extract from the twigs of Garcinia esculenta exhibited strong xanthine oxidase inhibition in vitro. Bioassay-guided purification led to the isolation of 1,3,6,7-tetrahydroxyxanthone (3) and griffipavixanthone (8) as the main xanthine oxidase inhibitors, along with six additional compounds (1, 2, 4-7), including two new compounds (1 and 2). This enzyme inhibition was dose dependent with an IC50 value of approximately 1.2 µM for 3 and 6.3 µM for 8. The inhibitory activity of 3 was stronger than the control allopurinol (IC50 value: 5.3 µM). To our knowledge, compound 8 is the first bixanthone that demonstrated potent XO inhibitory activity in vitro. The structures of the new compounds were established by spectroscopic analysis, and the optical properties and absolute stereochemistry of racemic (±) esculentin A (2) were further determined by the calculation of the DP4 probability and analysis of its MTPA ester derivatives. PMID:25340468

  16. Activation of antibacterial autophagy by NADPH oxidases

    PubMed Central

    Huang, Ju; Canadien, Veronica; Lam, Grace Y.; Steinberg, Benjamin E.; Dinauer, Mary C.; Magalhaes, Marco A. O.; Glogauer, Michael; Grinstein, Sergio; Brumell, John H.

    2009-01-01

    Autophagy plays an important role in immunity to microbial pathogens. The autophagy system can target bacteria in phagosomes, promoting phagosome maturation and preventing pathogen escape into the cytosol. Recently, Toll-like receptor (TLR) signaling from phagosomes was found to initiate their targeting by the autophagy system, but the mechanism by which TLR signaling activates autophagy is unclear. Here we show that autophagy targeting of phagosomes is not exclusive to those containing TLR ligands. Engagement of either TLRs or the Fcγ receptors (FcγRs) during phagocytosis induced recruitment of the autophagy protein LC3 to phagosomes with similar kinetics. Both receptors are known to activate the NOX2 NADPH oxidase, which plays a central role in microbial killing by phagocytes through the generation of reactive oxygen species (ROS). We found that NOX2-generated ROS are necessary for LC3 recruitment to phagosomes. Antibacterial autophagy in human epithelial cells, which do not express NOX2, was also dependent on ROS generation. These data reveal a coupling of oxidative and nonoxidative killing activities of the NOX2 NADPH oxidase in phagocytes through autophagy. Furthermore, our results suggest a general role for members of the NOX family in regulating autophagy. PMID:19339495

  17. Northwest Outward Bound Instructor's Manual.

    ERIC Educational Resources Information Center

    Northwest Outward Bound School, Portland, OR.

    Instructor responsibilities, procedures for completing activities safely, and instructional methods and techniques are outlined to assist instructors in the Northwest Outward Bound School (Portland, Oregon) as they strive for teaching excellence. Information is organized into six chapters addressing: history and philosophy of Outward Bound; course…

  18. Fractional diffusion on bounded domains

    DOE PAGESBeta

    Defterli, Ozlem; D'Elia, Marta; Du, Qiang; Gunzburger, Max Donald; Lehoucq, Richard B.; Meerschaert, Mark M.

    2015-03-13

    We found that the mathematically correct specification of a fractional differential equation on a bounded domain requires specification of appropriate boundary conditions, or their fractional analogue. In this paper we discuss the application of nonlocal diffusion theory to specify well-posed fractional diffusion equations on bounded domains.

  19. Bounds for Asian basket options

    NASA Astrophysics Data System (ADS)

    Deelstra, Griselda; Diallo, Ibrahima; Vanmaele, Michèle

    2008-09-01

    In this paper we propose pricing bounds for European-style discrete arithmetic Asian basket options in a Black and Scholes framework. We start from methods used for basket options and Asian options. First, we use the general approach for deriving upper and lower bounds for stop-loss premia of sums of non-independent random variables as in Kaas et al. [Upper and lower bounds for sums of random variables, Insurance Math. Econom. 27 (2000) 151-168] or Dhaene et al. [The concept of comonotonicity in actuarial science and finance: theory, Insurance Math. Econom. 31(1) (2002) 3-33]. We generalize the methods in Deelstra et al. [Pricing of arithmetic basket options by conditioning, Insurance Math. Econom. 34 (2004) 55-57] and Vanmaele et al. [Bounds for the price of discrete sampled arithmetic Asian options, J. Comput. Appl. Math. 185(1) (2006) 51-90]. Afterwards we show how to derive an analytical closed-form expression for a lower bound in the non-comonotonic case. Finally, we derive upper bounds for Asian basket options by applying techniques as in Thompson [Fast narrow bounds on the value of Asian options, Working Paper, University of Cambridge, 1999] and Lord [Partially exact and bounded approximations for arithmetic Asian options, J. Comput. Finance 10 (2) (2006) 1-52]. Numerical results are included and on the basis of our numerical tests, we explain which method we recommend depending on moneyness and time-to-maturity.

  20. The complex roles of NADPH oxidases in fungal infection

    PubMed Central

    Hogan, Deborah; Wheeler, Robert T.

    2014-01-01

    Summary NADPH oxidases play key roles in immunity and inflammation that go beyond the production of microbicidal reactive oxygen species (ROS). The past decade has brought a new appreciation for the diversity of roles played by ROS in signaling associated with inflammation and immunity. NADPH oxidase activity affects disease outcome during infections by human pathogenic fungi, an important group of emerging and opportunistic pathogens that includes Candida, Aspergillus and Cryptococcus species. Here we review how alternative roles of NADPH oxidase activity impact fungal infection and how ROS signaling affects fungal physiology. Particular attention is paid to roles for NADPH oxidase in immune migration, immunoregulation in pulmonary infection, neutrophil extracellular trap formation, autophagy and inflammasome activity. These recent advances highlight the power and versatility of spatiotemporally controlled redox regulation in the context of infection, and point to a need to understand the molecular consequences of NADPH oxidase activity in the cell. PMID:24905433

  1. Ascorbic acid and L-gulonolactone oxidase in lagomorphs.

    PubMed

    Jenness, R; Birney, E C; Ayaz, K L

    1978-01-01

    1. The activity of L-gulonolactone oxidase (EC 1.1.3.8) in the liver of eastern cottontail rabbits (Sylvilagus floridanus) is about 10-fold greater in winter than in summer. 2. L-gulonolactone oxidase activity is low and tissue ascorbate high during all seasons in snowshoe hares (Lepus americanus). 3. Liver contents of ascorbate fall to low levels in L. americanus fed on rabbit chow in the laboratory. 4. The activity of L-gulonolactone oxidase in liver of Sylvilagus and Oryctolagus is depressed by feeding high levels of L-ascorbic acid. 5. The New Zealand White breed of domestic rabbit (Oryctolagus cuniculus) has considerably higher levels of L-gulonolactone oxidase and liver ascorbate than does the Dutch breed. 6. In a wild population of Oryctolagus sampled in Australia L-gulonolactone oxidase levels were intermediate between those of the two domestic breeds and more variable than either. PMID:318384

  2. Respiratory conservation of energy with dioxygen: cytochrome C oxidase.

    PubMed

    Yoshikawa, Shinya; Shimada, Atsuhiro; Shinzawa-Itoh, Kyoko

    2015-01-01

    Cytochrome c oxidase (CcO) is the terminal oxidase of cell respiration which reduces molecular oxygen (O₂) to H2O coupled with the proton pump. For elucidation of the mechanism of CcO, the three-dimensional location and chemical reactivity of each atom composing the functional sites have been extensively studied by various techniques, such as crystallography, vibrational and time-resolved electronic spectroscopy, since the X-ray structures (2.8 Å resolution) of bovine and bacterial CcO have been published in 1995.X-ray structures of bovine CcO in different oxidation and ligand binding states showed that the O₂reduction site, which is composed of Fe (heme a 3) and Cu (CuB), drives a non-sequential four-electron transfer for reduction of O₂to water without releasing any reactive oxygen species. These data provide the crucial structural basis to solve a long-standing problem, the mechanism of the O₂reduction.Time-resolved resonance Raman and charge translocation analyses revealed the mechanism for coupling between O₂reduction and the proton pump: O₂is received by the O₂reduction site where both metals are in the reduced state (R-intermediate), giving the O₂-bound form (A-intermediate). This is spontaneously converted to the P-intermediate, with the bound O₂fully reduced to 2 O²⁻. Hereafter the P-intermediate receives four electron equivalents from the second Fe site (heme a), one at a time, to form the three intermediates, F, O, and E to regenerate the R-intermediate. Each electron transfer step from heme a to the O₂reduction site is coupled with the proton pump.X-ray structural and mutational analyses of bovine CcO show three possible proton transfer pathways which can transfer pump protons (H) and chemical (water-forming) protons (K and D). The structure of the H-pathway of bovine CcO indicates that the driving force of the proton pump is the electrostatic repulsion between the protons on the H-pathway and positive charges of heme a, created

  3. Evaluation and comparison of postoperative levels of serum bilirubin, serum transaminases and alkaline phosphatase in laparoscopic cholecystectomy versus open cholecystectomy

    PubMed Central

    Singal, Rajinder Pal; Sandhu, Karamjot; Singh, Bir; Bhatia, Gaurav; Khatri, Abhishek; Sharma, Bhanu Pratap

    2015-01-01

    Background Laparoscopic cholecystectomy (LC) requires the creation of a pneumoperitoneum via insufflations of carbon dioxide; resulting in increased partial pressure of carbon dioxide (CO2) and intraperitoneal pressure which leads to the changes in pulmonary function and hemodynamic measurements. Hypercarbia leads to visceral organ ischemia including liver and venous stasis/thromboembolism or both due to impaired flow. The present study has been undertaken to see the changes in liver function tests (LFTs) after laparoscopic/open cholecystectomy (OC), the incidences of such change, their relation to age, sex, duration of surgery and to know the clinical significances of such disturbances. Aims and objectives To compare and correlate the serum level of bilirubin, alanine transaminase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP) in patients who underwent LC to those who underwent OC. Materials and methods The present study was conducted in the Department of Surgery at MMIMSR, MM University, Mullana, Ambala. A total number of 200 patients diagnosed as cholelithiasis were included in the study from May 2012 to May 2014. These cases were randomly divided into two groups (A and B) consisting of 100 cases each. LC was performed in group A patients and OC was done in group B patients. Three blood samples were taken: (I) pre-operatively; (II) after 24 hours of surgery; and (III) after 72 hours of surgery for comparison of the enzyme level alterations. Results In LC patients, there were rise in the levels of serum bilirubin, AST and ALT after 24 hrs of surgery from the preoperative value and then again fall was noted (near to normal value) after 72 hrs of surgery except in that of ALP. ALP levels showed slight fall after 24 hrs of surgery and then slight rise after 72 hrs which was within the normal limit. Whereas in OC patients, there were slight variations in the liver enzymes (which were within the normal range). Conclusions Transient elevation of serum

  4. Computer simulation of water in cytochrome c oxidase.

    PubMed

    Zheng, Xuehe; Medvedev, Dmitry M; Swanson, Jessica; Stuchebrukhov, Alexei A

    2003-03-01

    Statistical mechanics and molecular dynamics simulations have been carried out to study the distribution and dynamics of internal water molecules in bovine heart cytochrome c oxidase (CcO). CcO is found to be capable of holding plenty of water, which in subunit I alone amounts to about 165 molecules. The dynamic characterization of these water molecules is carried out. The nascent water molecules produced in the redox reaction at the heme a(3)-CuB binuclear site form an intriguing chain structure. The chain begins at the position of Glu242 at the end of the D channel, and has a fork structure, one branch of which leads to the binuclear center, and the other to the propionate d of heme a(3). The branch that leads to the binuclear center has dynamic access both to the site where the formation of water occurs, and to delta-nitrogen of His291. From the binuclear center, the chain continues to run into the K channel. The stability of this hydrogen bond network is examined dynamically. The catalytic site is located at the hydrophobic region, and the nascent water molecules are produced at the top of the energy hill. The energy gradient is utilized as the mechanism of water removal from the protein. The water exit channels are explored using high-temperature dynamics simulations. Two putative channels for water exit from the catalytic site have been identified. One is leading directly toward Mg(2+) site. However, this channel is only open when His291 is dissociated from CuB. If His291 is bound to CuB, the only channel for water exit is the one that originates at E242 and leads toward the middle of the membrane. This is the same channel that is presumably used for oxygen supply. PMID:12615353

  5. Mitochondrial Copper Metabolism and Delivery to Cytochrome c Oxidase

    PubMed Central

    Horn, Darryl; Barrientos, Antoni

    2010-01-01

    Summary Metals are essential elements of all living organisms. Among them, copper is required for a multiplicity of functions including mitochondrial oxidative phosphorylation and protection against oxidative stress. Here we will focus on describing the pathways involved in the delivery of copper to cytochrome c oxidase (COX), a mitochondrial metalloenzyme acting as the terminal enzyme of the mitochondrial respiratory chain. The catalytic core of COX is formed by three mitochondrially-encoded subunits and contains three copper atoms. Two copper atoms bound to subunit 2 constitute the CuA site, the primary acceptor of electrons from ferrocytochrome c. The third copper, CuB, is associated with the high-spin heme a3 group of subunit 1. Recent studies, mostly performed in the yeast Saccharomyces cerevisiae, have provided new clues about 1- the source of the copper used for COX metallation; 2- the roles of Sco1p and Cox11p, the proteins involved in the direct delivery of copper to the CuA and CuB sites, respectively; 3- the action mechanism of Cox17p, a copper chaperone that provides copper to Sco1p and Cox11p; 4- the existence of at least four Cox17p homologues carrying a similar twin CX9C domain suggestive of metal binding, Cox19p, Cox23p, Pet191p and Cmc1p, that could be part of the same pathway; and 5- the presence of a disulfide relay system in the intermembrane space of mitochondria that mediates import of proteins with conserved cysteines motifs such as the CX9C characteristic of Cox17p and its homologues. The different pathways are reviewed and discussed in the context of both mitochondrial COX assembly and copper homeostasis. PMID:18459161

  6. Gene structure and quinol oxidase activity of a cytochrome bd-type oxidase from Bacillus stearothermophilus.

    PubMed

    Sakamoto, J; Koga, E; Mizuta, T; Sato, C; Noguchi, S; Sone, N

    1999-04-21

    Gram-positive thermophilic Bacillus species contain cytochrome caa3-type cytochrome c oxidase as their main terminal oxidase in the respiratory chain. We previously identified and purified an alternative oxidase, cytochrome bd-type quinol oxidase, from a mutant of Bacillus stearothermophilus defective in the caa3-type oxidase activity (J. Sakamoto et al., FEMS Microbiol. Lett. 143 (1996) 151-158). Compared with proteobacterial counterparts, B. stearothermophilus cytochrome bd showed lower molecular weights of the two subunits, shorter wavelength of alpha-band absorption maximum due to heme D, and lower quinol oxidase activity. Preincubation with menaquinone-2 enhanced the enzyme activity up to 40 times, suggesting that, besides the catalytic site, there is another quinone-binding site which largely affects the enzyme activity. In order to clarify the molecular basis of the differences of cytochromes bd between B. stearothermophilus and proteobacteria, the genes encoding for the B. stearothermophilus bd was cloned based on its partial peptide sequences. The gene for subunit I (cbdA) encodes 448 amino acid residues with a molecular weight of 50195 Da, which is 14 and 17% shorter than those of Escherichia coli and Azotobacter vinelandii, respectively, and CbdA lacks the C-terminal half of the long hydrophilic loop between the putative transmembrane segments V and VI (Q loop), which has been suggested to include the substrate quinone-binding site for the E. coli enzyme. The gene for subunit II (cbdB) encodes 342 residues with a molecular weight of 38992 Da. Homology search indicated that the B. stearothermophilus cbdAB has the highest sequence similarity to ythAB in B. subtilis genome rather than to cydAB, the first set of cytochrome bd genes identified in the genome. Sequence comparison of cytochromes bd and their homologs from various organisms demonstrates that the proteins can be classified into two subfamilies, a proteobacterial type including E. coli bd and a

  7. Saturating the holographic entropy bound

    SciTech Connect

    Bousso, Raphael; Freivogel, Ben; Leichenauer, Stefan

    2010-10-15

    The covariant entropy bound states that the entropy, S, of matter on a light sheet cannot exceed a quarter of its initial area, A, in Planck units. The gravitational entropy of black holes saturates this inequality. The entropy of matter systems, however, falls short of saturating the bound in known examples. This puzzling gap has led to speculation that a much stronger bound, S < or approx. A{sup 3/4}, may hold true. In this note, we exhibit light sheets whose entropy exceeds A{sup 3/4} by arbitrarily large factors. In open Friedmann-Robertson-Walker universes, such light sheets contain the entropy visible in the sky; in the limit of early curvature domination, the covariant bound can be saturated but not violated. As a corollary, we find that the maximum observable matter and radiation entropy in universes with positive (negative) cosmological constant is of order {Lambda}{sup -1} ({Lambda}{sup -2}), and not |{Lambda}|{sup -3/4} as had hitherto been believed. Our results strengthen the evidence for the covariant entropy bound, while showing that the stronger bound S < or approx. A{sup 3/4} is not universally valid. We conjecture that the stronger bound does hold for static, weakly gravitating systems.

  8. Polymer resins with amino acid containing pendants for sorption of bilirubin. II. Polyamide resins with various basic amino acids.

    PubMed

    Henning, D S; Brown, G R; St-Pierre, L E

    1986-01-01

    Short peptides, three to eight amino acids in length, containing various combinations of alanine, arginine, lysine, histidine and tyrosine have been synthesized onto water-swellable polyamide resin by the solid phase peptide synthesis method. The amount of bilirubin adsorbed from aqueous buffer solution (pH = 7.8) by the resins increases with increasing basicity of the amino acids in the pendant. As the number of basic amino acids on the pendant is increased from one to five a 4.7 fold enhancement in the adsorption capacity is seen for arginine while a 9.3 fold enhancement is obtained for lysine. A corresponding increase in length for the non-basic histidine results in a 6 fold enhancement. With alanine the adsorption capacity is uneffected by an increase in pendant length. PMID:3957453

  9. Correlation Between C-reactive Protein and Non-enzymatic Antioxidants (Albumin, Ferritin, Uric Acid and Bilirubin) in Hemodialysis Patients

    PubMed Central

    Beciragic, Amela; Resic, Halima; Prohic, Nejra; Karamehic, Jasenko; Smajlovic, Ajdin; Masnic, Fahrudin; Ajanovic, Selma; Coric, Aida

    2015-01-01

    Introduction: Increased levels of C-Reactive Protein are found in 30-60% on hemodialysis patients and it is closely associated with the progression of atherosclerosis, cardiovascular morbidity and mortality. Non enzymatic antioxidants are antioxidants which primarily retain potentially dangerous ions of iron and copper in their inactive form and thereby prevent its participation in the production of free radicals. Aim: The aim of the study was to examine the relationship of CRP and non enzymatic antioxidants (albumin, ferritin, uric acid and bilirubin) i.e. examine the importance of CRP as a serum biomarker in assessing the condition of inflammation and its relationship to antioxidant protection in patients on hemodialysis. Methods: The study was cross-sectional, clinical, comparative and descriptive. The study involved 100 patients (non diabetic) on chronic hemodialysis. The control group consisted of 50 subjects without subjective and objective indicators of chronic renal disease. In all patients, the concentration of CRP as well as concentrations of non enzymatic antioxidants were determined. Results: In the group of hemodialysis patients 60% were men and 40% women. The average age of hemodialysis patients was 54.13 ± 11.8 years and the average age of the control group 41.72 ± 9.8 years. The average duration of hemodialysis treatment was 91.42 ± 76.2 months. In the group of hemodialysis patients statistically significant, negative linear correlation was determined between the concentration of CRP in and albumin concentration (rho = -0.251, p = 0.012) as well as negative, statistics insignificant, linear correlation between serum CRP and the concentration of uric acid (r = -0.077, p = 0.448). Furthermore, the positive, linear correlation was determined between serum CRP and ferritin (r = 0.159, p = 0.114) and positive linear correlation between CRP and total serum bilirubin (r = 0.121, p = 0.230). In the control group was determined a statistically significant

  10. Expression of the alternative oxidase complements cytochrome c oxidase deficiency in human cells

    PubMed Central

    Dassa, Emmanuel P; Dufour, Eric; Gonçalves, Sérgio; Paupe, Vincent; Hakkaart, Gertjan A J; Jacobs, Howard T; Rustin, Pierre

    2009-01-01

    Cytochrome c oxidase (COX) deficiency is associated with a wide spectrum of clinical conditions, ranging from early onset devastating encephalomyopathy and cardiomyopathy, to neurological diseases in adulthood and in the elderly. No method of compensating successfully for COX deficiency has been reported so far. In vitro, COX-deficient human cells require additional glucose, pyruvate and uridine for normal growth and are specifically sensitive to oxidative stress. Here, we have tested whether the expression of a mitochondrially targeted, cyanide-resistant, alternative oxidase (AOX) from Ciona intestinalis could alleviate the metabolic abnormalities of COX-deficient human cells either from a patient harbouring a COX15 pathological mutation or rendered deficient by silencing the COX10 gene using shRNA. We demonstrate that the expression of the AOX, well-tolerated by the cells, compensates for both the growth defect and the pronounced oxidant-sensitivity of COX-deficient human cells. PMID:20049701

  11. Calculated Proton Uptake on Anaerobic Reduction of Cytochrome c Oxidase: Is the Reaction Electroneutral?†

    PubMed Central

    Song, Yifan; Michonova-Alexova, Ekaterina; Gunner, M. R.

    2009-01-01

    Cytochrome c oxidase is a transmembrane proton pump that builds an electrochemical gradient using chemical energy from the reduction of O2. Ionization states of all residues were calculated with Multi-Conformation Continuum Electrostatics (MCCE) in seven anaerobic oxidase redox states ranging from fully oxidized to fully reduced. One long-standing problem is how proton uptake is coupled to the reduction of the active site binuclear center (BNC). The BNC has two cofactors: heme a3 and CuB If the protein needs to maintain electroneutrality, then 2 protons will be bound when the BNC is reduced by 2 electrons in the reductive half of the reaction cycle. The effective pKas of ionizable residues around the BNC are evaluated in Rhodobacter sphaeroides cytochrome c oxidase. At pH 7, only a hydroxide coordinated to CuB shifts its pKa from below 7 to above 7 and so picks up a proton when heme a3 and CuB are reduced. Glu I-286, Tyr I-288, His I-334, and a second hydroxide on heme a3 all have pKas above 7 in all redox states, although they have only 1.6–3.5 ΔpK units energy cost for deprotonation. Thus, at equilibrium, they are protonated and cannot serve as proton acceptors. The propionic acids near the BNC are deprotonated with pKas well below 7. They are well stabilized in their anionic state and do not bind a proton upon BNC reduction. This suggests that electroneutrality in the BNC is not maintained during the anaerobic reduction. Proton uptake on reduction of CuA, heme a, heme a3, and CuB shows ≈2.5 protons bound per 4 electrons, in agreement with prior experiments. One proton is bound by a hydroxyl group in the BNC and the rest to groups far from the BNC. The electrochemical midpoint potential (Em) of heme a is calculated in the fully oxidized protein and with 1 or 2 electrons in the BNC. The Em of heme a shifts down when the BNC is reduced, which agrees with prior experiments. If the BNC reduction is electroneutral, then the heme a Em is independent of the BNC

  12. High Total Bilirubin as a Protective Factor for Diabetes Mellitus: An Analysis of NHANES Data From 1999 - 2006

    PubMed Central

    Cheriyath, Pramil; Gorrepati, Venkata Subhash; Peters, Ian; Nookala, Vinod; Murphy, Megan E; Srouji, Nadine; Fischman, Daniel

    2010-01-01

    Background Diabetes Mellitus (DM) is a rampantly growing epidemic in the United States, affecting nearly 10% of the adult population. Studies have shown that higher levels of Total Bilirubin (TBili) convey a protective effect with regard to cardiovascular risk. In this study, we will examine the relationship between TBili level and prevalence of DM to discern whether a similar relationship exists. Methods The National Health and Nutrition Examination Survey (NHANES) is a comprehensive survey performed regularly to evaluate the overall health and nutrition status of the United States population. For the purpose of this study, we combined NHANES data collected between 1999 and 2006. Totally 15,876 eligible participants were selected after excluding all patients younger than twenty years, those with a history of abnormal liver function tests, or those who disclosed a history of liver disease. The data collected on these individuals was adjusted for demographic characteristics, as well as risk factors for DM, and was analyzed via multivariate logistic regression, using SAS proc survey methodology. Results After age adjustment, increased TBili was associated with 26% reduction in diabetes risk (OR 0.74, 95% CI 0.64 - 0.88). Multivariate analysis, adjusting for all diabetes risk factors assessed, confirmed this association (OR 0.80, 95% CI 0.67 - 0.95). Conclusions Our results show that a higher level of serum TBili is associated with odds of having a lower incidence of DM. This finding supports the hypothesis that the antioxidant nature of TBili, demonstrating a protective effect with regard to the risk of stroke, atherosclerosis, and vasculitis in prior research, also extends to DM risk. Furthermore, research has shown that higher levels of TBili increase glucose mobilization into the cells, leading to more efficient, biologic glucose utilization. There is no doubt that the beneficial effect of TBili is multifactorial; thus further investigation is warranted. Keywords

  13. Higher serum total bilirubin concentration is associated with lower risk of renal insufficiency in an adult population

    PubMed Central

    Lee, Ang-Tse; Wang, Ya-Yu; Lin, Shih-Yi; Liang, Jiin-Tsae; Sheu, Wayne Huey-Herng; Song, Yuh-Min; Chang, Wen-Dau

    2015-01-01

    Background: Chronic inflammation is proposed to play a central role in the pathogenesis of chronic kidney disease (CKD), and serum bilirubin has antioxidant and anti-inflammatory effects. We investigated the association between serum total bilirubin (Tb) concentration and renal function in an adult population. Methods: We conducted a cross-sectional study and collected anthropometric measurements, fasting blood tests, lifestyle habits and medical history of 3876 subjects attending a health examination. Renal insufficiency was defined as estimated glomerular filtration rate (eGFR) <60 mL/min/1.73 m2 calculated by using the CKD-EPI equation. Results: Serum Tb concentrations were higher in subjects without renal insufficiency than in those with renal insufficiency. Multivariable linear regression analysis showed that Tb concentration was positively associated with eGFR after adjusting for important CKD risk factors (P=0.04). Multivariable logistic regression analysis also revealed that higher Tb concentration (each increment of 1.71 μmol/L) (0.1 mg/dL) was associated with a reduced risk of renal insufficiency: odds ratios were 0.94 (P=0.005) for men and 0.90 (P=0.015) for women, respectively. When subjects were divided into quartiles of serum Tb, multivariable-adjusted odds ratios for renal insufficiency comparing the fourth to the first Tb quartile were 0.49 (P=0.001) for men and 0.35 (P=0.003) for women. A stepwise exclusion of subjects, first those with possible liver disease and second, those with CKD stage 4 and 5, showed consistent results. Conclusion: Higher serum Tb concentration was associated with lower risk of renal insufficiency, regardless of other conventional CKD risk factors. PMID:26770557

  14. Higher plasma bilirubin predicts veno-occlusive disease in early childhood undergoing hematopoietic stem cell transplantation with cyclosporine

    PubMed Central

    Kim, Kwi Suk; Moon, Aree; Kang, Hyoung Jin; Shin, Hee Young; Choi, Young Hee; Kim, Hyang Sook; Kim, Sang Geon

    2016-01-01

    AIM: To analyze the association between plasma bilirubin levels and veno-occlusive disease (VOD) in non-adult patients undergoing hematopoietic stem cell transplantation (HSCT) during cyclosporine therapy. METHODS: A total of 123 patients taking cyclosporine were evaluated using an electronic medical system at the Seoul National University Children’s Hospital from the years 2004 through 2011. Patients were grouped by age and analyzed for incidence and type of adverse drug reactions (ADRs) including VOD. RESULTS: The HSCT patients were divided into three age groups: G#1 ≥ 18; 9 ≤ G#2 ≤ 17; and G#3 ≤ 8 years of age). The majority of transplant donor types were cord blood transplantations. Most prevalent ADRs represented acute graft-vs-host disease (aGVHD) and VOD. Although the incidences of aGVHD did not vary among the groups, the higher frequency ratios of VOD in G#3 suggested that an age of 8 or younger is a risk factor for developing VOD in HSCT patients. After cyclosporine therapy, the trough plasma concentrations of cyclosporine were lower in G#3 than in G#1, indicative of its increased clearance. Moreover, in G#3 only, a maximal total bilirubin level (BILmax) of ≥ 1.4 mg/dL correlated with VOD incidence after cyclosporine therapy. CONCLUSION: HSCT patients 8 years of age or younger are more at risk for developing VOD, diagnosed as hyperbilirubinemia, tender hepatomegaly, and ascites/weight gain after cyclosporine therapy, which may be represented by a criterion of plasma BILmax being ≥ 1.4 mg/dL, suggestive of more sensitive VOD indication in this age group. PMID:27358786

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

    PubMed

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

    2016-01-01

    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. PMID:27409598

  16. Endogenously elevated bilirubin modulates kidney function and protects from circulating oxidative stress in a rat model of adenine-induced kidney failure

    PubMed Central

    Boon, Ai-Ching; Lam, Alfred K.; Gopalan, Vinod; Benzie, Iris F.; Briskey, David; Coombes, Jeff S.; Fassett, Robert G.; Bulmer, Andrew C.

    2015-01-01

    Mildly elevated bilirubin is associated with a reduction in the presence and progression of chronic kidney disease and related mortality, which may be attributed to bilirubin’s antioxidant properties. This study investigated whether endogenously elevated bilirubin would protect against adenine-induced kidney damage in male hyperbilirubinaemic Gunn rats and littermate controls. Animals were orally administered adenine or methylcellulose solvent (vehicle) daily for 10 days and were then monitored for 28 days. Serum and urine were assessed throughout the protocol for parameters of kidney function and antioxidant/oxidative stress status and kidneys were harvested for histological examination upon completion of the study. Adenine-treated animals experienced weight-loss, polyuria and polydipsia; however, these effects were significantly attenuated in adenine-treated Gunn rats. No difference in the presence of dihydroadenine crystals, lymphocytic infiltration and fibrosis were noted in Gunn rat kidneys versus controls. However, plasma protein carbonyl and F2-isoprostane concentrations were significantly decreased in Gunn rats versus controls, with no change in urinary 8-oxo-7,8-dihydro-2′-deoxyguanosine or kidney tissue F2-isoprostane concentrations. These data indicated that endogenously elevated bilirubin specifically protects from systemic oxidative stress in the vascular compartment. These data may help to clarify the protective relationship between bilirubin, kidney function and cardiovascular mortality in clinical investigations. PMID:26498893

  17. NADPH Oxidases in Chronic Liver Diseases

    PubMed Central

    Jiang, Joy X.; Török, Natalie J.

    2015-01-01

    Oxidative stress is a common feature observed in a wide spectrum of chronic liver diseases including viral hepatitis, alcoholic, and nonalcoholic steatohepatitis. The nicotinamide adenine dinucleotide phosphate (NADPH) oxidases (NOXs) are emerging as major sources of reactive oxygen species (ROS). Several major isoforms are expressed in the liver, including NOX1, NOX2, and NOX4. While the phagocytic NOX2 has been known to play an important role in Kupffer cell and neutrophil phagocytic activity and inflammation, the nonphagocytic NOX homologues are increasingly recognized as key enzymes in oxidative injury and wound healing. In this review, we will summarize the current advances in knowledge on the regulatory pathways of NOX activation, their cellular distribution, and their role in the modulation of redox signaling in liver diseases. PMID:26436133

  18. Degradation of pentachlorophenol by potato polyphenol oxidase.

    PubMed

    Hou, Mei-Fang; Tang, Xiao-Yan; Zhang, Wei-De; Liao, Lin; Wan, Hong-Fu

    2011-11-01

    In this study, polyphenol oxidase (PPO) was extracted from commercial potatoes. Degradation of pentachlorophenol by potato PPO was investigated. The experimental results show that potato PPO is more active in weak acid than in basic condition and that the optimum pH for the reaction is 5.0. The degradation of pentachlorophenol by potato PPO reaches a maximum at 298 K. After reaction for 1 h, the removal of both pentachlorophenol and total organic carbon is >70% with 6.0 units/mL potato PPO at pH 5.0 and 298 K. Pentachlorophenol can be degraded through dechlorination and ring-opening by potato PPO. The work demonstrates that pentachlorophenol can be effectively eliminated by crude potato PPO. PMID:21967325

  19. Visualization of monoamine oxidase in human brain

    SciTech Connect

    Fowler, J.S.; Volkow, N.D.; Wang, G.J.; Pappas, N.; Shea, C.; MacGregor, R.R.; Logan, J.

    1996-12-31

    Monoamine oxidase is a flavin enzyme which exists in two subtypes, MAO A and MAO B. In human brain MAO B predominates and is largely compartmentalized in cell bodies of serotonergic neurons and glia. Regional distribution of MAO B was determined by positron computed tomography with volunteers after the administration of deuterium substituted [11C]L-deprenyl. The basal ganglia and thalamus exhibited the greatest concentrations of MAO B with intermediate levels in the frontal cortex and cingulate gyrus while lowest levels were observed in the parietal and temporal cortices and cerebellum. We observed that brain MAO B increases with are in health normal subjects, however the increases were generally smaller than those revealed with post-mortem studies.

  20. Drugs related to monoamine oxidase activity.

    PubMed

    Fišar, Zdeněk

    2016-08-01

    Progress in understanding the role of monoamine neurotransmission in pathophysiology of neuropsychiatric disorders was made after the discovery of the mechanisms of action of psychoactive drugs, including monoamine oxidase (MAO) inhibitors. The increase in monoamine neurotransmitter availability, decrease in hydrogen peroxide production, and neuroprotective effects evoked by MAO inhibitors represent an important approach in the development of new drugs for the treatment of mental disorders and neurodegenerative diseases. New drugs are synthesized by acting as multitarget-directed ligands, with MAO, acetylcholinesterase, and iron chelation as targets. Basic information is summarized in this paper about the drug-induced regulation of monoaminergic systems in the brain, with a focus on MAO inhibition. Desirable effects of MAO inhibition include increased availability of monoamine neurotransmitters, decreased oxidative stress, decreased formation of neurotoxins, induction of pro-survival genes and antiapoptotic factors, and improved mitochondrial functions. PMID:26944656

  1. NADPH Oxidases in Lung Health and Disease

    PubMed Central

    Bernard, Karen; Hecker, Louise; Luckhardt, Tracy R.; Cheng, Guangjie

    2014-01-01

    Abstract Significance: The evolution of the lungs and circulatory systems in vertebrates ensured the availability of molecular oxygen (O2; dioxygen) for aerobic cellular metabolism of internal organs in large animals. O2 serves as the physiologic terminal acceptor of mitochondrial electron transfer and of the NADPH oxidase (Nox) family of oxidoreductases to generate primarily water and reactive oxygen species (ROS), respectively. Recent advances: The purposeful generation of ROS by Nox family enzymes suggests important roles in normal physiology and adaptation, most notably in host defense against invading pathogens and in cellular signaling. Critical issues: However, there is emerging evidence that, in the context of chronic stress and/or aging, Nox enzymes contribute to the pathogenesis of a number of lung diseases. Future Directions: Here, we review evolving functions of Nox enzymes in normal lung physiology and emerging pathophysiologic roles in lung disease. Antioxid. Redox Signal. 20, 2838–2853. PMID:24093231

  2. Modeling dioxygen reduction at multicopper oxidase cathodes.

    PubMed

    Agbo, Peter; Heath, James R; Gray, Harry B

    2014-10-01

    We report a general kinetics model for catalytic dioxygen reduction on multicopper oxidase (MCO) cathodes. Our rate equation combines Butler-Volmer (BV) electrode kinetics and the Michaelis-Menten (MM) formalism for enzymatic catalysis, with the BV model accounting for interfacial electron transfer (ET) between the electrode surface and the MCO type 1 copper site. Extending the principles of MM kinetics to this system produced an analytical expression incorporating the effects of subsequent intramolecular ET and dioxygen binding to the trinuclear copper cluster into the cumulative model. We employed experimental electrochemical data on Thermus thermophilus laccase as benchmarks to validate our model, which we suggest will aid in the design of more efficient MCO cathodes. In addition, we demonstrate the model's utility in determining estimates for both the electronic coupling and average distance between the laccase type-1 active site and the cathode substrate. PMID:25188422

  3. POLYAMINE OXIDASE 1 from rice (Oryza sativa) is a functional ortholog of Arabidopsis POLYAMINE OXIDASE 5

    PubMed Central

    Liu, Taibo; Wook Kim, Dong; Niitsu, Masaru; Berberich, Thomas; Kusano, Tomonobu

    2014-01-01

    POLYAMINE OXIDASE 1 (OsPAO1), from rice (Oryza sativa), and POLYAMINE OXIDASE 5 (AtPAO5), from Arabidopsis (Arabidopsis thaliana), are enzymes sharing high identity at the amino acid level and with similar characteristics, such as polyamine specificity and pH preference; furthermore, both proteins localize to the cytosol. A loss-of-function Arabidopsis mutant, Atpao5–2, was hypersensitive to low doses of exogenous thermospermine but this phenotype could be rescued by introduction of the wild-type AtPAO5 gene. Introduction of OsPAO1, under the control of a constitutive promoter, into Atpao5–2 mutants also restored normal thermospermine sensitivity, allowing growth in the presence of low levels of thermospermine, along with a concomitant decrease in thermospermine content in plants. By contrast, introduction of OsPAO3, which encodes a peroxisome-localized polyamine oxidase, into Atpao5–2 plants could not rescue any of the mutant phenotypes in the presence of thermospermine. These results suggest that OsPAO1 is the functional ortholog of AtPAO5. PMID:25763711

  4. POLYAMINE OXIDASE 1 from rice (Oryza sativa) is a functional ortholog of Arabidopsis POLYAMINE OXIDASE 5.

    PubMed

    Liu, Taibo; Wook Kim, Dong; Niitsu, Masaru; Berberich, Thomas; Kusano, Tomonobu

    2014-07-25

    POLYAMINE OXIDASE 1 (OsPAO1), from rice (Oryza sativa), and POLYAMINE OXIDASE 5 (AtPAO5), from Arabidopsis (Arabidopsis thaliana), are enzymes sharing high identity at the amino acid level and with similar characteristics, such as polyamine specificity and pH preference; furthermore, both proteins localize to the cytosol. A loss-of-function Arabidopsis mutant, Atpao5-2, was hypersensitive to low doses of exogenous thermospermine but this phenotype could be rescued by introduction of the wild-type AtPAO5 gene. Introduction of OsPAO1, under the control of a constitutive promoter, into Atpao5-2 mutants also restored normal thermospermine sensitivity, allowing growth in the presence of low levels of thermospermine, along with a concomitant decrease in thermospermine content in plants. By contrast, introduction of OsPAO3, which encodes a peroxisome-localized polyamine oxidase, into Atpao5-2 plants could not rescue any of the mutant phenotypes in the presence of thermospermine. These results suggest that OsPAO1 is the functional ortholog of AtPAO5. PMID:25061821

  5. Lysyl oxidase binds transforming growth factor-beta and regulates its signaling via amine oxidase activity.

    PubMed

    Atsawasuwan, Phimon; Mochida, Yoshiyuki; Katafuchi, Michitsuna; Kaku, Masaru; Fong, Keith S K; Csiszar, Katalin; Yamauchi, Mitsuo

    2008-12-01

    Lysyl oxidase (LOX), an amine oxidase critical for the initiation of collagen and elastin cross-linking, has recently been shown to regulate cellular activities possibly by modulating the functions of growth factors. In this study, we investigated the interaction between LOX and transforming growth factor-beta1 (TGF-beta1), a potent growth factor abundant in bone, the effect of LOX on TGF-beta1 signaling, and its potential mechanism. The specific binding between mature LOX and mature TGF-beta1 was demonstrated by immunoprecipitation and glutathione S-transferase pulldown assay in vitro. Both proteins were colocalized in the extracellular matrix in an osteoblastic cell culture system, and the binding complex was identified in the mineral-associated fraction of bone matrix. Furthermore, LOX suppressed TGF-beta1-induced Smad3 phosphorylation likely through its amine oxidase activity. The data indicate that LOX binds to mature TGF-beta1 and enzymatically regulates its signaling in bone and thus may play an important role in bone maintenance and remodeling. PMID:18835815

  6. Phagocyte NADPH oxidase and specific immunity.

    PubMed

    Cachat, Julien; Deffert, Christine; Hugues, Stephanie; Krause, Karl-Heinz

    2015-05-01

    The phagocyte NADPH oxidase NOX2 produces reactive oxygen species (ROS) and is a well-known player in host defence. However, there is also increasing evidence for a regulatory role of NOX2 in adaptive immunity. Deficiency in phagocyte NADPH oxidase causes chronic granulomatous disease (CGD) in humans, a condition that can also be studied in CGD mice. Clinical observations in CGD patients suggest a higher susceptibility to autoimmune diseases, in particular lupus, idiopathic thrombocytopenic purpura and rheumatoid arthritis. In mice, a strong correlation exists between a polymorphism in a NOX2 subunit and the development of autoimmune arthritis. NOX2 deficiency in mice also favours lupus development. Both CGD patients and CGD mice exhibit increased levels of immunoglobulins, including autoantibodies. Despite these phenotypes suggesting a role for NOX2 in specific immunity, mechanistic explanations for the typical increase of CGD in autoimmune disease and antibody levels are still preliminary. NOX2-dependent ROS generation is well documented for dendritic cells and B-lymphocytes. It is unclear whether T-lymphocytes produce ROS themselves or whether they are exposed to ROS derived from dendritic cells during the process of antigen presentation. ROS are signalling molecules in virtually any cell type, including T- and B-lymphocytes. However, knowledge about the impact of ROS-dependent signalling on T- and B-lymphocyte phenotype and response is still limited. ROS might contribute to Th1/Th2/Th17 cell fate decisions during T-lymphocyte activation and might enhance immunoglobulin production by B-lymphocytes. In dendritic cells, NOX2-derived ROS might be important for antigen processing and cell activation. PMID:25760962

  7. Polyphenol Oxidase Activity Expression in Ralstonia solanacearum

    PubMed Central

    Hernández-Romero, Diana; Solano, Francisco; Sanchez-Amat, Antonio

    2005-01-01

    Sequencing of the genome of Ralstonia solanacearum revealed several genes that putatively code for polyphenol oxidases (PPOs). To study the actual expression of these genes, we looked for and detected all kinds of PPO activities, including laccase, cresolase, and catechol oxidase activities, in cellular extracts of this microorganism. The conditions for the PPO assays were optimized for the phenolic substrate, pH, and sodium dodecyl sulfate concentration used. It was demonstrated that three different PPOs are expressed. The genes coding for the enzymes were unambiguously correlated with the enzymatic activities detected by generation of null mutations in the genes by using insertional mutagenesis with a suicide plasmid and estimating the changes in the levels of enzymatic activities compared to the levels in the wild-type strain. The protein encoded by the RSp1530 locus is a multicopper protein with laccase activity. Two other genes, RSc0337 and RSc1501, code for nonblue copper proteins exhibiting homology to tyrosinases. The product of RSc0337 has strong tyrosine hydroxylase activity, and it has been shown that this enzyme is involved in melanin synthesis by R. solanacearum. The product of the RSc1501 gene is an enzyme that shows a clear preference for oxidation of o-diphenols. Preliminary characterization of the mutants obtained indicated that PPOs expressed by R. solanacearum may participate in resistance to phenolic compounds since the mutants exhibited higher sensitivity to l-tyrosine than the wild-type strain. These results suggest a possible role in the pathogenic process to avoid plant resistance mechanisms involving the participation of phenolic compounds. PMID:16269713

  8. Evaluation of oxalate decarboxylase and oxalate oxidase for industrial applications.

    PubMed

    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. PMID:19763895

  9. Dephenolization of industrial wastewaters catalyzed by polyphenol oxidase

    SciTech Connect

    Atlow, S.C.; Bonadonna-Aparo, L.; Klibanov, A.M.

    1984-01-01

    A new enzymatic method for the removal of phenols from industrial aqueous effluents has been developed. The method uses the enzyme polyphenol oxidase which oxidizes phenols to the corresponding o-quinones; the latter then undergo a nonenzymatic polymerization to form water-insoluble aggregates. Therefore, the enzyme in effect precipitates phenols from water. Polyphenol oxidase has been found to nearly completely dephenolize solutions of phenol in the concentration range from 0.01 to 1.0 g/L. The enzymatic treatment is effective over a wide range of pH and temperature; a crude preparation of polyphenol oxidase (mushroom extract) is as effective as a purified, commercially obtained version. In addition to phenol itself, polyphenol oxidase is capable of precipitating from water a number of substituted phenols (cresols, chlorophenols, naphthol, etc.). Also, even pollutants which are unreactive towards polyphenol oxidase can be enzymatically coprecipitated with phenol. The polyphenol oxidase treatment has been successfully used to dephenolize two different real industrial wastewater samples, from a plant producing triarylphosphates and from a coke plant. The advantage of the polyphenol oxidase dephenolization over the peroxidase-catalyzed one previously elaborated by the authors is that the former enzyme uses molecular oxygen instead of costly hydrogen peroxide (used by peroxidase) as an oxidant.

  10. Surface enhanced Raman scattering investigation of protein-bound flavin adenine dinucleotide structure

    NASA Astrophysics Data System (ADS)

    Maskevich, S. A.; Strekal, N. D.; Artsukevich, I. M.; Kivach, L. N.; Chernikevich, I. P.

    1995-04-01

    The SERS spectra of alcohol oxidase from Pichia pastoris adsorbed on a silver electrode were obtained. The similarities and differences of these spectra with the SERS spectrum of free flavin adenine dinucleiotide were considered. The dependence of relative intensity of 1258 cm -1 band from the electrode potential in the protein SERS spectra differed from that of free flavin. From the data on this band being sensitive to the protein-flavin interaction a suggestion was made about incomplete dissociation of flavin from the protein. This conclusion is confirmed both by the fluorescence data and the SERS data on alcohol oxidase purified from Candida boidinii. The results of the SERS investigation of the interaction between the substrate, ethanol and the cofactor, FAD, as well as between protein-bound cofactor with the substrate are presented. The problem of retaining the protein enzyme activity is discussed.

  11. A Highly Stable D-Amino Acid Oxidase of the Thermophilic Bacterium Rubrobacter xylanophilus.

    PubMed

    Takahashi, Shouji; Furukawara, Makoto; Omae, Keishi; Tadokoro, Namiho; Saito, Yayoi; Abe, Katsumasa; Kera, Yoshio

    2014-12-01

    d-Amino acid oxidase (DAO) is a biotechnologically attractive enzyme that can be used in a variety of applications, but its utility is limited by its relatively poor stability. A search of a bacterial genome database revealed a gene encoding a protein homologous to DAO in the thermophilic bacterium Rubrobacter xylanophilus (RxDAO). The recombinant protein expressed in Escherichia coli was a monomeric protein containing noncovalently bound flavin adenine dinucleotide as a cofactor. This protein exhibited oxidase activity against neutral and basic d-amino acids and was significantly inhibited by a DAO inhibitor, benzoate, but not by any of the tested d-aspartate oxidase (DDO) inhibitors, thus indicating that the protein is DAO. RxDAO exhibited higher activities and affinities toward branched-chain d-amino acids, with the highest specific activity toward d-valine and catalytic efficiency (kcat/Km) toward d-leucine. Substrate inhibition was observed in the case of d-tyrosine. The enzyme had an optimum pH range and temperature of pH 7.5 to 10 and 65°C, respectively, and was stable between pH 5.0 and pH 8.0, with a T50 (the temperature at which 50% of the initial enzymatic activity is lost) of 64°C. No loss of enzyme activity was observed after a 1-week incubation period at 30°C. This enzyme was markedly inactivated by phenylmethylsulfonyl fluoride but not by thiol-modifying reagents and diethyl pyrocarbonate, which are known to inhibit certain DAOs. These results demonstrated that RxDAO is a highly stable DAO and suggested that this enzyme may be valuable for practical applications, such as the determination and quantification of branched-chain d-amino acids, and as a scaffold to generate a novel DAO via protein engineering. PMID:25217016

  12. Structure and proposed mechanism of α-glycerophosphate oxidase from Mycoplasma pneumoniae

    DOE PAGESBeta

    Elkhal, Callia K.; Kean, Kelsey M.; Parsonage, Derek; Maenpuen, Somchart; Chaiyen, Pimchai; Claiborne, Al; Karplus, P. Andrew

    2015-03-14

    In this study, the formation of hydrogen peroxide (H₂O₂) by the FAD-dependent α-glycerophosphate oxidase (GlpO), is important for the pathogenesis of Streptococcus pneumoniae and Mycoplasma pneumoniae. The structurally known GlpO from Streptococcus sp. (SspGlpO) is similar to the pneumococcal protein (SpGlpO) and provides a guide for drug design against that target. However, M. pneumoniae GlpO (MpGlpO), having <20% sequence identity with structurally known GlpOs, appears to represent a second type of GlpO we designate as Type II GlpOs. Here, the recombinant His-tagged MpGlpO structure is described at ~2.5 Å resolution, solved by molecular replacement using as a search model themore » Bordetella pertussis protein 3253 (Bp3253) a protein of unknown function solved by structural genomics efforts. Recombinant MpGlpO is an active oxidase with a turnover number of ~580 min⁻¹ while Bp3253 showed no GlpO activity. No substantial differences exist between the oxidized and dithionite-reduced MpGlpO structures. Although, no liganded structures were determined, a comparison with the tartrate-bound Bp3253 structure and consideration of residue conservation patterns guided the construction of a model for α-glycerophosphate (Glp) recognition and turnover by MpGlpO. The predicted binding mode also appears relevant for the type I GlpOs (such as SspGlpO) despite differences in substrate recognition residues, and it implicates a histidine conserved in type I and II Glp oxidases and dehydrogenases as the catalytic acid/base. This work provides a solid foundation for guiding further studies of the mitochondrial Glp dehydrogenases as well as for continued studies of M. pneumoniae and S. pneumoniae glycerol metabolism and the development of novel therapeutics targeting MpGlpO and SpGlpO.« less

  13. A Highly Stable d-Amino Acid Oxidase of the Thermophilic Bacterium Rubrobacter xylanophilus

    PubMed Central

    Furukawara, Makoto; Omae, Keishi; Tadokoro, Namiho; Saito, Yayoi; Abe, Katsumasa; Kera, Yoshio

    2014-01-01

    d-Amino acid oxidase (DAO) is a biotechnologically attractive enzyme that can be used in a variety of applications, but its utility is limited by its relatively poor stability. A search of a bacterial genome database revealed a gene encoding a protein homologous to DAO in the thermophilic bacterium Rubrobacter xylanophilus (RxDAO). The recombinant protein expressed in Escherichia coli was a monomeric protein containing noncovalently bound flavin adenine dinucleotide as a cofactor. This protein exhibited oxidase activity against neutral and basic d-amino acids and was significantly inhibited by a DAO inhibitor, benzoate, but not by any of the tested d-aspartate oxidase (DDO) inhibitors, thus indicating that the protein is DAO. RxDAO exhibited higher activities and affinities toward branched-chain d-amino acids, with the highest specific activity toward d-valine and catalytic efficiency (kcat/Km) toward d-leucine. Substrate inhibition was observed in the case of d-tyrosine. The enzyme had an optimum pH range and temperature of pH 7.5 to 10 and 65°C, respectively, and was stable between pH 5.0 and pH 8.0, with a T50 (the temperature at which 50% of the initial enzymatic activity is lost) of 64°C. No loss of enzyme activity was observed after a 1-week incubation period at 30°C. This enzyme was markedly inactivated by phenylmethylsulfonyl fluoride but not by thiol-modifying reagents and diethyl pyrocarbonate, which are known to inhibit certain DAOs. These results demonstrated that RxDAO is a highly stable DAO and suggested that this enzyme may be valuable for practical applications, such as the determination and quantification of branched-chain d-amino acids, and as a scaffold to generate a novel DAO via protein engineering. PMID:25217016

  14. Photochemical and ligand-exchange properties of the cyanide complex of fully reduced cytochrome c oxidase.

    PubMed Central

    Hill, B C; Marmor, S

    1991-01-01

    Cytochrome oxidase, in its fully reduced state, forms a complex with CN having a Kd of 230 microM with a stoicheiometry of 1 CN molecule per cytochrome oxidase. We do not detect a second CN-binding site as seen by i.r. spectroscopy [Yoshikawa & Caughey (1990) J. Biol. Chem. 265, 7945-7958]. The ferrocytochrome a3-CN complex, like the analogous ferrocytochrome a3-CO complex, is photosensitive but with a 15-fold lower quantum yield for photolysis. Analysis of the recombination kinetics after CN photolysis establishes a simple bimolecular binding constant of 235 M-1.s-1, in agreement with the value obtained from stopped-flow studies [Antonini, Brunori, Greenwood, Malmström & Rotillo (1971) Eur. J. Biochem. 23, 396-400]. A rate of 0.07 s-1 for the first-order dissociation of CN from cytochrome a3 is found by the rate of exchange of CO with ferrocytochrome a3-CN, and is consistent with the value calculated from the equilibrium binding constant and the CN on rate. However, O2 is able to oxidize the fully reduced CN compound at a rate well in excess of the CN off rate. The product of this oxidation reaction is a partially reduced CN complex. This implies that O2 either promotes CN dissociation or is able to oxidize the CN-bound enzyme directly. These results are discussed in the context of the structure and dynamics of the ligand-binding site of cytochrome oxidase. PMID:1659377

  15. Bounding the elliptic Mahler measure

    NASA Astrophysics Data System (ADS)

    Pinner, Christopher

    1998-11-01

    We give a simple inequality relating the elliptic Mahler measure of a polynomial to the traditional Mahler measure (via the length of the polynomial). These bounds are essentially sharp. We also give the corresponding result for polynomials in several variables.

  16. Immobilization of Pichia pastoris cells containing alcohol oxidase activity

    PubMed Central

    Maleknia, S; Ahmadi, H; Norouzian, D

    2011-01-01

    Background and Objectives The attempts were made to describe the development of a whole cell immobilization of P. pastoris by entrapping the cells in polyacrylamide gel beads. The alcohol oxidase activity of the whole cell Pichia pastoris was evaluated in comparison with yeast biomass production. Materials and Methods Methylotrophic yeast P. pastoris was obtained from Collection of Standard Microorganisms, Department of Bacterial Vaccines, Pasteur Institute of Iran (CSMPI). Stock culture was maintained on YPD agar plates. Alcohol oxidase was strongly induced by addition of 0.5% methanol as the carbon source. The cells were harvested by centrifugation then permeabilized. Finally the cells were immobilized in polyacrylamide gel beads. The activity of alcohol oxidase was determined by method of Tane et al. Results At the end of the logarithmic phase of cell culture, the alcohol oxidase activity of the whole cell P. Pastoris reached the highest level. In comparison, the alcohol oxidase activity was measured in an immobilized P. pastoris when entrapped in polyacrylamide gel beads. The alcohol oxidase activity of cells was induced by addition of 0.5% methanol as the carbon source. The cells were permeabilized by cetyltrimethylammonium bromide (CTAB) and immobilized. CTAB was also found to increase the gel permeability. Alcohol oxidase activity of immobilized cells was then quantitated by ABTS/POD spectrophotometric method at OD 420. There was a 14% increase in alcohol oxidase activity in immobilized cells as compared with free cells. By addition of 2-butanol as a substrate, the relative activity of alcohol oxidase was significantly higher as compared with other substrates added to the reaction media. Conclusion Immobilization of cells could eliminate lengthy and expensive procedures of enzyme separation and purification, protect and stabilize enzyme activity, and perform easy separation of the enzyme from the reaction media. PMID:22530090

  17. CotA, a Multicopper Oxidase from Bacillus pumilus WH4, Exhibits Manganese-Oxidase Activity

    PubMed Central

    Su, Jianmei; Bao, Peng; Bai, Tenglong; Deng, Lin; Wu, Hui; Liu, Fan; He, Jin

    2013-01-01

    Multicopper oxidases (MCOs) are a family of enzymes that use copper ions as cofactors to oxidize various substrates. Previous research has demonstrated that several MCOs such as MnxG, MofA and MoxA can act as putative Mn(II) oxidases. Meanwhile, the endospore coat protein CotA from Bacillus species has been confirmed as a typical MCO. To study the relationship between CotA and the Mn(II) oxidation, the cotA gene from a highly active Mn(II)-oxidizing strain Bacillus pumilus WH4 was cloned and overexpressed in Escherichia coli strain M15. The purified CotA contained approximately four copper atoms per molecule and showed spectroscopic properties typical of blue copper oxidases. Importantly, apart from the laccase activities, the CotA also displayed substantial Mn(II)-oxidase activities both in liquid culture system and native polyacrylamide gel electrophoresis. The optimum Mn(II) oxidase activity was obtained at 53°C in HEPES buffer (pH 8.0) supplemented with 0.8 mM CuCl2. Besides, the addition of o-phenanthroline and EDTA both led to a complete suppression of Mn(II)-oxidizing activity. The specific activity of purified CotA towards Mn(II) was 0.27 U/mg. The Km, Vmax and kcat values towards Mn(II) were 14.85±1.17 mM, 3.01×10−6±0.21 M·min−1 and 0.32±0.02 s−1, respectively. Moreover, the Mn(II)-oxidizing activity of the recombinant E. coli strain M15-pQE-cotA was significantly increased when cultured both in Mn-containing K liquid medium and on agar plates. After 7-day liquid cultivation, M15-pQE-cotA resulted in 18.2% removal of Mn(II) from the medium. Furthermore, the biogenic Mn oxides were clearly observed on the cell surfaces of M15-pQE-cotA by scanning electron microscopy. To our knowledge, this is the first report that provides the direct observation of Mn(II) oxidation with the heterologously expressed protein CotA, Therefore, this novel finding not only establishes the foundation for in-depth study of Mn(II) oxidation mechanisms, but also offers a

  18. The terminal oxidase in the marine bacterium Pseudomonas nautica 617.

    PubMed

    Simpson, H; Denis, M; Malatesta, F

    1997-06-01

    The molecular properties of a novel membrane quinol oxidase from the marine bacterium Pseudomonas nautica 617 are presented. The protein contains 2b hemes/mole which may be distinguished by EPR spectroscopy but not by optical spectroscopy and electrochemistry. Respiration, though being cyanide insensitive, is not inhibited by carbon monoxide and oxygen reduction is carried out only half-way with production of hydrogen peroxide. The terminal oxidase represents, therefore, a unique example in the large family of terminal oxidases known up to date. PMID:9337488

  19. Identification of yeasts from clinical specimens by oxidase test.

    PubMed

    Kumar, S; Arora, B S; Mathur, M D

    2000-10-01

    A total of 100 yeasts and yeast like fungi isolates from clinical specimens were negative for oxidase production on Sabouraud dextrose agar. When grown on Columbia agar, chocolate agar, tryptose agar, Mueller-Hinton agar, brain heart infusion and a medium resembling Sabouraud's dextrose agar but with starch instead of dextrose, all the isolate of Candida albicans (55), C. guilliermondii (6), C. parapsilosis (14), C. tropicalis (6), C. pseudotropicalis (6) and Crytococcus neoformans (2) were positive for oxidase producation. Torulopsis glabrata (2), Saccharomyces cervisiae (2) and two out of seven isolates of C. krusei were negative for oxidase test. PMID:11344606

  20. Aiding and abetting roles of NOX oxidases in cellular transformation

    PubMed Central

    Block, Karen; Gorin, Yves

    2013-01-01

    NADPH oxidases of the NADPH oxidase (NOX) family are dedicated reactive oxygen species-generating enzymes that broadly and specifically regulate redox-sensitive signalling pathways that are involved in cancer development and progression. They act at specific cellular membranes and microdomains through the activation of oncogenes and the inactivation of tumour suppressor proteins. In this Review, we discuss primary targets and redox-linked signalling systems that are influenced by NOX-derived ROS, and the biological role of NOX oxidases in the aetiology of cancer. PMID:22918415

  1. Multilayered Polyelectrolyte Microcapsules: Interaction with the Enzyme Cytochrome C Oxidase

    PubMed Central

    Pastorino, Laura; Dellacasa, Elena; Noor, Mohamed R.; Soulimane, Tewfik; Bianchini, Paolo; D'Autilia, Francesca; Antipov, Alexei; Diaspro, Alberto; Tofail, Syed A. M.; Ruggiero, Carmelina

    2014-01-01

    Cell-sized polyelectrolyte capsules functionalized with a redox-driven proton pump protein were assembled for the first time. The interaction of polyelectrolyte microcapsules, fabricated by electrostatic layer-by-layer assembly, with cytochrome c oxidase molecules was investigated. We found that the cytochrome c oxidase retained its functionality, that the functionalized microcapsules interacting with cytochrome c oxidase were permeable and that the permeability characteristics of the microcapsule shell depend on the shell components. This work provides a significant input towards the fabrication of an integrated device made of biological components and based on specific biomolecular functions and properties. PMID:25372607

  2. Comparison of coupling subsites and inhibition effects of piperidine alkaloids and aminoketones on plant amine oxidases.

    PubMed

    Pec, P; Frébort, I

    1991-01-01

    In the present work we compare the binding subsites of inhibitors from a series of alkaloids and aminoketones on pea and sainfoin diamine oxidase (EC 1.4.3.6; DAO) by the graphical method. As standard competitive inhibitors we have chosen oxoanalogs of the substrates, namely, 1,4-diamino-2-butanone and 1,5-diamino-3-pentanone, which were compared with the alkaloids (+)-sedamine, (-)-norallosedamine, (-)-norsedamine, L-lobeline, cinchonine and aromatic analogs of aliphatic aminoketones such as 1-amino-3-phenyl-3-propanone and 1-amino-3-phenyl-2-propanone. In the case of pea DAO all inhibitors compete for the same subsites with 1,4-diamino-2-butanone and 1,5-diamino-3-pentanone (alpha = infinity). In the case of sainfoin enzyme they are bound to other subsites and the interaction constants (0 < alpha < 1) point to a positive attraction between these two types of inhibitors. With sainfoin DAO, 1-amino-3-phenyl-3-propanone is bound into the same subsite as 1,4-diamino-2-butanone. Cinchonine and 1-amino-3-phenyl-3-propanone are bound to two different subsites and the value of the interaction constant (1 < alpha < infinity) shows repulsion between the inhibitors. PMID:1669832

  3. Three Redox States of Trypanosoma brucei Alternative Oxidase Identified by Infrared Spectroscopy and Electrochemistry

    PubMed Central

    Maréchal, Amandine; Kido, Yasutoshi; Kita, Kiyoshi; Moore, Anthony L.; Rich, Peter R.

    2009-01-01

    Electrochemistry coupled with Fourier transform infrared (IR) spectroscopy was used to investigate the redox properties of recombinant alternative ubiquinol oxidase from Trypanosoma brucei, the organism responsible for African sleeping sickness. Stepwise reduction of the fully oxidized resting state of recombinant alternative ubiquinol oxidase revealed two distinct IR redox difference spectra. The first of these, signal 1, titrates in the reductive direction as an n = 2 Nernstian component with an apparent midpoint potential of 80 mV at pH 7.0. However, reoxidation of signal 1 in the same potential range under anaerobic conditions did not occur and only began with potentials in excess of 500 mV. Reoxidation by introduction of oxygen was also unsuccessful. Signal 1 contained clear features that can be assigned to protonation of at least one carboxylate group, further perturbations of carboxylic and histidine residues, bound ubiquinone, and a negative band at 1554 cm−1 that might arise from a radical in the fully oxidized protein. A second distinct IR redox difference spectrum, signal 2, appeared more slowly once signal 1 had been reduced. This component could be reoxidized with potentials above 100 mV. In addition, when both signals 1 and 2 were reduced, introduction of oxygen caused rapid oxidation of both components. These data are interpreted in terms of the possible active site structure and mechanism of oxygen reduction to water. PMID:19767647

  4. Structural analysis of aliphatic versus aromatic substrate specificity in a copper amine oxidase from Hansenula polymorpha.

    PubMed

    Klema, Valerie J; Solheid, Corinne J; Klinman, Judith P; Wilmot, Carrie M

    2013-04-01

    Copper amine oxidases (CAOs) are responsible for the oxidative deamination of primary amines to their corresponding aldehydes. The CAO catalytic mechanism can be divided into two half-reactions: a reductive half-reaction in which a primary amine substrate is oxidized to its corresponding aldehyde with the concomitant reduction of the organic cofactor 2,4,5-trihydroxyphenylalanine quinone (TPQ) and an oxidative half-reaction in which reduced TPQ is reoxidized with the reduction of molecular oxygen to hydrogen peroxide. The reductive half-reaction proceeds via Schiff base chemistry, in which the primary amine substrate first attacks the C5 carbonyl of TPQ, forming a series of covalent Schiff base intermediates. The X-ray crystal structures of copper amine oxidase-1 from the yeast Hansenula polymorpha (HPAO-1) in complex with ethylamine and benzylamine have been determined to resolutions of 2.18 and 2.25 Å, respectively. These structures reveal the two amine substrates bound at the back of the active site coincident with TPQ in its two-electron-reduced aminoquinol form. Rearrangements of particular amino acid side chains within the substrate channel and specific protein-substrate interactions provide insight into the substrate specificity of HPAO-1. These changes begin to account for this CAO's kinetic preference for small, aliphatic amines over the aromatic amines or whole peptides preferred by some of its homologues. PMID:23452079

  5. Structural Snapshots from the Oxidative Half-reaction of a Copper Amine Oxidase

    PubMed Central

    Johnson, Bryan J.; Yukl, Erik T.; Klema, Valerie J.; Klinman, Judith P.; Wilmot, Carrie M.

    2013-01-01

    The mechanism of molecular oxygen activation is the subject of controversy in the copper amine oxidase family. At their active sites, copper amine oxidases contain both a mononuclear copper ion and a protein-derived quinone cofactor. Proposals have been made for the activation of molecular oxygen via both a Cu(II)-aminoquinol catalytic intermediate and a Cu(I)-semiquinone intermediate. Using protein crystallographic freeze-trapping methods under low oxygen conditions combined with single-crystal microspectrophotometry, we have determined structures corresponding to the iminoquinone and semiquinone forms of the enzyme. Methylamine reduction at acidic or neutral pH has revealed protonated and deprotonated forms of the iminoquinone that are accompanied by a bound oxygen species that is likely hydrogen peroxide. However, methylamine reduction at pH 8.5 has revealed a copper-ligated cofactor proposed to be the semiquinone form. A copper-ligated orientation, be it the sole identity of the semiquinone or not, blocks the oxygen-binding site, suggesting that accessibility of Cu(I) may be the basis of partitioning O2 activation between the aminoquinol and Cu(I). PMID:23940035

  6. Surface functionalization of silica-coated magnetic nanoparticles for covalent attachment of cholesterol oxidase

    NASA Astrophysics Data System (ADS)

    Šulek, Franja; Drofenik, Miha; Habulin, Maja; Knez, Željko

    2010-01-01

    A systematic approach towards the fabrication of highly functionalized silica shell magnetic nanoparticles, presently used for enzyme immobilization, is herein fully presented. The synthesis of bare maghemite (γ-Fe 2O 3) nanoparticles was accomplished by thermal co-precipitation of iron ions in ammonia alkaline solution at harsh reaction conditions, respectively. Primary surface engineering of maghemite nanoparticles was successfully performed by the proper deposition of silica onto nanoparticles surface under strictly regulated reaction conditions. Next, the secondary surface functionalization of the particles was achieved by coating the particles with organosilane followed by glutaraldehyde activation in order to enhance protein immobilization. Covalent immobilization of cholesterol oxidase was attempted afterwards. The structural and magnetic properties of magnetic silica nanocomposites were characterized by TEM and vibrating sample magnetometer (VSM) instruments. X-ray diffraction measurements confirmed the spinel structure and average size of uncoated maghemite nanoparticles to be around 20 nm in diameter. SEM-EDS spectra indicated a strong signal for Si, implying the coating procedure of silica onto the particles surface to be successfully accomplished. Fourier transform infrared (FT-IR) spectra analysis confirmed the binding of amino silane molecules onto the surface of the maghemite nanoparticles mediated Si-O-Si chemical bonds. Compared to the free enzyme, the covalently bound cholesterol oxidase retained 50% of its activity. Binding of enzyme onto chemically modified magnetic nanoparticles via glutaraldehyde activation is a promising method for developing biosensing components in biomedicine.

  7. Analysis of water channels by molecular dynamics simulation of heterotetrameric sarcosine oxidase

    PubMed Central

    Watanabe, Go; Nakajima, Daisuke; Hiroshima, Akinori; Suzuki, Haruo; Yoneda, Shigetaka

    2015-01-01

    A precise 100-ns molecular dynamics simulation in aquo was performed for the heterotetrameric sarcosine oxidase bound with a substrate analogue, dimethylglycine. The spatial region including the protein was divided into small rectangular cells. The average number of the water molecules locating within each cell was calculated based on the simulation trajectory. The clusters of the cells filled with water molecules were used to determine the water channels. The narrowness of the channels, the average hydropathy indices of the residues of the channels, and the number of migration events of water molecules through the channels were consistent with the selective transport hypothesis whereby tunnel T3 is the pathway for the exit of the iminium intermediate of the enzyme reaction. PMID:27493862

  8. Parkin degrades estrogen-related receptors to limit the expression of monoamine oxidases.

    PubMed

    Ren, Yong; Jiang, Houbo; Ma, Dingyuan; Nakaso, Kazuhiro; Feng, Jian

    2011-03-15

    Parkin, whose mutations cause Parkinson disease (PD), controls oxidative stress by limiting the expression of monoamine oxidases (MAO)--mitochondrial enzymes responsible for the oxidative de-amination of dopamine. Here, we show that parkin performed this function by increasing the ubiquitination and degradation of estrogen-related receptors (ERR), orphan nuclear receptors that play critical roles in the transcription regulation of many nuclear-encoded mitochondrial proteins. All three ERRs (α, β and γ) increased the transcription of MAOs A and B; the effects were abolished by parkin, but not by its PD-linked mutants. Parkin bound to ERRs and increased their ubiquitination and degradation. In fibroblasts from PD patients with parkin mutations or brain slices from parkin knockout mice, degradation of ERRs was significantly attenuated. The results reveal the molecular mechanism by which parkin suppresses the transcription of MAOs to control oxidative stress induced by dopamine oxidation. PMID:21177257

  9. Parkin degrades estrogen-related receptors to limit the expression of monoamine oxidases

    PubMed Central

    Ren, Yong; Jiang, Houbo; Ma, Dingyuan; Nakaso, Kazuhiro; Feng, Jian

    2011-01-01

    Parkin, whose mutations cause Parkinson disease (PD), controls oxidative stress by limiting the expression of monoamine oxidases (MAO)—mitochondrial enzymes responsible for the oxidative de-amination of dopamine. Here, we show that parkin performed this function by increasing the ubiquitination and degradation of estrogen-related receptors (ERR), orphan nuclear receptors that play critical roles in the transcription regulation of many nuclear-encoded mitochondrial proteins. All three ERRs (α, β and γ) increased the transcription of MAOs A and B; the effects were abolished by parkin, but not by its PD-linked mutants. Parkin bound to ERRs and increased their ubiquitination and degradation. In fibroblasts from PD patients with parkin mutations or brain slices from parkin knockout mice, degradation of ERRs was significantly attenuated. The results reveal the molecular mechanism by which parkin suppresses the transcription of MAOs to control oxidative stress induced by dopamine oxidation. PMID:21177257

  10. Cholesterol Oxidase Functionalised Polyaniline/Carbon Nanotube Hybrids for an Amperometric Biosensor.

    PubMed

    Shukla, Sudheesh K; Turner, Anthony P F; Tiwari, Ashutosh

    2015-05-01

    Functional carbon nanotubes (CNT) have attracted much attention for analytical and biomedical applications. This paper describes the fabrication of a cholesterol oxidase (ChOx) immobilised polyaniline (PANI)/CNT composite electrode for the amperometric detection of cholesterol. The prepared ChOx/PANI/CNT/Au bioelectrode bound ChOx via the available functionalties of PANI (-NH2) and CNT (-COOH). Moreover, the CNT creates a network inside the matrix that strengthens the mechanical property of the bioelectrode. The multifunctional matrix is presumed to provide a 3D-mesoporous surface, which substantially enhances enzyme activity. The linear range of the biosensor for cholesterol oleate was 30-280 μM with a response time of 10 sec. PMID:26504954

  11. Beyond brown: polyphenol oxidases as enzymes of plant specialized metabolism

    PubMed Central

    Sullivan, Michael L.

    2015-01-01

    Most cloned and/or characterized plant polyphenol oxidases (PPOs) have catechol oxidase activity (i.e., they oxidize o-diphenols to o-quinones) and are localized or predicted to be localized to plastids. As a class, they have broad substrate specificity and are associated with browning of produce and other plant materials. Because PPOs are often induced by wounding or pathogen attack, they are most generally believed to play important roles in plant defense responses. However, a few well-characterized PPOs appear to have very specific roles in the biosynthesis of specialized metabolites via both tyrosinase (monophenol oxidase) and catechol oxidase activities. Here we detail a few examples of these and explore the possibility that there may be many more “biosynthetic” PPOs. PMID:25642234

  12. Inhibition of plant and mammalian diamine oxidase by substrate analogues.

    PubMed

    Biegański, T; Osińska, Z; Masliński, C

    1982-04-01

    Imidazoles, aliphatic substrate analogues and the natural dipeptides, carnosine and anserine, were investigated as inhibitors of diamine oxidase from the pig kidney, human pregnancy plasma and pea seedlings. Imidazole, methylimidazoles, N-acetylimidazole, histamine and N tau-methylhistamine are relatively potent inhibitors of mammalian diamine oxidase showing no influence on plant enzymes. Anserine and carnosine are inhibitors of pig kidney and pea seedling enzymes. Ki values are 2 microM and 10 microM respectively. Investigated natural derivatives of putrescine and cadaverine have no influence on diamine oxidase of different origin. In conclusion, we present some evidence to suggest that mammalian diamine oxidase, despite a high reaction rate with putrescine, is better adapted to histamine oxidation, whereas for plant enzymes the diamines are preferred substrates. PMID:6805264

  13. [Heterogeneity of molecular forms of phenol oxidase from grape leaves].

    PubMed

    Pruidze, G N; Zaprometov, M N; Durmishidze, S V; Kintsurashvili, D F

    1983-07-01

    The substrate specificity and some kinetic properties of the monomeric (Mr = 26 000--35 000) and dimeric (Mr = 55 000--70 000) forms of phenol oxidase from vine leaves were studied. These forms possess different hydroxylating and o-diphenol oxidase activities. A kinetic analysis demonstrated that the monomeric form of the enzyme possesses a higher affinity for monophenols and can more effectively accomplish the hydroxylation reaction as compared to the dimeric one. During vine vegetation the ratio of molecular forms of phenol oxidase is altered manifesting itself in quantitative and qualitative changes of enzymatic activity. During plant maturation the dimeric fraction is predominant. The maturation process is associated with a sharp rise of the o-phenol oxidase activity, a disappearance of the hydroxylating activity and a substantial deceleration of phenol compounds production. PMID:6412775

  14. Reactions of bovine serum amine oxidase with NN-diethyldithiocarbamate. Selective removal of one copper ion.

    PubMed Central

    Morpurgo, L; Agostinelli, E; Befani, O; Mondovì, B

    1987-01-01

    NN-Diethyldithiocarbamate (DDC) was able to bind, at 1.0 mM concentration, only about 50% the Cu(II) ions of bovine plasma amine oxidase. Under reducing conditions, this Cu(II) was removed with inactivation of the enzyme. Up to 90% activity could be recovered by treatment with excess Cu(II). The organic cofactor, sensitive to carbonyl reagents, was reduced in the half-Cu-depleted protein and no longer bound phenylhydrazine. The fully reacted protein, in the presence of 10 mM-DDC, lost 50% Cu(II) upon storage at -20 degrees C, but in this case the residual Cu(II) was in the DDC-bound form and the cofactor was in the oxidized state, as it could still bind phenylhydrazine. In the presence of DDC, the rate of reaction with phenylhydrazine was always low, even at 50% DDC saturation, and all derivatives showed identical modifications of the optical and e.p.r. spectra with respect to the phenylhydrazone of the native protein. It is concluded that the two Cu(II) ions are not equivalent, that removal of a single Cu(II) is sufficient to inhibit the re-oxidation of the organic cofactor, and that both Cu(II) ions are in some way involved in the reaction with phenylhydrazine. After reaction with DDC, the optical and e.p.r. spectra of 63Cu(II)-amine oxidase and of 63Cu(II)-carbonic anhydrase [Morpurgo, Desideri, Rigo, Viglino & Rotilio (1983) Biochim. Biophys. Acta 746, 168-175] are very similar and show distorted equatorial co-ordination to Cu(II) of two sulphur atoms and two magnetically equivalent nitrogen atoms. PMID:2829844

  15. Structural characterization of acyl-CoA oxidases reveals a direct link between pheromone biosynthesis and metabolic state in Caenorhabditis elegans.

    PubMed

    Zhang, Xinxing; Li, Kunhua; Jones, Rachel A; Bruner, Steven D; Butcher, Rebecca A

    2016-09-01

    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

  16. Confirmation of a blocked amino terminus of sulfhydryl oxidase

    SciTech Connect

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

    1990-09-01

    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 ({sup 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 ({plus minus} 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.

  17. Lysyl oxidase activity regulates oncogenic stress response and tumorigenesis.

    PubMed

    Wiel, C; Augert, A; Vincent, D F; Gitenay, D; Vindrieux, D; Le Calvé, B; Arfi, V; Lallet-Daher, H; Reynaud, C; Treilleux, I; Bartholin, L; Lelievre, E; Bernard, D

    2013-01-01

    Cellular senescence, a stable proliferation arrest, is induced in response to various stresses. Oncogenic stress-induced senescence (OIS) results in blocked proliferation and constitutes a fail-safe program counteracting tumorigenesis. The events that enable a tumor in a benign senescent state to escape from OIS and become malignant are largely unknown. We show that lysyl oxidase activity contributes to the decision to maintain senescence. Indeed, in human epithelial cell the constitutive expression of the LOX or LOXL2 protein favored OIS escape, whereas inhibition of lysyl oxidase activity was found to stabilize OIS. The relevance of these in vitro observations is supported by in vivo findings: in a transgenic mouse model of aggressive pancreatic ductal adenocarcinoma (PDAC), increasing lysyl oxidase activity accelerates senescence escape, whereas inhibition of lysyl oxidase activity was found to stabilize senescence, delay tumorigenesis, and increase survival. Mechanistically, we show that lysyl oxidase activity favors the escape of senescence by regulating the focal-adhesion kinase. Altogether, our results demonstrate that lysyl oxidase activity participates in primary tumor growth by directly impacting the senescence stability. PMID:24113189

  18. Bound polarons in semiconductor nanostructures

    NASA Astrophysics Data System (ADS)

    Woggon, U.; Miller, D.; Kalina, F.; Gerlach, B.; Kayser, D.; Leonardi, K.; Hommel, D.

    2003-01-01

    Bound polarons are discrete, confined electronic states, spatially localized due to a local potential V(r) but sharing a common phonon state of the surrounding crystal. We study the energy states of polarons bound in a potential and determine the local optical absorption spectrum up to first-order time-dependent perturbation theory with respect to the electron-photon interaction. The model is applied to describe the optical properties of submonolayer CdSe insertions epitaxially grown between ZnSe layers. As a typical signature of bound polarons we found excited-state energies equidistantly separated by the LO phonon energy and with optical transition probabilities determined by the anisotropies in V(r).

  19. Unitarity bound for gluon shadowing

    SciTech Connect

    Kopeliovich, B. Z.; Levin, E.; Potashnikova, I. K.; Schmidt, Ivan

    2009-06-15

    Although at small Bjorken x gluons originated from different nucleons in a nucleus overlap in the longitudinal direction, most of them are still well separated in the transverse plane and therefore cannot fuse. For this reason the gluon density in nuclei cannot drop at small x below a certain bottom bound, which we evaluated in a model independent manner assuming the maximal strength of gluon fusion. We also calculated gluon shadowing in the saturated regime using the Balitsky-Kovchegov equation and found the nuclear ratio to be well above the unitarity bound. The recently updated analysis of parton distributions in nuclei, including BNL Relativistic Heavy Ion Collider (RHIC) data on high-p{sub T} hadron production at forward rapidities, led to strong gluon shadowing. Such strong shadowing and therefore the interpretation of the nuclear modification of the p{sub T} spectra in dA collisions at RHIC seem to be inconsistent with this unitarity bound.

  20. Molecular aspects of monoamine oxidase B.

    PubMed

    Ramsay, Rona R

    2016-08-01

    Monoamine oxidases (MAO) influence the monoamine levels in brain by virtue of their role in neurotransmitter breakdown. MAO B is the predominant form in glial cells and in platelets. MAO B structure, function and kinetics are described as a background for the effect of alterations in its activity on behavior. The need to inhibit MAO B to combat decreased brain amines continues to drive the search for new drugs. Reversible and irreversible inhibitors are now designed using data-mining, computational screening, docking and molecular dynamics. Multi-target ligands designed to combat the elevated activity of MAO B in Alzheimer's and Parkinson's Diseases incorporate MAO inhibition (usually irreversible) as well as iron chelation, antioxidant or neuroprotective properties. The main focus of drug design is the catalytic activity of MAO, but the imidazoline I2 site in the entrance cavity of MAO B is also a pharmacological target. Endogenous regulation of MAO B expression is discussed briefly in light of new studies measuring mRNA, protein, or activity in healthy and degenerative samples, including the effect of DNA methylation on the expression. Overall, this review focuses on examples of recent research on the molecular aspects of the expression, activity, and inhibition of MAO B. PMID:26891670

  1. Polyphenol oxidase from yacon roots (Smallanthus sonchifolius).

    PubMed

    Neves, Valdir Augusto; da Silva, Maraiza Aparecida

    2007-03-21

    Polyphenol oxidase (E.C. 1.14.18.1) (PPO) extracted from yacon roots (Smallanthus sonchifolius) was partially purified by ammonium sulfate fractionation and separation on Sephadex G-100. The enzyme had a molecular weight of 45 490+/-3500 Da and Km values of 0.23, 1.14, 1.34, and 5.0 mM for the substrates caffeic acid, chlorogenic acid, 4-methylcatechol, and catechol, respectively. When assayed with resorcinol, DL-DOPA, pyrogallol, protocatechuic, p-coumaric, ferulic, and cinnamic acids, catechin, and quercetin, the PPO showed no activity. The optimum pH varied from 5.0 to 6.6, depending on substrate. PPO activity was inhibited by various phenolic and nonphenolic compounds. p-Coumaric and cinnamic acids showed competitive inhibition, with Ki values of 0.017 and 0.011 mM, respectively, using chlorogenic acid as substrate. Heat inactivation from 60 to 90 degrees C showed the enzyme to be relatively stable at 60-70 degrees C, with progressive inactivation when incubated at 80 and 90 degrees C. The Ea (apparent activation energy) for inactivation was 93.69 kJ mol-1. Sucrose, maltose, glucose, fructose, and trehalose at high concentrations appeared to protect yacon PPO against thermal inactivation at 75 and 80 degrees C. PMID:17316020

  2. Origin and evolution of lysyl oxidases

    PubMed Central

    Grau-Bové, Xavier; Ruiz-Trillo, Iñaki; Rodriguez-Pascual, Fernando

    2015-01-01

    Lysyl oxidases (LOX) are copper-dependent enzymes that oxidize primary amine substrates to reactive aldehydes. The best-studied role of LOX enzymes is the remodeling of the extracellular matrix (ECM) in animals by cross-linking collagens and elastin, although intracellular functions have been reported as well. Five different LOX enzymes have been identified in mammals, LOX and LOX-like (LOXL) 1 to 4, showing a highly conserved catalytic carboxy terminal domain and more divergence in the rest of the sequence. Here we have surveyed a wide selection of genomes in order to infer the evolutionary history of LOX. We identified LOX proteins not only in animals, but also in many other eukaryotes, as well as in bacteria and archaea – which reveals a pre-metazoan origin for this gene family. LOX genes expanded during metazoan evolution resulting in two superfamilies, LOXL2/L3/L4 and LOX/L1/L5. Considering the current knowledge on the function of mammalian LOX isoforms in ECM remodeling, we propose that LOXL2/L3/L4 members might have preferentially been involved in making cross-linked collagen IV-based basement membrane, whereas the diversification of LOX/L1/L5 forms contributed to chordate/vertebrate-specific ECM innovations, such as elastin and fibronectin. Our work provides a novel view on the evolution of this family of enzymes. PMID:26024311

  3. Origin and evolution of lysyl oxidases.

    PubMed

    Grau-Bové, Xavier; Ruiz-Trillo, Iñaki; Rodriguez-Pascual, Fernando

    2015-01-01

    Lysyl oxidases (LOX) are copper-dependent enzymes that oxidize primary amine substrates to reactive aldehydes. The best-studied role of LOX enzymes is the remodeling of the extracellular matrix (ECM) in animals by cross-linking collagens and elastin, although intracellular functions have been reported as well. Five different LOX enzymes have been identified in mammals, LOX and LOX-like (LOXL) 1 to 4, showing a highly conserved catalytic carboxy terminal domain and more divergence in the rest of the sequence. Here we have surveyed a wide selection of genomes in order to infer the evolutionary history of LOX. We identified LOX proteins not only in animals, but also in many other eukaryotes, as well as in bacteria and archaea - which reveals a pre-metazoan origin for this gene family. LOX genes expanded during metazoan evolution resulting in two superfamilies, LOXL2/L3/L4 and LOX/L1/L5. Considering the current knowledge on the function of mammalian LOX isoforms in ECM remodeling, we propose that LOXL2/L3/L4 members might have preferentially been involved in making cross-linked collagen IV-based basement membrane, whereas the diversification of LOX/L1/L5 forms contributed to chordate/vertebrate-specific ECM innovations, such as elastin and fibronectin. Our work provides a novel view on the evolution of this family of enzymes. PMID:26024311

  4. MONOAMINE OXIDASE: RADIOTRACER DEVELOPMENT AND HUMAN STUDIES.

    SciTech Connect

    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.

  5. Photodynamic therapy using a protoporphyrinogen oxidase inhibitor.

    PubMed

    Fingar, V H; Wieman, T J; McMahon, K S; Haydon, P S; Halling, B P; Yuhas, D A; Winkelman, J W

    1997-10-15

    The use of endogenously created porphyrins as an alternative to photosensitizer injection for photodynamic therapy is a rapidly evolving area of study. One common method to induce porphyrin synthesis and accumulation in cells is the topical, oral, or parenteral administration of 5-aminolevulinic acid, a precursor for heme biosynthesis. Porphyrin accumulation may also be elicited by the use of enzyme inhibitors of the heme biosynthetic pathway. Groups of DBA/2 mice bearing SMT-F mammary tumors were placed on a diet containing 0-4000 ppm of a protoporphyrinogen oxidase inhibitor, FP-846. This agent blocks a critical step in porphyrin metabolism and results in elevated intracellular levels of protoporphyrin IX. Light treatment of tumors produced both initial and long-term regression that was dependent on the amount of inhibitor, the duration of inhibitor exposure to animals, and the amount of light used in PDT. Tumor regression occurred without significant destruction of normal tissues in the treatment field and without initial vascular constriction or blood flow stasis. Tumor cure in animals given 4000 ppm FP-846 in feed for 3 days and 300 J/cm2 602-670 nm light (23% cure) was similar to the response in animals given 10 mg/kg Photofrin and the same light dose (20%). PMID:9377568

  6. Analyzing the electrogenicity of cytochrome c oxidase.

    PubMed

    Kim, Ilsoo; Warshel, Arieh

    2016-07-12

    Measurements of voltage changes in response to charge separation within membrane proteins can offer fundamental information on spectroscopically "invisible" steps. For example, results from studies of voltage changes associated with electron and proton transfer in cytochrome c oxidase could, in principle, be used to discriminate between different theoretical models describing the molecular mechanism of proton pumping. Earlier analyses of data from these measurements have been based on macroscopic considerations that may not allow for exploring the actual molecular mechanisms. Here, we have used a coarse-grained model describing the relation between observed voltage changes and specific charge-transfer reactions, which includes an explicit description of the membrane, the electrolytes, and the electrodes. The results from these calculations offer mechanistic insights at the molecular level. Our main conclusion is that previously assumed mechanistic evidence that was based on electrogenic measurements is not unique. However, the ability of our calculations to obtain reliable voltage changes means that we have a tool that can be used to describe a wide range of electrogenic charge transfers in channels and transporters, by combining voltage measurements with other experiments and simulations to analyze new mechanistic proposals. PMID:27357681

  7. Molecularly "wired" cholesterol oxidase for biosensing.

    PubMed

    Leonida, Mihaela D; Aurian-Blajeni, Benedict

    2015-02-01

    The influence of several factors on the activity of cholesterol oxidase (ChOx) transiently exposed to a room temperature ionic liquid (RTIL) was studied. Presence of flavin adenine dinucleotide (FAD, prosthetic group of ChOx) during exposure to RTIL makes the procedure enzyme-friendly, while the use of RTIL (green reagent) makes it environmentally-friendly. Following exposure to RTIL and its subsequent removal, FAD becomes part of the molecular structure of the refolded protein (a molecular "wire"). This makes the procedure used here a molecular one. The factors studied were: FAD presence in RTIL during modification, water presence during exposure to RTIL, and ratio FAD:RTIL during "wiring". Performance parameters monitored were: enzyme activity before and after "wiring" (expressed as (dA/dt)/mg enzyme, and measured spectrophotometrically), peak current in an amperometric biosensor for cholesterol detection, and linearity of the biosensor response depending on cholesterol concentration. After RTIL removal, the modified enzyme (ME) retained a high percentage of the added FAD, which supplemented that of the native enzyme (functioning as a "wire" and enhancing electron transfer kinetics), and a fraction of the initial activity. Used in an amperometric biosensor, ME showed catalytic activity, linear behavior as a function of cholesterol concentration, and stability. PMID:25579496

  8. Crystallization of carbohydrate oxidase from Microdochium nivale.

    PubMed

    Dusková, Jarmila; Dohnálek, Jan; Skálová, Tereza; Østergaard, Lars Henrik; Fuglsang, Claus Crone; Kolenko, Petr; Stepánková, Andrea; Hasek, Jindrich

    2009-06-01

    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 A resolution, with unit-cell parameters a = b = 55.7, c = 610.4 A 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 A, beta = 95.7 degrees . Data sets were collected to a resolution of 2.4 A. The structure was solved by the molecular-replacement method. Model refinement is currently in progress. PMID:19478452

  9. Crystallization of carbohydrate oxidase from Microdochium nivale

    PubMed Central

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

    2009-01-01

    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 P6222. 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. PMID:19478452

  10. Monoamine oxidase: Radiotracer chemistry and human studies

    DOE PAGESBeta

    Fowler, Joanna S.; Logan, Jean; Shumay, Elena; Alia-Klein, Nelly; Wang, Gene-Jack; Volkow, Nora D.

    2015-03-01

    Monoamine oxidase (MAO) oxidizes amines from both endogenous and exogenous sources thereby regulating the concentration of neurotransmitter amines such as serot onin, norepinephrine and dopamine as well as many xenobiotics. MAO inhibitor drugs are used in the treatment of Parkinson’s disease and in depression stimulating the development of radiotracer tools to probe the role of MAO in normal human biology and in disease. Over the past 30 since the first radiotracers were developed and the first PET images of MAO in humans were carried out, PET studies of brain MAO in healthy volunteers and in patients have identified different variablesmore » which have contributed to different MAO levels in brain and in peripheral organs. MAO radiotracers and PET have also been used to study the current and developing MAO inhibitor drugs including the selection of doses for clinical trials. In this article, we describe (1) the development of MAO radiotracers; (2) human studies including the relationship of brain MAO levels to genotype, personality, neurological and psychiatric disorders; (3) examples of the use of MAO radiotracers in drug research and development. We will conclude with outstanding needs to improve the radiotracers which are currently used and possible new applications.« less

  11. Monoamine oxidase: Radiotracer chemistry and human studies

    SciTech Connect

    Fowler, Joanna S.; Logan, Jean; Shumay, Elena; Alia-Klein, Nelly; Wang, Gene-Jack; Volkow, Nora D.

    2015-03-01

    Monoamine oxidase (MAO) oxidizes amines from both endogenous and exogenous sources thereby regulating the concentration of neurotransmitter amines such as serot onin, norepinephrine and dopamine as well as many xenobiotics. MAO inhibitor drugs are used in the treatment of Parkinson’s disease and in depression stimulating the development of radiotracer tools to probe the role of MAO in normal human biology and in disease. Over the past 30 since the first radiotracers were developed and the first PET images of MAO in humans were carried out, PET studies of brain MAO in healthy volunteers and in patients have identified different variables which have contributed to different MAO levels in brain and in peripheral organs. MAO radiotracers and PET have also been used to study the current and developing MAO inhibitor drugs including the selection of doses for clinical trials. In this article, we describe (1) the development of MAO radiotracers; (2) human studies including the relationship of brain MAO levels to genotype, personality, neurological and psychiatric disorders; (3) examples of the use of MAO radiotracers in drug research and development. We will conclude with outstanding needs to improve the radiotracers which are currently used and possible new applications.

  12. An ultrafiltration assay for lysyl oxidase.

    PubMed

    Shackleton, D R; Hulmes, D J

    1990-03-01

    A modification of the original microdistillation assay for lysyl oxidase is described in which Amicon C-10 microconcentrators are used to separate, by ultrafiltration, the 3H-labeled products released from a [4,5-3H]-lysine-labeled elastin substrate. Enzyme activity is determined by scintillation counting of the ultrafiltrate, after subtraction of radioactivity released in the presence of beta-aminopropionitrile, a specific inhibitor of the enzyme. Conditions are described which optimize both the sensitivity and the efficient use of substrate. The assay shows linear inhibition of activity in up to 1 M urea; hence, as the enzyme is normally diluted in the assay, samples in 6 M urea can be assayed directly, without prior dialysis, and corrected for partial inhibition. Comparable results are obtained when enzyme activity is assayed by ultrafiltration or microdistillation. The assay is simple and convenient and, by using disposable containers throughout, it eliminates the need for time-consuming decontamination of radioactive glassware. PMID:1971160

  13. Bounds for nonlocality distillation protocols

    SciTech Connect

    Forster, Manuel

    2011-06-15

    Nonlocality can be quantified by the violation of a Bell inequality. Since this violation may be amplified by local operations, an alternative measure has been proposed--distillable nonlocality. The alternative measure is difficult to calculate exactly due to the double exponential growth of the parameter space. In this paper, we give a way to bound the distillable nonlocality of a resource by the solutions to a related optimization problem. Our upper bounds are exponentially easier to compute than the exact value and are shown to be meaningful in general and tight in some cases.

  14. A simple purification procedure of D-amino-acid oxidase from Candida guilliermondii H(see symbol)-4.

    PubMed

    Gevorgyan, G K; Davtyan, M A; Hambardzumyan, A A

    2012-01-01

    D-amino-acid oxidase (EC 1.4.3.3) was purified about 1480-fold from the yeast Candida guilliermondii H(see symbol)-4 using chromatofocusing method. The purification procedure gave an enzyme preparation which is greater than 90% homogenous on SDS-polyacrylamide gels with a specific activity of 11.54 U/mg at 30 degrees C with D-proline as substrate with the yield of total activity 9.3%. The molecular weights of subunit and native enzyme were determined to be 38.4 and 78.6 kDa by SDS-polyacrylamide gel electrophoresis and gel-filtration, respectively, suggesting that the native enzyme exists as a homodimer. A single molecular form with an isoelectric point of 6.85 was detected in analytical isoelectrofocusing. The optimum pH and temperature were 8.0 and 33 degrees C. An enzyme shows stability in the pH range from 7.4 to 9.0 and at the temperature no higher than 38 degrees C. Activation energy for D-amino-acid oxidase reaction was calculated to be 60 kJ/mol at 30 degrees C. The strict D-isomer specificity of the enzyme is confirmed, since no reaction could be detected with L-amino acids, and a large number of D-amino acids could be substrates for this enzyme. K(m) and V(max) values were determined for D-proline and D-alanine, which, among 22 tested, were the best substrates of the enzyme. D-amino-acid oxidase from the yeast C. guilliermondii is a flavoprotein oxidase in which the prosthetic group is tightly, but not covalently, bound FAD. The enzyme is completely inhibited by sodium benzoate, SH-oxidizing agents, but not by sodium azide, toluene or chloroform. PMID:23156699

  15. Deciphering the role of NADPH oxidase in complex interactions between maize (Zea mays L.) genotypes and cereal aphids.

    PubMed

    Sytykiewicz, Hubert

    2016-07-22

    Plant NADPH oxidases (NOXs) encompass a group of membrane-bound enzymes participating in formation of reactive oxygen species (ROS) under physiological conditions as well as in response to environmental stressors. The purpose of the survey was to unveil the role of NADPH oxidase in pro-oxidative responses of maize (Zea mays L.) seedling leaves exposed to cereal aphids' infestation. The impact of apteral females of bird cherry-oat aphid (Rhopalosiphum padi L.) and grain aphid (Sitobion avenae F.) feeding on expression levels of all four NADPH oxidase genes (rbohA, rbohB, rbohC, rbohD) and total activity of NOX enzyme in maize plants were investigated. In addition, inhibitory effect of diphenylene iodonium (DPI) pre-treatment on NOX activity and hydrogen peroxide content in aphid-stressed maize seedlings was studied. Leaf infestation biotests were accomplished on 14-day-old seedlings representing two aphid-resistant varieties (Ambrozja and Waza) and two aphid-susceptible ones (Tasty Sweet and Złota Karłowa). Insects' attack led to profound upregulation of rbohA and rbohD genes in tested host plants, lower elevations were noted in level of rbohB mRNA, whereas abundance of rbohC transcript was not significantly altered. It was uncovered aphid-induced enhancement of NOX activity in examined plants. Higher increases in expression of all investigated rboh genes and activity of NADPH oxidase occurred in tissues of more resistant maize cultivars than in susceptible ones. Furthermore, DPI treatment resulted in strong reduction of NOX activity and H2O2 accumulation in aphid-infested Z. mays plants, thus evidencing circumstantial role of the enzyme in insect-elicited ROS generation. PMID:27178208

  16. Effects of crystallization on the heme-carbon monoxide moiety of bovine heart cytochrome c oxidase carbonyl.

    PubMed Central

    Tsubaki, M; Shinzawa, K; Yoshikawa, S

    1992-01-01

    Cytochrome c oxidase isolated from bovine heart was crystallized in the fully reduced carbon monoxide (CO)-bound form. To evaluate the structure of the O2 reaction site in crystals and in solution, the bound C-O stretch infrared band in protein crystals was compared with the band for protein solution. In solution, the C-O stretch band could be deconvoluted into two extremely narrow bands, one at 1963.6 cm-1 with delta v1/2 = 3.4 cm-1 of 60% Gaussian/40% Lorentzian character represented 86% of the total band area and the other at 1960.3 cm-1 with delta v1/2 = 3.0 cm-1 of 47% Gaussian/53% Lorentzian character represented 14% of the total band area. The crystals exhibited two deconvoluted C-O infrared bands having very similar band parameters with those in solution. These findings support the presence of two structurally similar conformers in both crystals and solution. Thus crystallization of this enzyme does not affect the structure at the CO-binding site to as great extent as has been noted for myoglobin and hemoglobin carbonyls, indicating that the active (CO- or O2-binding) site of cytochrome c oxidase must be conformationally very stable and highly ordered compared to other hemoproteins such as hemoglobin. Images FIGURE 1 PMID:1336988

  17. New Functional Sulfide Oxidase-Oxygen Reductase Supercomplex in the Membrane of the Hyperthermophilic Bacterium Aquifex aeolicus*

    PubMed Central

    Prunetti, Laurence; Infossi, Pascale; Brugna, Myriam; Ebel, Christine; Giudici-Orticoni, Marie-Thérèse; Guiral, Marianne

    2010-01-01

    Aquifex aeolicus, a hyperthermophilic and microaerophilic bacterium, obtains energy for growth from inorganic compounds alone. It was previously proposed that one of the respiratory pathways in this organism consists of the electron transfer from hydrogen sulfide (H2S) to molecular oxygen. H2S is oxidized by the sulfide quinone reductase, a membrane-bound flavoenzyme, which reduces the quinone pool. We have purified and characterized a novel membrane-bound multienzyme supercomplex that brings together all the molecular components involved in this bioenergetic chain. Our results indicate that this purified structure consists of one dimeric bc1 complex (complex III), one cytochrome c oxidase (complex IV), and one or two sulfide quinone reductases as well as traces of the monoheme cytochrome c555 and quinone molecules. In addition, this work strongly suggests that the cytochrome c oxidase in the supercomplex is a ba3-type enzyme. The supercomplex has a molecular mass of about 350 kDa and is enzymatically functional, reducing O2 in the presence of the electron donor, H2S. This is the first demonstration of the existence of such a respirasome carrying a sulfide oxidase-oxygen reductase activity. Moreover, the kinetic properties of the sulfide quinone reductase change slightly when integrated in the supercomplex, compared with the free enzyme. We previously purified a complete respirasome involved in hydrogen oxidation and sulfur reduction from Aquifex aeolicus. Thus, two different bioenergetic pathways (sulfur reduction and sulfur oxidation) are organized in this bacterium as supramolecular structures in the membrane. A model for the energetic sulfur metabolism of Aquifex aeolicus is proposed. PMID:20971847

  18. An alcohol oxidase of Phanerochaete chrysosporium with a distinct glycerol oxidase activity.

    PubMed

    Linke, Diana; Lehnert, Nicole; Nimtz, Manfred; Berger, Ralf G

    2014-01-01

    An intracellular alcohol oxidase (AOX) was isolated from the white-rot basidiomycete Phanerochaete chrysosporium (Pch), grown on l-lactate induction medium, and purified to electrophoretic homogeneity. The dimeric protein consisted of two identical 75kDa subunits. The open reading frame of 1,956bp resulted in a monomer consisting of 651 amino acids. The enzyme showed a pI at 5.4, a pH optimum of 9, a temperature optimum at 50°C, possessed putative conserved domains of the GMC superfamily, a FAD binding domain, and showed up to 86% homology to alcohol oxidase sequences of Gloeophyllum trabeum and Coprinopsis cinerea. As was shown for the first time for an AOX from a basidiomycete, not only methanol, but also lower primary alcohols and glycerol were accepted as substrates. An assay based on aldehyde dehydrogenase confirmed d-glyceraldehyde as the product of the reaction. A bioprocess based on this enzyme could alleviate the problems associated with the huge side-stream of glycerol occurring during the manufacture of biodiesel, yielding the green oxidant hydrogen peroxide. PMID:24910330

  19. Wronskian Method for Bound States

    ERIC Educational Resources Information Center

    Fernandez, Francisco M.

    2011-01-01

    We propose a simple and straightforward method based on Wronskians for the calculation of bound-state energies and wavefunctions of one-dimensional quantum-mechanical problems. We explicitly discuss the asymptotic behaviour of the wavefunction and show that the allowed energies make the divergent part vanish. As illustrative examples we consider…

  20. Teacher Education in Outward Bound.

    ERIC Educational Resources Information Center

    Robinson, Richard A.

    A series of Outward Bound programs and experiences was planned for El Paso County, Colorado, school teachers to increase their awareness of their personal characteristics, especially those that might enhance learning on the part of their students. Part of the planning for the program involved a survey of county high school teachers, counselors,…

  1. Loosely-Bound Diatomic Molecules.

    ERIC Educational Resources Information Center

    Balfour, W. J.

    1979-01-01

    Discusses concept of covalent bonding as related to homonuclear diatomic molecules. Article draws attention to the existence of bound rare gas and alkaline earth diatomic molecules. Summarizes their molecular parameters and offers spectroscopic data. Strength and variation with distance of interatomic attractive forces is given. (Author/SA)

  2. Definition of the catalytic site of cytochrome c oxidase: specific ligands of heme a and the heme a3-CuB center.

    PubMed Central

    Shapleigh, J P; Hosler, J P; Tecklenburg, M M; Kim, Y; Babcock, G T; Gennis, R B; Ferguson-Miller, S

    1992-01-01

    The three-subunit aa3-type cytochrome c oxidase (EC 1.9.3.1) of Rhodobacter sphaeroides is structurally and functionally homologous to the more complex mitochondrial oxidase. The largest subunit, subunit I, is highly conserved and predicted to contain 12 transmembrane segments that provide all the ligands for three of the four metal centers: heme a, heme a3, and CuB. A variety of spectroscopic techniques identify these ligands as histidines. We have used site-directed mutagenesis to change all the conserved histidines within subunit I of cytochrome c oxidase from Rb. sphaeroides. Analysis of the membrane-bound and purified mutant proteins by optical absorption and resonance Raman spectroscopy indicates that His-102 and His-421 are the ligands of heme a, while His-284, His-333, His-334, and His-419 ligate the heme a3-CuB center. To satisfy this ligation assignment, helices II, VI, VII, and X, which contain these histidine residues, must be in close proximity. These data provide empirical evidence regarding the three-dimensional protein structure at the catalytic core of cytochrome c oxidase. Images PMID:1317571

  3. Research Update: Facile synthesis of CoFe2O4 nano-hollow spheres for efficient bilirubin adsorption

    NASA Astrophysics Data System (ADS)

    Rakshit, Rupali; Pal, Monalisa; Chaudhuri, Arka; Mandal, Madhuri; Mandal, Kalyan

    2015-11-01

    Herein, we report an unprecedented bilirubin (BR) adsorption efficiency of CoFe2O4 (CFO) nanostructures in contrast to the commercially available activated carbon and resin which are generally used for haemoperfusion and haemodialysis. We have synthesized CFO nanoparticles of diameter 100 nm and a series of nano-hollow spheres of diameter 100, 160, 250, and 350 nm using a simple template free solvothermal technique through proper variation of reaction time and capping agent, oleylamine (OLA), respectively, and carried out SiO2 coating by employing Stöber method. The comparative BR adsorption study of CFO and SiO2 coated CFO nanostructures indicates that apart from porosity and hollow configuration of nanostructures, the electrostatic affinity between anionic carboxyl group of BR and cationic amine group of OLA plays a significant role in adsorbing BR. Finally, we demonstrate that the BR adsorption capacity of the nanostructures can be tailored by varying the morphology as well as size of the nanostructures. We believe that our developed magnetic nanostructures could be considered as a potential material towards therapeutic applications against hyperbilirubinemia.

  4. Study of Model Systems for Bilirubin and Bilin Chromophores: Determination and Modification of Thermal and Photochemical Properties.

    PubMed

    García-Iriepa, Cristina; Ernst, Hanna A; Liang, Yu; Unterreiner, Andreas-Neil; Frutos, Luis Manuel; Sampedro, Diego

    2016-08-01

    Bilin chromophores and bilirubin are involved in relevant biological functions such as light perception in plants and as protective agents against Alzheimer and other diseases. Despite their extensive use, a deep rationalization of the main factors controlling the thermal and photochemical properties has not been performed yet, which in turn hampers further applications of these versatile molecules. In an effort to understand those factors and allow control of the relevant properties, a combined experimental and computational study has been carried out for diverse model systems to understand the interconversion between Z and E isomers. In this study, we have demonstrated the crucial role of steric hindrance and hydrogen-bond interactions in thermal stability and the ability to control them by designing novel compounds. We also determined several photochemical properties and studied the photodynamics of two model systems in more detail, observing a fast relaxation of the excited state shorter than 2 ps in both cases. Finally, the computational study allowed us to rationalize the experimental evidence. PMID:27391671

  5. Titania bound sodium titanate ion exchanger

    DOEpatents

    DeFilippi, Irene C. G.; Yates, Stephen Frederic; Shen, Jian-Kun; Gaita, Romulus; Sedath, Robert Henry; Seminara, Gary Joseph; Straszewski, Michael Peter; Anderson, David Joseph

    1999-03-23

    This invention is method for preparing a titania bound ion exchange composition comprising admixing crystalline sodium titanate and a hydrolyzable titanium compound and, thereafter drying the titania bound crystalline sodium titanate and subjecting the dried titania bound ion exchange composition to optional compaction and calcination steps to improve the physical strength of the titania bound composition.

  6. The effect of premature and delayed birth on the development of UDP-glucuronosyltransferase activities towards bilirubin, morphine and testosterone in the rat.

    PubMed Central

    Campbell, M T; Wishart, G J

    1980-01-01

    In the rat, UDP-glucuronosyltransferase activities towards bilirubin, morphine and testosterone increase markedly after normal or premature birth. This rapid development is superimposed upon a much slower maturation of activity which occurs in utero during the last 2 days of normal gestation and gestation when birth is delayed. Development of all three activities is similar under these different conditions, suggesting a common developmenpal regulatory mechanism. PMID:6769435

  7. Identification of a genetic alteration in the code for bilirubin UDP-glucuronosyltransferase in the UGT1 gene complex of a Crigler-Najjar type I patient.

    PubMed Central

    Ritter, J K; Yeatman, M T; Ferreira, P; Owens, I S

    1992-01-01

    Patients with Crigler-Najjar syndrome (CN) type I inherit an autosomal recessive trait for hyperbilirubinemia, which is characterized by the total absence of bilirubin UDP-glucuronosyltransferase (transferase) activity. The recent identification of two bilirubin transferase isoforms with identical carboxyl termini (Ritter, J. K., J. M. Crawford, and I. S. Owens. 1991. J. Biol. Chem. 266:1043-1047) led to the discovery of a unique locus, UGT1, which encodes a family of UDP-glucuronosyltransferase isozymes, including the two bilirubin forms (Ritter, J. K., F. Chen, Y. Y. Sheen, H. M. Tran, S. Kimura, M. T. Yeatman, and I. S. Owens. 1992. J. Biol. Chem. 267:3257-3261). The UGT1 locus features a complex of six overlapping transcriptional units encoding transferases, each of which shares the four most 3' exons (2, 3, 4, and 5) specifying the 3' half of the transferase coding regions (condons 289-533) and the entire 3' untranslated region of each mRNA. This gene model predicts that a single critical mutation in any of these four "common" exons may inactivate the entire family of encoded transferases. In agreement with this prediction, we show here that in the first CN type I individual analyzed (patient F.B.), a 13-bp deletion has occurred in exon 2. Analysis of product generated by the polymerase chain reaction and genomic DNA demonstrated that F.B. is homozygous for the defective allele (UGT1*FB), and that the consanguineous parents are both heterozygotic at this locus. The mutation is predicted to result in the synthesis of severely truncated bilirubin transferase isozymes that are lacking a highly conserved sequence in the carboxyl-terminus and the characteristic membrane (endoplasmic reticulum)-anchoring segment of the protein molecule. Images PMID:1634606

  8. Monoamine oxidase and agitation in psychiatric patients.

    PubMed

    Nikolac Perkovic, Matea; Svob Strac, Dubravka; Nedic Erjavec, Gordana; Uzun, Suzana; Podobnik, Josip; Kozumplik, Oliver; Vlatkovic, Suzana; Pivac, Nela

    2016-08-01

    Subjects with schizophrenia or conduct disorder display a lifelong pattern of antisocial, aggressive and violent behavior and agitation. Monoamine oxidase (MAO) is an enzyme involved in the degradation of various monoamine neurotransmitters and neuromodulators and therefore has a role in various psychiatric and neurodegenerative disorders and pathological behaviors. Platelet MAO-B activity has been associated with psychopathy- and aggression-related personality traits, while variants of the MAOA and MAOB genes have been associated with diverse clinical phenotypes, including aggressiveness, antisocial problems and violent delinquency. The aim of the study was to evaluate the association of platelet MAO-B activity, MAOB rs1799836 polymorphism and MAOA uVNTR polymorphism with severe agitation in 363 subjects with schizophrenia and conduct disorder. The results demonstrated significant association of severe agitation and smoking, but not diagnosis or age, with platelet MAO-B activity. Higher platelet MAO-B activity was found in subjects with severe agitation compared to non-agitated subjects. Platelet MAO-B activity was not associated with MAOB rs1799836 polymorphism. These results suggested the association between increased platelet MAO-B activity and severe agitation. No significant association was found between severe agitation and MAOA uVNTR or MAOB rs1799836 polymorphism, revealing that these individual polymorphisms in MAO genes are not related to severe agitation in subjects with schizophrenia and conduct disorder. As our study included 363 homogenous Caucasian male subjects, our data showing this negative genetic association will be a useful addition to future meta-analyses. PMID:26851573

  9. Structure and function of mammalian aldehyde oxidases.

    PubMed

    Terao, Mineko; Romão, Maria João; Leimkühler, Silke; Bolis, Marco; Fratelli, Maddalena; Coelho, Catarina; Santos-Silva, Teresa; Garattini, Enrico

    2016-04-01

    Mammalian aldehyde oxidases (AOXs; EC1.2.3.1) are a group of conserved proteins belonging to the family of molybdo-flavoenzymes along with the structurally related xanthine dehydrogenase enzyme. AOXs are characterized by broad substrate specificity, oxidizing not only aromatic and aliphatic aldehydes into the corresponding carboxylic acids, but also hydroxylating a series of heteroaromatic rings. The number of AOX isoenzymes expressed in different vertebrate species is variable. The two extremes are represented by humans, which express a single enzyme (AOX1) in many organs and mice or rats which are characterized by tissue-specific expression of four isoforms (AOX1, AOX2, AOX3, and AOX4). In vertebrates each AOX isoenzyme is the product of a distinct gene consisting of 35 highly conserved exons. The extant species-specific complement of AOX isoenzymes is the result of a complex evolutionary process consisting of a first phase characterized by a series of asynchronous gene duplications and a second phase where the pseudogenization and gene deletion events prevail. In the last few years remarkable advances in the elucidation of the structural characteristics and the catalytic mechanisms of mammalian AOXs have been made thanks to the successful crystallization of human AOX1 and mouse AOX3. Much less is known about the physiological function and physiological substrates of human AOX1 and other mammalian AOX isoenzymes, although the importance of these proteins in xenobiotic metabolism is fairly well established and their relevance in drug development is increasing. This review article provides an overview and a discussion of the current knowledge on mammalian AOX. PMID:26920149

  10. Monoamine oxidase inhibitory activities of heterocyclic chalcones.

    PubMed

    Minders, Corné; Petzer, Jacobus P; Petzer, Anél; Lourens, Anna C U

    2015-11-15

    Studies have shown that natural and synthetic chalcones (1,3-diphenyl-2-propen-1-ones) possess monoamine oxidase (MAO) inhibition activities. Of particular importance to the present study is a report that a series of furanochalcones acts as MAO-B selective inhibitors. Since the effect of heterocyclic substitution, other than furan (and more recently thiophene, piperidine and quinoline) on the MAO inhibitory properties of the chalcone scaffold remains unexplored, the aim of this study was to synthesise and evaluate further heterocyclic chalcone analogues as inhibitors of the human MAOs. For this purpose, heterocyclic chalcone analogues that incorporate pyrrole, 5-methylthiophene, 5-chlorothiophene and 6-methoxypyridine substitution were examined. Seven of the nine synthesised compounds exhibited IC50 values <1 μM for the inhibition of MAO-B, with all compounds exhibiting higher affinities for MAO-B compared to the MAO-A isoform. The most potent MAO-B inhibitor (4h) displays an IC50 value of 0.067 μM while the most potent MAO-A inhibitor (4e) exhibits an IC50 value of 3.81 μM. It was further established that selected heterocyclic chalcones are reversible and competitive MAO inhibitors. 4h, however, may exhibit tight-binding to MAO-B, a property linked to its thiophene moiety. We conclude that high potency chalcones such as 4h represent suitable leads for the development of MAO-B inhibitors for the treatment of Parkinson's disease and possibly other neurodegenerative disorders. PMID:26432037

  11. Role of NADPH Oxidases in Liver Fibrosis

    PubMed Central

    Paik, Yong-Han; Kim, Jonghwa; Aoyama, Tomonori; De Minicis, Samuele; Bataller, Ramon

    2014-01-01

    Abstract Significance: Hepatic fibrosis is the common pathophysiologic process resulting from chronic liver injury, characterized by the accumulation of an excessive extracellular matrix. Multiple lines of evidence indicate that oxidative stress plays a pivotal role in the pathogenesis of liver fibrosis. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) is a multicomponent enzyme complex that generates reactive oxygen species (ROS) in response to a wide range of stimuli. In addition to phagocytic NOX2, there are six nonphagocytic NOX proteins. Recent Advances: In the liver, NOX is functionally expressed both in the phagocytic form and in the nonphagocytic form. NOX-derived ROS contributes to various kinds of liver disease caused by alcohol, hepatitis C virus, and toxic bile acids. Recent evidence indicates that both phagocytic NOX2 and nonphagocytic NOX isoforms, including NOX1 and NOX4, mediate distinct profibrogenic actions in hepatic stellate cells, the main fibrogenic cell type in the liver. The critical role of NOX in hepatic fibrogenesis provides a rationale to assess pharmacological NOX inhibitors that treat hepatic fibrosis in patients with chronic liver disease. Critical Issues: Although there is compelling evidence indicating a crucial role for NOX-mediated ROS generation in hepatic fibrogenesis, little is known about the expression, subcellular localization, regulation, and redox signaling of NOX isoforms in specific cell types in the liver. Moreover, the exact mechanism of NOX-mediated fibrogenic signaling is still largely unknown. Future Directions: A better understanding through further research about NOX-mediated fibrogenic signaling may enable the development of novel anti-fibrotic therapy using NOX inhibition strategy. Antioxid. Redox Signal. 20, 2854–2872. PMID:24040957

  12. Transversely bounded DFB lasers. [bounded distributed-feedback lasers

    NASA Technical Reports Server (NTRS)

    Elachi, C.; Evans, G.; Yeh, C.

    1975-01-01

    Bounded distributed-feedback (DFB) lasers are studied in detail. Threshold gain and field distribution for a number of configurations are derived and analyzed. More specifically, the thin-film guide, fiber, diffusion guide, and hollow channel with inhomogeneous-cladding DFB lasers are considered. Optimum points exist and must be used in DFB laser design. Different-modes feedback and the effects of the transverse boundaries are included. A number of applications are also discussed.

  13. Does Serum Bilirubin level on admission predict TIMI flow grade and in-hospital MACE in patients with STEMI undergoing primary PCI.

    PubMed

    Celik, Turgay; Kaya, Mehmet G; Akpek, Mahmut; Yarlioglues, Mikail; Sarli, Bahadir; Topsakal, Ramazan; Gibson, Charles Michael

    2014-03-01

    We evaluated the association of total bilirubin with post-percutaneous coronary intervention (PCI) coronary blood flow and in-hospital major adverse cardiac events (MACEs) in patients with ST-segment elevation myocardial infarction (STEMI) undergoing primary PCI. A total of 536 consecutive patients with STEMI (male 79%, mean age = 59.9 ± 12.6 years) admitted within 6 hours from symptom onset were enrolled. Patients were divided into 2 groups based on the thrombolysis in myocardial infarction (MI) flow grade. In-stent thrombosis, nonfatal MI, and in-hospital mortality were significantly higher in no-reflow group (P = .007, P = .002, and P < .001, respectively). On multivariate regression, the total bilirubin levels remained independent predictors of no-reflow (odds ratio [OR] 1.586, 95% confidence interval [CI] 1.02-2.47; P = .042) and in-hospital MACE (OR 1.399, 95% CI 1.053-1.857; P = .020). Serum bilirubin levels were independently associated with no-reflow and in-hospital MACE in patients with STEMI undergoing primary PCI. PMID:23378197

  14. Terminal Oxidases of Bacillus subtilis Strain 168: One Quinol Oxidase, Cytochrome aa3 or Cytochrome bd, Is Required for Aerobic Growth

    PubMed Central

    Winstedt, Lena; von Wachenfeldt, Claes

    2000-01-01

    The gram-positive endospore-forming bacterium Bacillus subtilis has, under aerobic conditions, a branched respiratory system comprising one quinol oxidase branch and one cytochrome oxidase branch. The system terminates in one of four alternative terminal oxidases. Cytochrome caa3 is a cytochrome c oxidase, whereas cytochrome bd and cytochrome aa3 are quinol oxidases. A fourth terminal oxidase, YthAB, is a putative quinol oxidase predicted from DNA sequence analysis. None of the terminal oxidases are, by themselves, essential for growth. However, one quinol oxidase (cytochrome aa3 or cytochrome bd) is required for aerobic growth of B. subtilis strain 168. Data indicating that cytochrome aa3 is the major oxidase used by exponentially growing cells in minimal and rich medium are presented. We show that one of the two heme-copper oxidases, cytochrome caa3 or cytochrome aa3, is required for efficient sporulation of B. subtilis strain 168 and that deletion of YthAB in a strain lacking cytochrome aa3 makes the strain sporulation deficient. PMID:11073895

  15. Crystal Structure of Alcohol Oxidase from Pichia pastoris

    PubMed Central

    Valerius, Oliver; Feussner, Ivo; Ficner, Ralf

    2016-01-01

    FAD-dependent alcohol oxidases (AOX) are key enzymes of methylotrophic organisms that can utilize lower primary alcohols as sole source of carbon and energy. Here we report the crystal structure analysis of the methanol oxidase AOX1 from Pichia pastoris. The crystallographic phase problem was solved by means of Molecular Replacement in combination with initial structure rebuilding using Rosetta model completion and relaxation against an averaged electron density map. The subunit arrangement of the homo-octameric AOX1 differs from that of octameric vanillyl alcohol oxidase and other dimeric or tetrameric alcohol oxidases, due to the insertion of two large protruding loop regions and an additional C-terminal extension in AOX1. In comparison to other alcohol oxidases, the active site cavity of AOX1 is significantly reduced in size, which could explain the observed preference for methanol as substrate. All AOX1 subunits of the structure reported here harbor a modified flavin adenine dinucleotide, which contains an arabityl chain instead of a ribityl chain attached to the isoalloxazine ring. PMID:26905908

  16. NADPH OXIDASE: STRUCTURE AND ACTIVATION MECHANISMS (REVIEW). NOTE I.

    PubMed

    Filip-Ciubotaru, Florina; Manciuc, Carmen; Stoleriu, Gabriela; Foia, Liliana

    2016-01-01

    NADPH oxidase (nicotinamide adenine dinucleotide phosphate-oxidase), with its generically termed NOX isoforms, is the major source of ROS (reactive oxigen species) in biological systems. ROS are small oxygen-derived molecules with an important role in various biological processes (physiological or pathological). If under physiological conditions some processes are beneficial and necessary for life, under pathophysiological conditions they are noxious, harmful. NADPH oxidases are present in phagocytes and in a wide variety of nonphagocytic cells. The enzyme generates superoxide by transferring electrons from NADPH inside the cell across the membrane and coupling them to molecular oxygen to produce superoxide anion, a reactive free-radical. Structurally, NADPH oxidase is a multicomponent enzyme which includes two integral membrane proteins, glycoprotein gp9 1 Phox and adaptor protein p22(phox), which together form the heterodimeric flavocytochrome b558 that constitutes the core of the enzyme. During the resting state, the multidomain regulatory subunits p40P(phox), p47(phox), p67(Phox) are located in the cytosol organized as a complex. The activation of phagocytic NADPH oxidase occurs through a complex series of protein interactions. PMID:27125069

  17. Sulfide inhibition of and metabolism by cytochrome c oxidase.

    PubMed

    Nicholls, Peter; Marshall, Doug C; Cooper, Chris E; Wilson, Mike T

    2013-10-01

    Hydrogen sulfide (H2S), a classic cytochrome c oxidase inhibitor, is also an in vitro oxidase substrate and an in vivo candidate hormonal ('gasotransmitter') species affecting sleep and hibernation. H2S, nitric oxide (NO) and carbon monoxide (CO) share some common features. All are low-molecular-mass physiological effectors and also oxidase inhibitors, capable of binding more than one enzyme site, and each is an oxidizable 'substrate'. The oxidase oxidizes CO to CO2, NO to nitrite and sulfide to probable persulfide species. Mitochondrial cytochrome c oxidase in an aerobic steady state with ascorbate and cytochrome c is rapidly inhibited by sulfide in a biphasic manner. At least two successive inhibited species are involved, probably partially reduced. The oxidized enzyme, in the absence of turnover, occurs in at least two forms: the 'pulsed' and 'resting' states. The pulsed form reacts aerobically with sulfide to form two intermediates, 'P' and 'F', otherwise involved in the reaction of oxygen with reduced enzyme. Sulfide can directly reduce the oxygen-reactive a3CuB binuclear centre in the pulsed state. The resting enzyme does not undergo such a step, but only a very slow one-electron reduction of the electron-transferring haem a. In final reactivation phases, both the steady-state inhibition of catalysis and the accumulation of P and F states are reversed by slow sulfide oxidation. A model for this complex reaction pattern is presented. PMID:24059525

  18. Crystal Structure of Alcohol Oxidase from Pichia pastoris.

    PubMed

    Koch, Christian; Neumann, Piotr; Valerius, Oliver; Feussner, Ivo; Ficner, Ralf

    2016-01-01

    FAD-dependent alcohol oxidases (AOX) are key enzymes of methylotrophic organisms that can utilize lower primary alcohols as sole source of carbon and energy. Here we report the crystal structure analysis of the methanol oxidase AOX1 from Pichia pastoris. The crystallographic phase problem was solved by means of Molecular Replacement in combination with initial structure rebuilding using Rosetta model completion and relaxation against an averaged electron density map. The subunit arrangement of the homo-octameric AOX1 differs from that of octameric vanillyl alcohol oxidase and other dimeric or tetrameric alcohol oxidases, due to the insertion of two large protruding loop regions and an additional C-terminal extension in AOX1. In comparison to other alcohol oxidases, the active site cavity of AOX1 is significantly reduced in size, which could explain the observed preference for methanol as substrate. All AOX1 subunits of the structure reported here harbor a modified flavin adenine dinucleotide, which contains an arabityl chain instead of a ribityl chain attached to the isoalloxazine ring. PMID:26905908

  19. Bone marrow purging by a xanthine oxidase-antibody conjugate.

    PubMed

    Dinota, A; Tazzari, P L; Abbondanza, A; Battelli, M G; Gobbi, M; Stirpe, F

    1990-07-01

    The selective cytotoxicity of the xanthine oxidase conjugated to an 8A monoclonal antibody recognizing a human plasma cell-associated antigen has been described. The selectivity and the toxicity of the hypoxanthine/conjugated xanthine oxidase system was increased by removing the excess of conjugate and by adding chelated iron. Under these experimental conditions the cytotoxicity of the conjugate exceeded that of free xanthine oxidase by one order of magnitude. The conjugate effectively purged bone marrow from infiltrating neoplastic plasma cells and added target Raji cells, provided blood was removed and bone marrow peroxidases were exhausted. In conditions of purging effectiveness the conjugate had no toxicity to CFU-GM. No toxicity to mice was observed after i.v. injection of xanthine oxidase-antibody conjugate up to 2.9 U/kg body weight. Thus the hypoxanthine/conjugated xanthine oxidase system could be an effective and nontoxic tool for the ex vivo bone marrow purging in multiple myeloma patients for autologous transplantation. PMID:2390631

  20. Modifications of the Griesmer bound

    NASA Technical Reports Server (NTRS)

    Mceliece, R. J.; Solomon, G.

    1991-01-01

    The Griesmer bound is a classical technique (developed in 1960) for estimating the minimum length n required for a binary linear code with a given dimension k and minimum distance d. In this article, a unified derivation of the Griesmer bound and two new variations on it are presented. The first variation deals with linear codes which contain the all-ones vector; such codes are quite common and are useful in practice because of their 'transparent' properties. The second variation deals with codes that are constrained to contain a word of weight greater than or equal to M. In both cases these constraints (the all-ones word or a word of high weight) can increase the minimum length of a code with given k and d.

  1. Semiclassical bounds in magnetic bottles

    NASA Astrophysics Data System (ADS)

    Barseghyan, Diana; Exner, Pavel; Kovařík, Hynek; Weidl, Timo

    2016-02-01

    The aim of the paper is to derive spectral estimates into several classes of magnetic systems. They include three-dimensional regions with Dirichlet boundary as well as a particle in ℝ3 confined by a local change of the magnetic field. We establish two-dimensional Berezin-Li-Yau and Lieb-Thirring-type bounds in the presence of magnetic fields and, using them, get three-dimensional estimates for the eigenvalue moments of the corresponding magnetic Laplacians.

  2. Bilirubin UDP-Glucuronosyltransferase 1A1 (UGT1A1) Gene Promoter Polymorphisms and HPRT, Glycophorin A, and Micronuclei Mutant Frequencies in Human Blood

    SciTech Connect

    Grant, D; Hall, I J; Eastmond, D; Jones, I M; Bell, D A

    2004-10-06

    A dinucleotide repeat polymorphism (5-, 6-, 7-, or 8-TA units) has been identified within the promoter region of UDP-glucuronosyltransferase 1A1 gene (UGT1A1). The 7-TA repeat allele has been associated with elevated serum bilirubin levels that cause a mild hyperbilirubinemia (Gilbert's syndrome). Studies suggest that promoter transcriptional activity of UGT1A1 is inversely related to the number of TA repeats and that unconjugated bilirubin concentration increases directly with the number of TA repeat elements. Because bilirubin is a known antioxidant, we hypothesized that UGT1A1 repeats associated with higher bilirubin may be protective against oxidative damage. We examined the effect of UGT1A1 genotype on somatic mutant frequency in the hypoxanthine-guanine phosphoribosyl-transferase (HPRT) gene in human lymphocytes and the glycophorin A (GPA) gene of red blood cells (both N0, NN mutants), and the frequency of lymphocyte micronuclei (both kinetochore (K) positive or micronuclei K negative) in 101 healthy smoking and nonsmoking individuals. As hypothesized, genotypes containing 7-TA and 8-TA displayed marginally lower GPA{_}NN mutant frequency relative to 5/5, 5/6, 6/6 genotypes (p<0.05). In contrast, our analysis showed that lower expressing UGT1A1 alleles (7-TA and 8-TA) were associated with modestly increased HPRT mutation frequency (p<0.05) while the same low expression genotypes were not significantly associated with micronuclei frequencies (K-positive or K-negative) when compared to high expression genotypes (5-TA and 6-TA). We found weak evidence that UGT1A1 genotypes containing 7-TA and 8-TA were associated with increased GPA{_}N0 mutant frequency relative to 5/5, 5/6, 6/6 genotypes (p<0.05). These data suggest that UGT1A1 genotype may modulate somatic mutation of some types, in some cell lineages, by a mechanism not involving bilirubin antioxidant activity. More detailed studies examining UGT1A1 promoter variation, oxidant/antioxidant balance and genetic

  3. Regulation of Ascorbate Oxidase Expression in Pumpkin by Auxin and Copper 1

    PubMed Central

    Esaka, Muneharu; Fujisawa, Kouichi; Goto, Miwa; Kisu, Yasutomo

    1992-01-01

    Ascorbate oxidase expression in pumpkin (Cucurbita spp.) tissues was studied. Specific ascorbate oxidase activities in pumpkin leaf and stem tissues were about 2 and 1.5 times that in the fruit tissues, respectively. In seeds, little ascorbate oxidase activity was detected. Northern blot analyses showed an abundant ascorbate oxidase mRNA in leaf and stem tissues. Fruit tissues had lower levels of ascorbate oxidase mRNA than leaf and stem tissues. Ascorbate oxidase mRNA was not detected in seeds. Specific ascorbate oxidase activity gradually increased during early seedling growth of pumpkin seeds. The increase was accompanied by an increase in ascorbate oxidase mRNA. When ascorbate oxidase activity in developing pumpkin fruits was investigated, the activities in immature fruits that are rapidly growing at 0, 2, 4, and 7 d after anthesis were much higher than those in mature fruits at 14 and 30 d after anthesis. The specific activity and mRNA of ascorbate oxidase markedly increased after inoculation of pumpkin fruit tissues into Murashige and Skoog's culture medium in the presence of an auxin such as 2,4-dichlorophenoxyacetic acid (2,4-D) but not in the absence of 2,4-D. In the presence of 10 mg/L of 2,4-D, ascorbate oxidase mRNA was the most abundant. Thus, ascorbate oxidase is induced by 2,4-D. These results indicate that ascorbate oxidase is involved in cell growth. In pumpkin callus, ascorbate oxidase activity could be markedly increased by adding copper. Furthermore, immunological blotting showed that the amount of ascorbate oxidase protein was also increased by adding copper. However, northern blot analyses showed that ascorbate oxidase mRNA was not increased by adding copper. We suggest that copper may control ascorbate oxidase expression at translation or at a site after translation. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:16652952

  4. Regulation of ascorbate oxidase expression in pumpkin by auxin and copper.

    PubMed

    Esaka, M; Fujisawa, K; Goto, M; Kisu, Y

    1992-09-01

    Ascorbate oxidase expression in pumpkin (Cucurbita spp.) tissues was studied. Specific ascorbate oxidase activities in pumpkin leaf and stem tissues were about 2 and 1.5 times that in the fruit tissues, respectively. In seeds, little ascorbate oxidase activity was detected. Northern blot analyses showed an abundant ascorbate oxidase mRNA in leaf and stem tissues. Fruit tissues had lower levels of ascorbate oxidase mRNA than leaf and stem tissues. Ascorbate oxidase mRNA was not detected in seeds. Specific ascorbate oxidase activity gradually increased during early seedling growth of pumpkin seeds. The increase was accompanied by an increase in ascorbate oxidase mRNA. When ascorbate oxidase activity in developing pumpkin fruits was investigated, the activities in immature fruits that are rapidly growing at 0, 2, 4, and 7 d after anthesis were much higher than those in mature fruits at 14 and 30 d after anthesis. The specific activity and mRNA of ascorbate oxidase markedly increased after inoculation of pumpkin fruit tissues into Murashige and Skoog's culture medium in the presence of an auxin such as 2,4-dichlorophenoxyacetic acid (2,4-D) but not in the absence of 2,4-D. In the presence of 10 mg/L of 2,4-D, ascorbate oxidase mRNA was the most abundant. Thus, ascorbate oxidase is induced by 2,4-D. These results indicate that ascorbate oxidase is involved in cell growth. In pumpkin callus, ascorbate oxidase activity could be markedly increased by adding copper. Furthermore, immunological blotting showed that the amount of ascorbate oxidase protein was also increased by adding copper. However, northern blot analyses showed that ascorbate oxidase mRNA was not increased by adding copper. We suggest that copper may control ascorbate oxidase expression at translation or at a site after translation. PMID:16652952

  5. 78 FR 18326 - Agency Information Collection Activities; Comment Request; Upward Bound and Upward Bound Math...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-03-26

    ... Agency Information Collection Activities; Comment Request; Upward Bound and Upward Bound Math Science... Upward Bound Math Science Annual Performance Report. OMB Control Number: 1840-NEW. Type of Review: New... under the regular Upward Bound (UB) and Upward Bound Math and Science (UBMS) Programs. The Department...

  6. Hunting η-bound nuclei

    NASA Astrophysics Data System (ADS)

    Machner, H.

    2010-09-01

    The η meson can be bound to atomic nuclei. Experimental search is discussed in the form of final state interaction for the reactions dp → 3Heη and dd → 4Heη. For the latter case tensor polarized deuterons were used in order to extract the s-wave strength. For both reactions complex scattering lengths are deduced: a3Heη = [± (10.7 ± 0.8+0.1-0.5) + i. (1.5 ± 2.6 +1.0-0.9)] fm and a4Heη = [±(3.1 ±0.5) + i. (0 ±0.5)] fm. In a two-nucleon transfer reaction under quasi-free conditions, p27 Al → 3HeX, was investigated. The system X can be the bound 25Mgotimesη at rest. When a possible decay of an intermediate N* (1535) is required, a highly significant bump shows up in the missing mass spectrum. The data give for a bound state a binding energy of 13.3 ±1.6 MeV and a width of σ = 4.4±1.3 MeV.

  7. Expression of alternative oxidase in Drosophila ameliorates diverse phenotypes due to cytochrome oxidase deficiency

    PubMed Central

    Kemppainen, Kia K.; Rinne, Juho; Sriram, Ashwin; Lakanmaa, Matti; Zeb, Akbar; Tuomela, Tea; Popplestone, Anna; Singh, Satpal; Sanz, Alberto; Rustin, Pierre; Jacobs, Howard T.

    2014-01-01

    Mitochondrial dysfunction is a significant factor in human disease, ranging from systemic disorders of childhood to cardiomyopathy, ischaemia and neurodegeneration. Cytochrome oxidase, the terminal enzyme of the mitochondrial respiratory chain, is a frequent target. Lower eukaryotes possess alternative respiratory-chain enzymes that provide non-proton-translocating bypasses for respiratory complexes I (single-subunit reduced nicotinamide adenine dinucleotide dehydrogenases, e.g. Ndi1 from yeast) or III + IV [alternative oxidase (AOX)], under conditions of respiratory stress or overload. In previous studies, it was shown that transfer of yeast Ndi1 or Ciona intestinalis AOX to Drosophila was able to overcome the lethality produced by toxins or partial knockdown of complex I or IV. Here, we show that AOX can provide a complete or substantial rescue of a range of phenotypes induced by global or tissue-specific knockdown of different cIV subunits, including integral subunits required for catalysis, as well as peripheral subunits required for multimerization and assembly. AOX was also able to overcome the pupal lethality produced by muscle-specific knockdown of subunit CoVb, although the rescued flies were short lived and had a motility defect. cIV knockdown in neurons was not lethal during development but produced a rapidly progressing locomotor and seizure-sensitivity phenotype, which was substantially alleviated by AOX. Expression of Ndi1 exacerbated the neuronal phenotype produced by cIV knockdown. Ndi1 expressed in place of essential cI subunits produced a distinct residual phenotype of delayed development, bang sensitivity and male sterility. These findings confirm the potential utility of alternative respiratory chain enzymes as tools to combat mitochondrial disease, while indicating important limitations thereof. PMID:24293544

  8. Polyamine Oxidase5 Regulates Arabidopsis Growth through Thermospermine Oxidase Activity1[C][W

    PubMed Central

    Kim, Dong Wook; Watanabe, Kanako; Murayama, Chihiro; Izawa, Sho; Niitsu, Masaru; Michael, Anthony J.; Berberich, Thomas; Kusano, Tomonobu

    2014-01-01

    The major plant polyamines (PAs) are the tetraamines spermine (Spm) and thermospermine (T-Spm), the triamine spermidine, and the diamine putrescine. PA homeostasis is governed by the balance between biosynthesis and catabolism; the latter is catalyzed by polyamine oxidase (PAO). Arabidopsis (Arabidopsis thaliana) has five PAO genes, AtPAO1 to AtPAO5, and all encoded proteins have been biochemically characterized. All AtPAO enzymes function in the back-conversion of tetraamine to triamine and/or triamine to diamine, albeit with different PA specificities. Here, we demonstrate that AtPAO5 loss-of-function mutants (pao5) contain 2-fold higher T-Spm levels and exhibit delayed transition from vegetative to reproductive growth compared with that of wild-type plants. Although the wild type and pao5 are indistinguishable at the early seedling stage, externally supplied low-dose T-Spm, but not other PAs, inhibits aerial growth of pao5 mutants in a dose-dependent manner. Introduction of wild-type AtPAO5 into pao5 mutants rescues growth and reduces the T-Spm content, demonstrating that AtPAO5 is a T-Spm oxidase. Recombinant AtPAO5 catalyzes the conversion of T-Spm and Spm to triamine spermidine in vitro. AtPAO5 specificity for T-Spm in planta may be explained by coexpression with T-Spm synthase but not with Spm synthase. The pao5 mutant lacking T-Spm oxidation and the acl5 mutant lacking T-Spm synthesis both exhibit growth defects. This study indicates a crucial role for T-Spm in plant growth and development. PMID:24906355

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

    SciTech Connect

    Pan, Heng-Yen; Whittaker, Mei M.; Bouveret, Romaric; Berna, Anne; Bernier, Francois; Whittaker, James W. . E-mail: jim@ebs.ogi.edu

    2007-05-18

    High-level secretory expression of wheat (Triticum aestivum) germin/oxalate oxidase was achieved in Pichia pastoris fermentation cultures as an {alpha}-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{sup 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.

  10. X-ray absorption spectroscopy of chicken sulfite oxidase crystals

    SciTech Connect

    George, G.N.; Pickering, I.J.; Kisker, C.

    1999-05-17

    Sulfite oxidase catalyzes the physiologically vital oxidation of sulfite to sulfate. Recently, the crystal structure of chicken sulfite oxidase has been reported at 1.9 {angstrom} resolution. In contrast to the information available from previous X-ray absorption spectroscopic studies, the active site indicated by crystallography was a mono-oxo species. Because of this the possibility that the crystals did in fact contain a reduced molybdenum species was considered in the crystallographic work. The authors report herein an X-ray absorption spectroscopic study of polycrystalline sulfite oxidase prepared in the same manner as the previous single-crystal samples, and compare this with data for frozen solutions of oxidized and reduced enzyme.

  11. Protoporphyrinogen oxidase as a molecular target for diphenyl ether herbicides.

    PubMed Central

    Matringe, M; Camadro, J M; Labbe, P; Scalla, R

    1989-01-01

    Diphenyl ether herbicides induce an accumulation of protoporphyrin IX in plant tissues. By analogy to human porphyria, the accumulation could be attributed to decreased (Mg or Fe)-chelatase or protoporphyrinogen oxidase activities. Possible effects of acifluorfen-methyl on these enzymes were investigated in isolated corn (maize, Zea mays) etioplasts, potato (Solanum tuberosum) and mouse mitochondria, and yeast mitochondrial membranes. Acifluorfen-methyl was strongly inhibitory to protoporphyrinogen oxidase activities whatever their origins [concn. causing 50% inhibition (IC50) = 4 nM for the corn etioplast enzyme]. By contrast, it was roughly 100,000 times less active on (Mg or Fe)-chelatase activities (IC50 = 80-100 microM). Our results lead us to propose protoporphyrinogen oxidase as a cellular target for diphenyl ether herbicides. PMID:2775186

  12. [Amperometric enzyme biosensor with a glucose oxidase-polyaniline membrane].

    PubMed

    Dziadevich, S V; Doldatkin, A P; Rossokhatyĭ, V K; Shram, N F; Shul'ga, A A; Strikha, V I

    1994-01-01

    An amperometric glucose biosensor was made by electrochemical polymerization of aniline onto the gold electrodes in presence of the enzyme glucose oxidase in the phosphate buffer solution with pH 7.0. Aniline is easily polymerized forming a thin film, which adheres tightly on the electrodes surface. During the electropolymerization process glucose oxidase was entrapped into polyaniline film which then became the catalyst of the enzyme reaction of glucose hydrolysis. Experiments were performed to determine optimal conditions of polyaniline-glucose oxidase film preparation. Glucose was amperometrically determined with the electrochemically fabricated biosensor in the concentration range 10(-4) M to 2 x 10(-2) M. The linearity of the enzyme electrode response ranged from 2 x 10(-4) M to 6 x 10(-3) M. The electrochemical synthesis of a polyaniline-enzyme thin film a high-technologic one and this permits fabricating various microbiosensors and multisensors in the continuous technological cycle. PMID:7754558

  13. Hydrophilic bile acids protect human blood-brain barrier endothelial cells from disruption by unconjugated bilirubin: an in vitro study

    PubMed Central

    Palmela, Inês; Correia, Leonor; Silva, Rui F. M.; Sasaki, Hiroyuki; Kim, Kwang S.; Brites, Dora; Brito, Maria A.

    2015-01-01

    Ursodeoxycholic acid and its main conjugate glycoursodeoxycholic acid are bile acids with neuroprotective properties. Our previous studies demonstrated their anti-apoptotic, anti-inflammatory, and antioxidant properties in neural cells exposed to elevated levels of unconjugated bilirubin (UCB) as in severe jaundice. In a simplified model of the blood-brain barrier, formed by confluent monolayers of a cell line of human brain microvascular endothelial cells, UCB has shown to induce caspase-3 activation and cell death, as well as interleukin-6 release and a loss of blood-brain barrier integrity. Here, we tested the preventive and restorative effects of these bile acids regarding the disruption of blood-brain barrier properties by UCB in in vitro conditions mimicking severe neonatal hyperbilirubinemia and using the same experimental blood-brain barrier model. Both bile acids reduced the apoptotic cell death induced by UCB, but only glycoursodeoxycholic acid significantly counteracted caspase-3 activation. Bile acids also prevented the upregulation of interleukin-6 mRNA, whereas only ursodeoxycholic acid abrogated cytokine release. Regarding barrier integrity, only ursodeoxycholic acid abrogated UCB-induced barrier permeability. Better protective effects were obtained by bile acid pre-treatment, but a strong efficacy was still observed by their addition after UCB treatment. Finally, both bile acids showed ability to cross confluent monolayers of human brain microvascular endothelial cells in a time-dependent manner. Collectively, data disclose a therapeutic time-window for preventive and restorative effects of ursodeoxycholic acid and glycoursodeoxycholic acid against UCB-induced blood-brain barrier disruption and damage to human brain microvascular endothelial cells. PMID:25821432

  14. N-Methyl-d-Aspartate Receptor and Neuronal Nitric Oxide Synthase Activation Mediate Bilirubin-Induced Neurotoxicity

    PubMed Central

    Brito, Maria A; Vaz, Ana R; Silva, Sandra L; Falcão, Ana S; Fernandes, Adelaide; Silva, Rui FM; Brites, Dora

    2010-01-01

    Hyperbilirubinemia may lead to neurotoxicity and neuronal death. Although the mechanisms of nerve cell damage by unconjugated bilirubin (UCB) appear to involve a disruption of the redox status and excitotoxicity, the contribution of nitric oxide (NO·) and of N-methyl-d-aspartate (NMDA) glutamate receptors is unclear. We investigated the role of NO· and NMDA glutamate receptors in the pathways of nerve cell demise by UCB. Neurons were incubated with 100 μmol/L UCB, in the presence of 100 μmol/L human serum albumin for 4 h at 37ºC, alone or in combination with N-ω-nitro-l-arginine methyl ester (l-NAME) (an inhibitor of neuronal nitric oxide synthase [nNOS]), hemoglobin (an NO· scavenger) or (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine maleate (MK-801) (an NMDA-receptor antagonist). Exposure to UCB led to increased expression of nNOS and production of both NO· and cyclic guanosine 3′,5′-monophosphate (cGMP), along with protein oxidation and depletion of glutathione. These events concurred for cell dysfunction and death and were counteracted by l-NAME. Moreover, the UCB-induced loss of neuronal viability was abolished by hemoglobin, whereas the activation of nNOS and production of both NO· and cGMP were counteracted by MK-801, resulting in significant protection from cell dysfunction and death. These results reinforce the involvement of oxidative stress by showing that nerve cell damage by UCB is mediated by NO· and therefore is counteracted by NO· inhibitors or scavengers. Our findings strongly suggest that the activation of nNOS and neurotoxicity occur through the engagement of NMDA receptors. These data reveal a role for overstimulation of glutamate receptors in mediating oxidative damage by UCB. PMID:20593111

  15. Anti-cancer effects of blue-green alga Spirulina platensis, a natural source of bilirubin-like tetrapyrrolic compounds.

    PubMed

    Koníčková, Renata; Vaňková, Kateřina; Vaníková, Jana; Váňová, Kateřina; Muchová, Lucie; Subhanová, Iva; Zadinová, Marie; Zelenka, Jaroslav; Dvořák, Aleš; Kolář, Michal; Strnad, Hynek; Rimpelová, Silvie; Ruml, Tomáš; J Wong, Ronald; Vítek, Libor

    2014-01-01

    Spirulina platensis is a blue-green alga used as a dietary supplement because of its hypocholesterolemic properties. Among other bioactive substances, it is also rich in tetrapyrrolic compounds closely related to bilirubin molecule, a potent antioxidant and anti-proliferative agent. The aim of our study was to evaluate possible anticancer effects of S. platensis and S. platensis-derived tetrapyrroles using an experimental model of pancreatic cancer. The anti-proliferative effects of S. platensis and its tetrapyrrolic components [phycocyanobilin (PCB) and chlorophyllin, a surrogate molecule for chlorophyll A] were tested on several human pancreatic cancer cell lines and xenotransplanted nude mice. The effects of experimental therapeutics on mitochondrial reactive oxygen species (ROS) production and glutathione redox status were also evaluated. Compared to untreated cells, experimental therapeutics significantly decreased proliferation of human pancreatic cancer cell lines in vitro in a dose-dependent manner (from 0.16 g•L-1 [S. platensis], 60 μM [PCB], and 125 μM [chlorophyllin], p<0.05). The anti-proliferative effects of S. platensis were also shown in vivo, where inhibition of pancreatic cancer growth was evidenced since the third day of treatment (p < 0.05). All tested compounds decreased generation of mitochondrial ROS and glutathione redox status (p = 0.0006; 0.016; and 0.006 for S. platensis, PCB, and chlorophyllin, respectively). In conclusion, S. platensis and its tetrapyrrolic components substantially decreased the proliferation of experimental pancreatic cancer. These data support a chemopreventive role of this edible alga. Furthermore, it seems that dietary supplementation with this alga might enhance systemic pool of tetrapyrroles, known to be higher in subjects with Gilbert syndrome. PMID:24552870

  16. Hyperglycaemia promotes human brain microvascular endothelial cell apoptosis via induction of protein kinase C-ßI and prooxidant enzyme NADPH oxidase

    PubMed Central

    Shao, Beili; Bayraktutan, Ulvi

    2014-01-01

    Blood–brain barrier disruption represents a key feature in hyperglycaemia-aggravated cerebral damage after an ischaemic stroke. Although the underlying mechanisms remain largely unknown, activation of protein kinase C (PKC) is thought to play a critical role. This study examined whether apoptosis of human brain microvascular endothelial cells (HBMEC) might contribute to hyperglycaemia-evoked barrier damage and assessed the specific role of PKC in this phenomenon. Treatments with hyperglycaemia (25 mM) or phorbol myristate acetate (PMA, a protein kinase C activator, 100 nM) significantly increased NADPH oxidase activity, O2•- generation, proapoptotic protein Bax expression, TUNEL-positive staining and caspase-3/7 activities. Pharmacological inhibition of NADPH oxidase, PKC-a, PKC-ß or PKC-ßI via their specific inhibitors and neutralisation of O2•- by a cell-permeable superoxide dismutase mimetic, MnTBAP normalised all the aforementioned increases induced by hyperglycaemia. Suppression of these PKC isoforms also negated the stimulatory effects of hyperglycaemia on the protein expression of NADPH oxidase membrane-bound components, Nox2 and p22-phox which determine the overall enzymatic activity. Silencing of PKC-ßI gene through use of specific siRNAs abolished the effects of both hyperglycaemia and PMA on endothelial cell NADPH oxidase activity, O2•- production and apoptosis and consequently improved the integrity and function of an in vitro model of human cerebral barrier comprising HBMEC, astrocytes and pericytes. Hyperglycaemia-mediated apoptosis of HBMEC contributes to cerebral barrier dysfunction and is modulated by sequential activations of PKC-ßI and NADPH oxidase. PMID:24936444

  17. Lower bounds for randomized Exclusive Write PRAMs

    SciTech Connect

    MacKenzie, P.D.

    1995-05-02

    In this paper we study the question: How useful is randomization in speeding up Exclusive Write PRAM computations? Our results give further evidence that randomization is of limited use in these types of computations. First we examine a compaction problem on both the CREW and EREW PRAM models, and we present randomized lower bounds which match the best deterministic lower bounds known. (For the CREW PRAM model, the lower bound is asymptotically optimal.) These are the first non-trivial randomized lower bounds known for the compaction problem on these models. We show that our lower bounds also apply to the problem of approximate compaction. Next we examine the problem of computing boolean functions on the CREW PRAM model, and we present a randomized lower bound, which improves on the previous best randomized lower bound for many boolean functions, including the OR function. (The previous lower bounds for these functions were asymptotically optimal, but we improve the constant multiplicative factor.) We also give an alternate proof for the randomized lower bound on PARITY, which was already optimal to within a constant additive factor. Lastly, we give a randomized lower bound for integer merging on an EREW PRAM which matches the best deterministic lower bound known. In all our proofs, we use the Random Adversary method, which has previously only been used for proving lower bounds on models with Concurrent Write capabilities. Thus this paper also serves to illustrate the power and generality of this method for proving parallel randomized lower bounds.

  18. Expression of terminal oxidases under nutrient-starved conditions in Shewanella oneidensis: detection of the A-type cytochrome c oxidase.

    PubMed

    Le Laz, Sébastien; Kpebe, Arlette; Bauzan, Marielle; Lignon, Sabrina; Rousset, Marc; Brugna, Myriam

    2016-01-01

    Shewanella species are facultative anaerobic bacteria that colonize redox-stratified habitats where O2 and nutrient concentrations fluctuate. The model species Shewanella oneidensis MR-1 possesses genes coding for three terminal oxidases that can perform O2 respiration: a bd-type quinol oxidase and cytochrome c oxidases of the cbb3-type and the A-type. Whereas the bd- and cbb3-type oxidases are routinely detected, evidence for the expression of the A-type enzyme has so far been lacking. Here, we investigated the effect of nutrient starvation on the expression of these terminal oxidases under different O2 tensions. Our results reveal that the bd-type oxidase plays a significant role under nutrient starvation in aerobic conditions. The expression of the cbb3-type oxidase is also modulated by the nutrient composition of the medium and increases especially under iron-deficiency in exponentially growing cells. Most importantly, under conditions of carbon depletion, high O2 and stationary-growth, we report for the first time the expression of the A-type oxidase in S. oneidensis, indicating that this terminal oxidase is not functionally lost. The physiological role of the A-type oxidase in energy conservation and in the adaptation of S. oneidensis to redox-stratified environments is discussed. PMID:26815910

  19. Expression of terminal oxidases under nutrient-starved conditions in Shewanella oneidensis: detection of the A-type cytochrome c oxidase

    PubMed Central

    Le Laz, Sébastien; kpebe, Arlette; Bauzan, Marielle; Lignon, Sabrina; Rousset, Marc; Brugna, Myriam

    2016-01-01

    Shewanella species are facultative anaerobic bacteria that colonize redox-stratified habitats where O2 and nutrient concentrations fluctuate. The model species Shewanella oneidensis MR-1 possesses genes coding for three terminal oxidases that can perform O2 respiration: a bd-type quinol oxidase and cytochrome c oxidases of the cbb3-type and the A-type. Whereas the bd- and cbb3-type oxidases are routinely detected, evidence for the expression of the A-type enzyme has so far been lacking. Here, we investigated the effect of nutrient starvation on the expression of these terminal oxidases under different O2 tensions. Our results reveal that the bd-type oxidase plays a significant role under nutrient starvation in aerobic conditions. The expression of the cbb3-type oxidase is also modulated by the nutrient composition of the medium and increases especially under iron-deficiency in exponentially growing cells. Most importantly, under conditions of carbon depletion, high O2 and stationary-growth, we report for the first time the expression of the A-type oxidase in S. oneidensis, indicating that this terminal oxidase is not functionally lost. The physiological role of the A-type oxidase in energy conservation and in the adaptation of S. oneidensis to redox-stratified environments is discussed. PMID:26815910

  20. The stabilizing effects of immobilization in D-amino acid oxidase from Trigonopsis variabilis

    PubMed Central

    Dib, Iskandar; Nidetzky, Bernd

    2008-01-01

    Background Immobilization of Trigonopsis variabilis D-amino acid oxidase (TvDAO) on solid support is the key to a reasonably stable performance of this enzyme in the industrial process for the conversion of cephalosporin C as well as in other biocatalytic applications. Results To provide a mechanistic basis for the stabilization of the carrier-bound oxidase we analyzed the stabilizing effects of immobilization in TvDAO exposed to the stress of elevated temperature and operational conditions. Two different strategies of immobilization were used: multi-point covalent binding to epoxy-activated Sepabeads EC-EP; and non-covalent oriented immobilization of the enzyme through affinity of its N-terminal Strep-tag to Strep-Tactin coated on insoluble particles. At 50°C, the oriented immobilizate was not stabilized as compared to the free enzyme. The structure of TvDAO was stabilized via covalent attachment to Sepabeads EC-EP but concomitantly, binding of the FAD cofactor was weakened. FAD release from the enzyme into solution markedly reduced the positive effect of immobilization on the overall stability of TvDAO. Under conditions of substrate conversion in a bubble-aerated stirred tank reactor, both immobilization techniques as well as the addition of the surfactant Pluronic F-68 stabilized TvDAO by protecting the enzyme from the deleterious effect of gas-liquid interfaces. Immobilization of TvDAO on Sepabeads EC-EP however stabilized the enzyme beyond this effect and led to a biocatalyst that could be re-used in multiple cycles of substrate conversion. Conclusion Multi-point covalent attachment of TvDAO on an isoluble porous carrier provides stabilization against the denaturing effects of high temperature and exposure to a gas-liquid interface. Improvement of binding of the FAD cofactor, probably by using methods of protein engineering, would further enhance the stability of the immobilized enzyme. PMID:18798979

  1. CO impedes superfast O2 binding in ba3 cytochrome oxidase from Thermus thermophilus

    PubMed Central

    Szundi, Istvan; Funatogawa, Chie; Fee, James A.; Soulimane, Tewfik; Einarsdóttir, Ólöf

    2010-01-01

    Kinetic studies of heme-copper terminal oxidases using the CO flow-flash method are potentially compromised by the fate of the photodissociated CO. In this time-resolved optical absorption study, we compared the kinetics of dioxygen reduction by ba3 cytochrome c oxidase from Thermus thermophilus in the absence and presence of CO using a photolabile O2-carrier. A novel double-laser excitation is introduced in which dioxygen is generated by photolyzing the O2-carrier with a 355 nm laser pulse and the fully reduced CO-bound ba3 simultaneously with a second 532-nm laser pulse. A kinetic analysis reveals a sequential mechanism in which O2 binding to heme a3 at 90 μM O2 occurs with lifetimes of 9.3 and 110 μs in the absence and presence of CO, respectively, followed by a faster cleavage of the dioxygen bond (4.8 μs), which generates the P intermediate with the concomitant oxidation of heme b. The second-order rate constant of 1 × 109 M-1 s-1 for O2 binding to ba3 in the absence of CO is 10 times greater than observed in the presence of CO as well as for the bovine heart enzyme. The O2 bond cleavage in ba3 of 4.8 μs is also approximately 10 times faster than in the bovine enzyme. These results suggest important structural differences between the accessibility of O2 to the active site in ba3 and the bovine enzyme, and they demonstrate that the photodissociated CO impedes access of dioxygen to the heme a3 site in ba3, making the CO flow-flash method inapplicable. PMID:21097703

  2. Structure and proposed mechanism of α-glycerophosphate oxidase from Mycoplasma pneumoniae

    SciTech Connect

    Elkhal, Callia K.; Kean, Kelsey M.; Parsonage, Derek; Maenpuen, Somchart; Chaiyen, Pimchai; Claiborne, Al; Karplus, P. Andrew

    2015-03-14

    In this study, the formation of hydrogen peroxide (H₂O₂) by the FAD-dependent α-glycerophosphate oxidase (GlpO), is important for the pathogenesis of Streptococcus pneumoniae and Mycoplasma pneumoniae. The structurally known GlpO from Streptococcus sp. (SspGlpO) is similar to the pneumococcal protein (SpGlpO) and provides a guide for drug design against that target. However, M. pneumoniae GlpO (MpGlpO), having <20% sequence identity with structurally known GlpOs, appears to represent a second type of GlpO we designate as Type II GlpOs. Here, the recombinant His-tagged MpGlpO structure is described at ~2.5 Å resolution, solved by molecular replacement using as a search model the Bordetella pertussis protein 3253 (Bp3253) a protein of unknown function solved by structural genomics efforts. Recombinant MpGlpO is an active oxidase with a turnover number of ~580 min⁻¹ while Bp3253 showed no GlpO activity. No substantial differences exist between the oxidized and dithionite-reduced MpGlpO structures. Although, no liganded structures were determined, a comparison with the tartrate-bound Bp3253 structure and consideration of residue conservation patterns guided the construction of a model for α-glycerophosphate (Glp) recognition and turnover by MpGlpO. The predicted binding mode also appears relevant for the type I GlpOs (such as SspGlpO) despite differences in substrate recognition residues, and it implicates a histidine conserved in type I and II Glp oxidases and dehydrogenases as the catalytic acid/base. This work provides a solid foundation for guiding further studies of the mitochondrial Glp dehydrogenases as well as for continued studies of M. pneumoniae and S. pneumoniae glycerol metabolism and the development of novel therapeutics targeting MpGlpO and SpGlpO.

  3. Entropy bounds and dark energy

    NASA Astrophysics Data System (ADS)

    Hsu, Stephen D. H.

    2004-07-01

    Entropy bounds render quantum corrections to the cosmological constant Λ finite. Under certain assumptions, the natural value of Λ is of order the observed dark energy density ~10-10 eV4, thereby resolving the cosmological constant problem. We note that the dark energy equation of state in these scenarios is w≡p/ρ=0 over cosmological distances, and is strongly disfavored by observational data. Alternatively, Λ in these scenarios might account for the diffuse dark matter component of the cosmological energy density. Permanent address: Institute of Theoretical Science and Department of Physics, University of Oregon, Eugene, OR 97403.

  4. Bound potassium in muscle II.

    PubMed

    Hummel, Z

    1980-01-01

    Experiments were performed to decide between the alternatives a) the ionized K+ is in a dissolved state in the muscle water, or b) a part of the muscle potassium is in a "bound' state. Sartorius muscles of Rana esculenta were put into glicerol for about one hour at 0-2 degrees C. Most of muscle water came out, but most of muscle potassium remained in the muscles. In contrast to this: from muscle in heat rigor more potassium was released due to glicerol treating than from the intact ones. 1. Supposition a) is experimentally refuted. 2. Supposition b) corresponds to the experimental results. PMID:6969511

  5. Computer-controlled system for the study of oxidase reactions: application to the peroxidase-oxidase oscillator.

    PubMed

    McDonald, Andrew G; Tipton, Keith F

    2010-12-16

    An apparatus for the study of bisubstrate oxidase reactions at maintained steady-state substrate concentrations is described, and its specific application to the peroxidase-oxidase biochemical oscillator is reported. Instrument control and data acquisition are provided by custom software written in LabVIEW. The software allows measurement, recording, and control of dissolved oxygen through a Clark-type oxygen electrode, reaction monitoring by a UV/vis spectrophotometer, and controlled substrate delivery by a syringe infusion pump. For peroxidase from horseradish, the optimal pH for oscillatory behavior was found to be in the range 4.5-5.5. PMID:21049952

  6. Diiron centre mutations in Ciona intestinalis alternative oxidase abolish enzymatic activity and prevent rescue of cytochrome oxidase deficiency in flies

    PubMed Central

    Andjelković, Ana; Oliveira, Marcos T.; Cannino, Giuseppe; Yalgin, Cagri; Dhandapani, Praveen K.; Dufour, Eric; Rustin, Pierre; Szibor, Marten; Jacobs, Howard T.

    2015-01-01

    The mitochondrial alternative oxidase, AOX, carries out the non proton-motive re-oxidation of ubiquinol by oxygen in lower eukaryotes, plants and some animals. Here we created a modified version of AOX from Ciona instestinalis, carrying mutations at conserved residues predicted to be required for chelation of the diiron prosthetic group. The modified protein was stably expressed in mammalian cells or flies, but lacked enzymatic activity and was unable to rescue the phenotypes of flies knocked down for a subunit of cytochrome oxidase. The mutated AOX transgene is thus a potentially useful tool in studies of the physiological effects of AOX expression. PMID:26672986

  7. Transfusion of human volunteers with older, stored red blood cells produces extravascular hemolysis and circulating non–transferrin-bound iron

    PubMed Central

    Brittenham, Gary M.; Billote, Genia B.; Francis, Richard O.; Ginzburg, Yelena Z.; Hendrickson, Jeanne E.; Jhang, Jeffrey; Schwartz, Joseph; Sharma, Shruti; Sheth, Sujit; Sireci, Anthony N.; Stephens, Hannah L.; Stotler, Brie A.; Wojczyk, Boguslaw S.; Zimring, James C.; Spitalnik, Steven L.

    2011-01-01

    Transfusions of RBCs stored for longer durations are associated with adverse effects in hospitalized patients. We prospectively studied 14 healthy human volunteers who donated standard leuko-reduced, double RBC units. One unit was autologously transfused “fresh” (3-7 days of storage), and the other “older” unit was transfused after 40 to 42 days of storage. Of the routine laboratory parameters measured at defined times surrounding transfusion, significant differences between fresh and older transfusions were only observed in iron parameters and markers of extravascular hemolysis. Compared with fresh RBCs, mean serum total bilirubin increased by 0.55 mg/dL at 4 hours after transfusion of older RBCs (P = .0003), without significant changes in haptoglobin or lactate dehydrogenase. In addition, only after the older transfusion, transferrin saturation increased progressively over 4 hours to a mean of 64%, and non–transferrin-bound iron appeared, reaching a mean of 3.2μM. The increased concentrations of non–transferrin-bound iron correlated with enhanced proliferation in vitro of a pathogenic strain of Escherichia coli (r = 0.94, P = .002). Therefore, circulating non–transferrin-bound iron derived from rapid clearance of transfused, older stored RBCs may enhance transfusion-related complications, such as infection. The trial was registered with www.clinicaltrials.gov as #NCT01319552. PMID:22021369

  8. Consistent Tolerance Bounds for Statistical Distributions

    NASA Technical Reports Server (NTRS)

    Mezzacappa, M. A.

    1983-01-01

    Assumption that sample comes from population with particular distribution is made with confidence C if data lie between certain bounds. These "confidence bounds" depend on C and assumption about distribution of sampling errors around regression line. Graphical test criteria using tolerance bounds are applied in industry where statistical analysis influences product development and use. Applied to evaluate equipment life.

  9. Lower bound of concurrence for qubit systems

    NASA Astrophysics Data System (ADS)

    Zhu, Xue-Na; Fei, Shao-Ming

    2013-11-01

    We study the concurrence of four-qubit quantum states and provide analytical lower bounds of concurrence in terms of the monogamy inequality of concurrence for qubit systems. It is shown that these lower bounds are able to improve the existing bounds and detect entanglement better. The approach is generalized to arbitrary qubit systems.

  10. Quantum correlations beyond Tsirelson's bound

    NASA Astrophysics Data System (ADS)

    Berry, Dominic; Ringbauer, Martin; Fedrizzi, Alessandro; White, Andrew

    2014-03-01

    Violations of Bell inequalities show that there are correlations that cannot explained by any classical theory. Further violation, beyond Tsirelson's bound, shows that there are correlations that are not explained by quantum mechanics. Such super-quantum correlations would enable violation of information causality, where communication of one bit provides more than one bit of information [Nature 461, 1101 (2009)]. An unavoidable feature of all realistic Bell inequality experiments is loss. If one postselects on successful measurements, unentangled states can violate Bell inequalities. On the other hand, loss can be used to enhance the violation of Bell inequalities for entangled states. This can improve the ability to distinguish between entangled and unentangled states, despite loss. Here we report an optical experiment providing maximal violation of the CHSH-Bell inequality with entangled states. Due to loss and postselection, Tsirelson's bound is also violated. This enables us to more easily distinguish between entangled and unentangled states. In addition, it provides violation of information causality for the postselected data.

  11. Endurance bounds of aerial systems

    NASA Astrophysics Data System (ADS)

    Harrington, Aaron M.; Kroninger, Christopher M.

    2014-06-01

    Within the past few years micro aerial vehicles (MAVs) have received much more attention and are starting to proliferate into military as well as civilian roles. However, one of the major drawbacks for this technology currently, has been their poor endurance, usually below 10 minutes. This is a direct result of the inefficiencies inherent in their design. Often times, designers do not consider the various components in the vehicle design and match their performance to the desired mission for the vehicle. These vehicles lack a prescribed set of design guidelines or empirically derived design equations which often limits their design to selection of commercial off-the-shelf components without proper consideration of their affect on vehicle performance. In the current study, the design space for different vehicle configurations has been examined including insect flapping, avian flapping, rotary wing, and fixed wing, and their performance bounds are established. The propulsion system typical of a rotary wing vehicle is analyzed to establish current baselines for efficiency of vehicles at this scale. The power draw from communications is analyzed to determine its impact on vehicle performance. Finally, a representative fixed wing MAV is examined and the effects of adaptive structures as a means for increasing vehicle endurance and range are examined. This paper seeks to establish the performance bounds for micro air vehicles and establish a path forward for future designs so that efficiency may be maximized.

  12. IRON REGULATES XANTHINE OXIDASE ACTIVITY IN THE LUNG

    EPA Science Inventory

    The iron chelator deferoxamine has been reported to inhibit both xanthine oxidase (XO) and xanthine dehydrogenase activity, but the relationship of this effect to the availability of iron in the cellular and tissue environment remains unexplored. XO and total xanthine oxidoreduct...

  13. Beyond brown: polyphenol oxidases as enzymes of plant specialized metabolism

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Most cloned and/or characterized plant polyphenol oxidases (PPOs) have catecholase activity (i.e., they oxidize o-diphenols to o-quinones) and are localized or predicted to be localized to plastids. As a class, they have broad substrate specificity and are associated with browning of produce and oth...

  14. Reducing peanut allergens by high pressure combined with polyphenol oxidase

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polyphenol oxidase (PPO) has been shown to reduce major peanut allergens (Ara h 1 and Ara h 2). Because high pressure (HP) can increase enzyme activity, we postulated that further reduction of peanut allergens can be achieved through HP combined with PPO. Peanut extracts were treated with each of th...

  15. HypC, the anthrone oxidase involved in aflatoxin biosynthesis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Based on gene disruption and enzyme activity, hypC, an open reading frame in the pksA (aflC)/nor-1 (aflD) intergenic region in the aflatoxin biosynthesis cluster, encodes a 17 kDa oxidase that catalyzes the conversion of norsolorinic acid anthrone to norsolorinic acid....

  16. Platinum Nanoparticles: Efficient and Stable Catechol Oxidase Mimetics.

    PubMed

    Liu, Yi; Wu, Haohao; Chong, Yu; Wamer, Wayne G; Xia, Qingsu; Cai, Lining; Nie, Zhihong; Fu, Peter P; Yin, Jun-Jie

    2015-09-01

    Although enzyme-like nanomaterials have been extensively investigated over the past decade, most research has focused on the peroxidase-like, catalase-like, or SOD-like activity of these nanomaterials. Identifying nanomaterials having oxidase-like activities has received less attention. In this study, we demonstrate that platinum nanoparticles (Pt NPs) exhibit catechol oxidase-like activity, oxidizing polyphenols into the corresponding o-quinones. Four unique approaches are employed to demonstrate the catechol oxidase-like activity exerted by Pt NPs. First, UV-vis spectroscopy is used to monitor the oxidation of polyphenols catalyzed by Pt NPs. Second, the oxidized products of polyphenols are identified by ultrahigh-performance liquid chromatography (UHPLC) separation followed by high-resolution mass spectrometry (HRMS) identification. Third, electron spin resonance (ESR) oximetry techniques are used to confirm the O2 consumption during the oxidation reaction. Fourth, the intermediate products of semiquinone radicals formed during the oxidation of polyphenols are determined by ESR using spin stabilization. These results indicate Pt NPs possess catechol oxidase-like activity. Because polyphenols and related bioactive substances have been explored as potent antioxidants that could be useful for the prevention of cancer and cardiovascular diseases, and Pt NPs have been widely used in the chemical industry and medical science, it is essential to understand the potential effects of Pt NPs for altering or influencing the antioxidant activity of polyphenols. PMID:26305170

  17. Structural Changes and Proton Transfer in Cytochrome c Oxidase

    PubMed Central

    Vilhjálmsdóttir, Jóhanna; Johansson, Ann-Louise; Brzezinski, Peter

    2015-01-01

    In cytochrome c oxidase electron transfer from cytochrome c to O2 is linked to transmembrane proton pumping, which contributes to maintaining a proton electrochemical gradient across the membrane. The mechanism by which cytochrome c oxidase couples the exergonic electron transfer to the endergonic proton translocation is not known, but it presumably involves local structural changes that control the alternating proton access to the two sides of the membrane. Such redox-induced structural changes have been observed in X-ray crystallographic studies at residues 423–425 (in the R. sphaeroides oxidase), located near heme a. The aim of the present study is to investigate the functional effects of these structural changes on reaction steps associated with proton pumping. Residue Ser425 was modified using site-directed mutagenesis and time-resolved spectroscopy was used to investigate coupled electron-proton transfer upon reaction of the oxidase with O2. The data indicate that the structural change at position 425 propagates to the D proton pathway, which suggests a link between redox changes at heme a and modulation of intramolecular proton-transfer rates. PMID:26310633

  18. Glucose oxidase-magnetite nanoparticle bioconjugate for glucose sensing.

    PubMed

    Rossi, Liane M; Quach, Ashley D; Rosenzweig, Zeev

    2004-10-01

    Immobilization of bioactive molecules on the surface of magnetic nanoparticles is of great interest, because the magnetic properties of these bioconjugates promise to greatly improve the delivery and recovery of biomolecules in biomedical applications. Here we present the preparation and functionalization of magnetite (Fe3O4) nanoparticles 20 nm in diameter and the successful covalent conjugation of the enzyme glucose oxidase to the amino-modified nanoparticle surface. Functionalization of the magnetic nanoparticle surface with amino groups greatly increased the amount and activity of the immobilized enzyme compared with immobilization procedures involving physical adsorption. The enzymatic activity of the glucose oxidase-coated magnetic nanoparticles was investigated by monitoring oxygen consumption during the enzymatic oxidation of glucose using a ruthenium phenanthroline fluorescent complex for oxygen sensing. The glucose oxidase-coated magnetite nanoparticles could function as nanometric glucose sensors in glucose solutions of concentrations up to 20 mmol L(-1). Immobilization of glucose oxidase on the nanoparticles also increased the stability of the enzyme. When stored at 4 degrees C the nanoparticle suspensions maintained their bioactivity for up to 3 months. PMID:15448967

  19. Molecular dynamics in cytochrome c oxidase Moessbauer spectra deconvolution

    SciTech Connect

    Bossis, Fabrizio; Palese, Luigi L.

    2011-01-07

    Research highlights: {yields} Cytochrome c oxidase molecular dynamics serve to predict Moessbauer lineshape widths. {yields} Half height widths are used in modeling of Lorentzian doublets. {yields} Such spectral deconvolutions are useful in detecting the enzyme intermediates. -- Abstract: In this work low temperature molecular dynamics simulations of cytochrome c oxidase are used to predict an experimentally observable, namely Moessbauer spectra width. Predicted lineshapes are used to model Lorentzian doublets, with which published cytochrome c oxidase Moessbauer spectra were simulated. Molecular dynamics imposed constraints to spectral lineshapes permit to obtain useful information, like the presence of multiple chemical species in the binuclear center of cytochrome c oxidase. Moreover, a benchmark of quality for molecular dynamic simulations can be obtained. Despite the overwhelming importance of dynamics in electron-proton transfer systems, limited work has been devoted to unravel how much realistic are molecular dynamics simulations results. In this work, molecular dynamics based predictions are found to be in good agreement with published experimental spectra, showing that we can confidently rely on actual simulations. Molecular dynamics based deconvolution of Moessbauer spectra will lead to a renewed interest for application of this approach in bioenergetics.

  20. Polyphenol oxidase activity in co-ensiled temperate grasses

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polyphenol oxidase (PPO) and its o-diphenol substrates have been shown to effectively decrease proteolytic activity during the ensiling of forages such as red clover. Orchardgrass and smooth bromegrass both contain high levels of PPO activity, but lack appropriate levels of o-diphenols to adequately...