Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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    Effect of respiration and manganese on oxidative stress resistance of Lactobacillus plantarum WCFS1
    Watanabe, M. ; Veen, S. van der; Nakajima, H. ; Abee, T. - \ 2012
    Microbiology 158 (2012)1. - ISSN 1350-0872 - p. 293 - 300.
    lactic-acid bacteria - lactococcus-lactis - electron-transport - hydrogen-peroxide - escherichia-coli - catalase - expression - survival - tolerance - toxicity
    Lactobacillus plantarum is a facultatively anaerobic bacterium that can perform respiration under aerobic conditions in the presence of haem, with vitamin K2 acting as a source of menaquinone. We investigated growth performance and oxidative stress resistance of Lb. plantarum WCFS1 cultures grown in de Man, Rogosa and Sharpe (MRS) medium without and with added manganese under fermentative, aerobic, aerobic with haem, and respiratory conditions. Previous studies showed that Lb. plantarum WCFS1 lacks a superoxide dismutase and requires high levels of manganese for optimum fermentative and aerobic growth. In this study, respiratory growth with added manganese resulted in significantly higher cell densities compared to the other growth conditions, while without manganese added, similar but lower cell densities were reached. Notably, cells derived from the respiratory cultures showed the highest hydrogen peroxide resistance in all conditions tested, although similar activity levels of haem-dependent catalase were detected in cells grown under aerobic conditions with haem. These results indicate that oxidative stress resistance of Lb. plantarum is affected by respiratory growth, growth phase, haem and manganese. As levels of haem and manganese can differ considerably in the raw materials used in fermentation processes, including those of milk, meat and vegetables, the insight gained here may provide tools to increase the performance and robustness of starter bacteria
    Purification and characterization of a chlorite dismutase from Pseudomonas chloritidismutans
    Mehboob, F. ; Wolterink, A.F.W.M. ; Vermeulen, A.J. ; Jiang, B. ; Hagedoorn, P.L. ; Stams, A.J.M. ; Kengen, S.W.M. - \ 2009
    FEMS Microbiology Letters 293 (2009)1. - ISSN 0378-1097 - p. 115 - 121.
    desulfovibrio-vulgaris hildenborough - (per)chlorate-reducing bacteria - strain gr-1 - reductase - catalase
    The chlorite dismutase (Cld) of Pseudomonas chloritidismutans was purified from the periplasmic fraction in one step by hydroxyapatite chromatography. The enzyme has a molecular mass of 110 kDa and consists of four 31-kDa subunits. Enzyme catalysis followed Michaelis-Menten kinetics, with Vmax and K(m) values of 443 U mg(-1) and 84 microM, respectively. A pyridine-NaOH-dithionite-reduced Cld revealed a Soret peak at 418 nm, indicative for protoheme IX. The spectral data indicate the presence of 1.5 mol protoheme IX mol(-1) tetrameric enzyme while metal analysis revealed 2.2 mol iron mol(-1) tetrameric enzyme. High concentrations of chlorite resulted in the disappearance of the Soret peak, which coincided with loss in activity. Electron paramagnetic resonance analyses showed an axial high-spin ferric iron signal. Cld was inhibited by cyanide, azide, but not by hydroxylamine or 3-amino-1,2,3-triazole. Remarkably, the activity was drastically enhanced by kosmotropic salts, and chaotropic salts decreased the activity, in accordance with the Hofmeister series. Chlorite conversion in the presence of 18O-labeled water did not result in the formation of oxygen with a mass of 34 (16O-18O) or a mass of 36 ((18)O-(18)O), indicating that water is not a substrate in the reaction and that both oxygen atoms originate from chlorite
    Internal browning in pear fruit (Pyrus communis L. cv Conference) may be a result of a limited availability of energy and antioxidants
    Veltman, R.H. ; Lenthéric, I. ; Plas, L.H.W. van der; Peppelenbos, H.W. - \ 2003
    Postharvest Biology and Technology 28 (2003). - ISSN 0925-5214 - p. 295 - 302.
    superoxide-dismutase - atp production - ascorbic-acid - elevated co2 - apple fruit - mitochondria - respiration - catalase
    Storage of pears (Pyrus communis) under hypoxia, especially in the presence of increased CO2 partial pressures, can lead to development of brown core. Disorder development, concentrations of ascorbic acid (AA) and adenosine triphosphate (ATP), and respiration were examined under various O-2 (0-21 kPa) and CO2 (0 and 5 kPa) atmospheres during 31 days of storage. ATP production was estimated using the respiration data. Hypoxia increased brown core incidence, decreased AA and ATP concentrations, and lowered ATP production. AA concentrations decreased before brown core became visible. Adding CO2 to the storage atmosphere increased the severity of brown core. CO2 addition also decreased AA levels by about 46% at O-2 partial pressures of 2.5 kPa and higher. CO2, however, had variable effects on ATP production. No brown core was found in fruit kept at 0 kPa O-2 with or without CO2, nor decreased AA levels. These results support the hypothesis that brown core initiation is a consequence of membrane damage caused by a combination of oxygen free radical action and a lack of maintenance energy. This combination may lead to decompartmentation of intracellular structures and the initiation of brown pigmentation, visible in pears with disorders. (C) 2002 Elsevier Science B.V. All rights reserved.
    Antioxidants and myocardial infarction : the EURAMIC study
    Kardinaal, A.F.M. - \ 1994
    Agricultural University. Promotor(en): F.J. Kok; R.J.J. Hermus; P. van 't Veer. - S.l. : Kardinaal - ISBN 9789054852841 - 157
    hartinfarct - geneesmiddelen - hematologische middelen - immunosuppressieve middelen - peroxidasen - katalase - myocardial infarction - drugs - haematologic agents - immunosuppressive agents - peroxidases - catalase

    This thesis reports the background, design and results of a multi-centre study on the relationship between diet-derived antioxidants and the risk of acute myocardial infarction (MI) in men. Levels of α-tocopherol and β-carotene in adipose tissue and of selenium in toenails were compared between almost 700 patients with first MI and a similar number of control subjects, recruited in 8 European countries and Israel. The concentration of β-carotene in adipose tissue, expressed in quintiles of the distribution in controls, was inversely associated with the risk of MI (p for trend 0.001), independently of other risk factors. This association was strongest in current cigarette smokers and in subjects with a high proportion of polyunsaturated fatty acids in the adipose tissue. The risk of MI was not related to α-tocopherol in adipose tissue. In persons with low vitamin E levels, low toenail selenium was associated with a 2.5-fold increased risk of MI, compared to high selenium levels. An additional study among 85 healthy, non-smoking volunteers, aged 50-70, showed only a modest correlation of adipose tissue concentrations with dietary intake of α-tocopherol and β-carotene assessed by a food frequency questionnaire.

    Randomized controlled trials with varying doses and combinations of antioxidant nutrients should clarify whether the observed associations are causal. Until then, supplement use is not recommended, but a generous consumption of fruits and vegetables may be encouraged.

    Physiological roles and metabolism of fungal aryl alcohols
    Jong, E. de - \ 1993
    Agricultural University. Promotor(en): J.A.M. de Bont, co-promotor(en): M.M.G.R. Bol; J.A. Field. - S.l. : De Jong - ISBN 9789054851943 - 224
    enzymen - bosbouw - pulpbereiding - biodegradatie - oxidoreductasen - peroxidasen - katalase - basidiomycotina - scheurvorming - decompositie - degradatie - lignine - chemische verbindingen - enzymes - forestry - pulping - biodegradation - oxidoreductases - peroxidases - catalase - basidiomycotina - cracking - decomposition - degradation - lignin - chemical compounds

    The major structural elements of wood and other vascular tissues are cellulose, hemicellulose and generally 20-30% lignin. Lignin gives the plant strength, it serves as a barrier against microbial attack and it acts as a water impermeable seal across cell walls of the xylem tissue. However, the presence of lignin has practical drawbacks for some of the applications of lignocellulosic materials. First, lignin has to be removed for the production of high quality pulps. Second, lignin reduces the digestibility of lignocellulosic materials. High quality pulps can be produced with chemical methods, however the abundant use of chemicals and energy, and the formation of an enormous waste stream has led scientists to investigate the possibilities of biodelignification. White-rot fungi give the most rapid and extensive degradation and have become subject of intensive research. Results obtained with the model organism Phanerochaete chrysosporium and other strains have revealed that lignin biodegradation is an extracellular, oxidative and non-specific process. This unique biodegradative potential has been considered for broader applications such as waste water treatment and the degradation of xenobiotic compounds. The research described in this thesis concentrates on the function of aryl alcohols in fungal physiology.

    Aryl alcohols In the physiology of white-rot fungi. White-rot fungi have a versatile machinery of enzymes, including peroxidases and oxidases, which work in harmony with secondary aryl alcohol metabolites to degrade the recalcitrant, aromatic biopolymer lignin. In chapter 2 literature concerning the important physiological roles of aryl (veratryl, anisyl and chlorinated anisyl) alcohols in the ligninolytic enzyme system has been reviewed. Their functions include stabilization of lignin peroxidase, charge-transfer reactions and as substrate for oxidases generating extracellular H 2 O 2 .

    The experimental research described in this thesis was initiated to evaluate the possibilities of white-rot fungi in the biopulping of hemp stem wood. Sixty-seven basidiomycetes were isolated and screened for high peroxidative activity (chapter 3). Several of the new isolates were promising manganese peroxidase-producing white-rot fungi. Enzyme assays indicated that for the production of H 2 O 2 either extracellular glyoxal or aryl alcohol oxidase were present. In contrast, lignin peroxidase was only detected in P. chrysosporium , despite attempts to induce this enzyme in other strains with oxygen and oxygen/veratryl alcohol additions. A highly significant correlation was found between two ligninolytic indicators: ethene formation from α-keto-γ- methylthiolbutyric acid and the decolorization of a polymeric dye, Poly R-478. Three of the new isolates had significantly higher Poly R decolorizing activities compared to P. chrysosporium .

    One of the best Poly R decolorizing strains, Bjerkandera sp. BOS55 was selected for further characterization. A novel enzyme activity (manganese independent peroxidase) was detected in the extracellular fluid of Bjerkandera sp. BOS55 (chapter 4). The purified enzyme could oxidize several compounds like phenol red, 2,6-dimethoxyphenol (DMP), Poly R-478, 2,2'-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS) and guaiacol with H 2 O 2 as an electron acceptor. In contrast, veratryl alcohol was not a substrate. This enzyme also had the capacity to oxidize DMP in the absence of H 2 O 2 . Bjerkandera sp. BOS55 also produced de novo several aromatic metabolites. Besides veratryl alcohol and veratraldehyde, compounds which are known to be involved in the ligninolytic system of several other white-rot fungi (chapter 2), other metabolites were formed. These included anisaldehyde, 3-chloro-anisaldehyde, 3,5-dichloro-anisaldehyde and small amounts of the corresponding anisyl, 3-chloro-anisyl and 3,5-dichloro-anisyl alcohol (chapters 5 and 6). This was the first report of de novo biosynthesis of simple chlorinated aromatic compounds by a white-rot fungus. These unexpected findings led us investigate the physiological role(s) of the denovo biosynthesized chlorinated anisyl alcohols (chapter 6). All metabolites were produced simultaneously with the extracellular ligninolytic enzymes. The monoand dichlorinated anisyl alcohols appeared to be excellent substrates for the extracellular aryl alcohol oxidases. The formed aldehydes were readily recycled via reduction by washed fungal mycelium, thus creating an extracellular H 2 O 2 production system regulated by intracellular enzymes. Lignin peroxidase does not oxidize the chlorinated anisyl alcohols both in the absence and in the presence of veratryl alcohol. It was therefore concluded that the chlorinated anisyl alcohols are well protected against the fungus's own aggressive ligninolytic enzymes. The relative amounts of veratryl alcohol and the chlorinated anisyl alcohols differ significantly depending on the growth conditions, indicating that the production of veratryl alcohol and the (chlorinated) anisyl metabolites are independently regulated.

    It was concluded that the chlorinated anisyl metabolites, biosynthesized by the white-rot fungus Bjerkandera sp. BOS55, are purposeful for ecologically significant processes such as lignin degradation. These results made us speculate if a significant biogenesis of chlorinated aromatics by fungi occurs in natural environments (chapter 7). Many common wood- and forest litter-degrading fungi were indeed detected that produced chlorinated anisyl metabolites (CAM). These compounds, which are structurally related to xenobiotic chloroaromatics, were present in the environment and occur at high concentrations of approximately 75 mg CAM kg -1wood or litter. The ubiquity among common fungi to produce large amounts of chlorinated aromatic compounds in the environment leads to the conclusion that these kind of compounds can no longer be considered to originate from anthropogenic sources only.

    Degradation of aryl alcohols by fungi. In chapter 2 the anabolic and catabolic routes of aryl alcohols by white-rot fungi has been reviewed. These fungi can not use veratryl alcohol as sole source of carbon and energy. However, several bacteria, yeasts and fungi were selectively isolated from paper mill waste water that grew on veratryl alcohol (chapter 8). Penicilliumsimplicissimum was selected for the characterization of the veratryl alcohol degradation route. P. simplicissimum oxidized veratryl alcohol via a NAD(P) +-dependent veratryl alcohol dehydrogenase to veratraldehyde which was further oxidized to veratric acid in a NAD(P) +-dependent reaction. Veratric acid-grown cells contained NAD(P)H-dependent O -demethylase activity for veratrate, vanillate and isovanillate. Ring-cleavage of protocatechuate was by a protocatechuate 3,4-dioxygenase. An interesting aspect of P. simplicissimum is the production of vanillyl alcohol oxidase with covalently bound FAD (chapter 9). The intracellular enzyme was purified 32-fold. SDS-PAGE of the purified enzyme revealed a single fluorescent band of 65 Kda. Gel filtration and sedimentation-velocity experiments indicated that the purified enzyme exists in solution as an octamer, containing 1 molecule flavin/subunit. The covalently bound prosthetic group of the enzyme was identified as 8α-(N 3-histidyl)FAD from pH dependent fluorescence quenching (p Ka = 4.85) and no decrease in fluorescence upon reduction with sodium borohydride. The enzyme showed a narrow and rather peculiar substrate specificity. In addition to vanillyl alcohol and 4-hydroxybenzyl alcohol, eugenol and chavicol are substrates for the enzyme (chapter 10). The formed products, coniferyl and coumaryl alcohol are the natural precursors of lignin in plants. This reaction has a potential application to produce coniferyl alcohol and subsequent synthetic lignin (DHP) from the inexpensive precursor eugenol.

    Occurrence and properties of Petunia peroxidase a
    Hendriks, T. - \ 1989
    Agricultural University. Promotor(en): J. Bruinsma; L.C. van Loon. - S.l. : Hendriks - 159
    sierplanten - solanaceae - peroxidasen - katalase - petunia - ornamental plants - solanaceae - peroxidases - catalase - petunia

    Peroxidases are probably the most extensively studied enzymes in higher plants. Various isoenzymes occur as soluble proteins in the apoplast and in the vacuole, or are bound to membranes and cell walls. Their occurrence is often organ-specific and developmentally controlled, and there is circumstantial evidence that they function in growth, differentiation and defence. In Chapter I biochemical, physiological and genetic aspects of higher-plant peroxidases are reviewed, particularly in relation to the peroxidase system in petunia ( Stimoryne Rafin., formerly Petunia Juss.). It is pointed out that only by studying genetically characterized systems a full understanding of the biochemical and physiological characteristics of the various peroxidase isoenzymes can be obtained.

    In petunia eight different peroxidase isoenzymes have been genetically characterized and the occurrence of two more is inferred (Chapters 1 and 2). Of these , the most prominent one in leaves and stems is peroxidase a (PRXa), which migrates upon gel electrophoresis as a fast-moving, anionic set of three (sometimes four) molecular forms (mozymes). PRXa also occurs in petals, but is absent from roots. Leaves also contain the intermediatemoving anionic isoenzyme PRXb and the fast-moving cationic peroxidase bands of PRXc. PRXa and PRXc were shown to be the major peroxidases present in extracellular extracts obtained by the vacuum infiltration method, whereas PRXb was the most active peroxidase in isolated mesophyll protoplasts. The three isoenzymes differed in their affinity for Concanavalin A (Con A)-Sepharose; whereas PRXb bound firmly, and was released only in the presence of mannose or glucose, PRXa and PRXc were only partly retained. Since all peroxidases are glycoproteins, these results suggest that the nature of the oligosaccharide chains could be important in determining the cellular location of the peroxidases (Chapter 3).

    The localization of PRXa in the apoplast was exploited for its purification by preferentially extracting the enzyme from leaves by the vacuum infiltration method (Chapter 4). The electrophoretic variant (allozyme) PRXa1, the product of the allele prxA1 , present in the petunia cv. Roter Vogel, was purified over 1300-fold by two sequential acetone precipitations, gel filtration and chromatofocusing. The purified enzyme had a RZ (Reinheits Zahl; A 404nm /A 280nm ) of 3.6, an apparent molecular weight of about 37 kD and an isoelectric point of 3.8. The three molecular forms had slightly different molecular weights and were separated by affinity chromatography on a Con A-Sepharose column. The absorption spectrum of PRXa showed maxima at 404 (Soret band), 496 and 636 nm. Further spectral analy sis revealed that freshly isolated PRXa contains a bound hydrogen donor which is lost upon storage. The pH optimum for the reaction with hydrogen peroxide and guaiacol was 5.0 and the specific activity of the enzyme 61 mkat/g protein. The rate constants for its reaction with hydrogen peroxide and guaiacol were 2.6 x 10 6and 1.9 x 10 6M -1s -1, respectively. The reaction with guaiacol was inhibited by various aromatic compounds, notably by trihydroxylated and ortho - and para -dihydroxylated phenolic derivatives. Cinnamic-acid derivatives were more inhibitory than phenol or benzoic-acid derivatives. Coniferyl alcohol was completely inhibitory and was polymerized to presumed lignin-like material. Together with its localization in the apoplast, this substrate preference suggests that PRXa functions in the polymerization of lignin in the cell wall (Chapter 4).

    A specific antiserum raised in a rabbit against the purified PRXa was used to investigate as to how far peroxidases in other Solanaceous species are antigenically related to the petunia enzyme. Polyacrylamide gel electrophoresis in the presence of SDS (SDS-PAGE) followed by immunoblotting revealed the presence of cross-reacting proteins in all species tested. To determine whether these proteins represented peroxidases, thorn-apple ( Datura stramonium L.), tobacco ( Nicotiana tabacum L.), sweet pepper ( Capsicum annuum L.), potato ( Solanum tuberosum L.) and tomato ( Lycopersicon esculentum Mill.) were selected for further analysis. Immunoblots after native PAGE revealed that the antiserum specifically recognized the fastmoving anionic peroxidases that are localized in the apoplast. Despite their antigenic relatedness, these peroxidases differed with respect to heat stability and apparent molecular weight, possibly as a result of differences in glycosylation. Apparently, the Solanaceae contain orthologous genes encoding peroxidase isoenzymes homologous to petunia PRXa. The antiserum did not react with peroxidases from horseradish, turnip, African marigold, maize and oats (Chapter 5).

    Since PRXa is encoded by a single gene, the occurrence of three molecular forms must result from post-transcriptional or post-translational modifications. In Chapter 6 the nature of this heterogeneity was further analysed. Upon incubation with α-mannosidase the larger PRXa1.1 and PRXa1.2 were converted into immunoreactive products resembling the smaller PRXa1.3, suggesting that the three forms of PRXa1 differ in the number of terminal mannose residues. Some α-mannosidese activity was found to be present in the apoplast of leaves and may be responsible for the conversion of PRXa1.1 into PRXa1-2 and PRXa1.3 during leaf development. Lectin-affinity blotting suggested that PRXa contains small, complex-type carbohydrate chains; gas-chromatographic analysis indicated the presence of xylose, arabinose, fucose, mannose, glucose and galactose (Chapter 6).

    In Chapter 7 the distribution of the soluble peroxidase activity in tissues from petals, leaves and stems was investigated. In petals from fully expanded flowers, the upper epidermis contained nearly 20 % of the peroxidase activity, but no activity was present in protoplasts isolated from this tissue. The peroxidase activity in the upper epidermis was extra-cellular and consisted of PRXa. In protoplasts from the remaining tissue (mesophyll and lower epidermis) PM was the only peroxidase present; it was responsible for about 45 % of the total activity in petals. In leaves, the lower epidermis contained 14 - 40 % of the peroxidase activity, depending on their stage of development. Up to half of this activity was localized extracellularly. In stems, the epidermis contained nearly 80 0 of the peroxidase activity and 30 % of this activity was localized extracellularly. All extracellular extracts contained merely PRXa, suggesting that PRXa is localized mainly in the epidermis. By pressing transversely cut leaves and stems on membrane filters and using these dip-blots to localize the enzyme insitu , it was shown that both total peroxidase activity and immunoreactive PRXa are localized mainly in the epidermis, in leaves particularly at the site of the midrib and veins. The epidermal localization of PRXa is suggestive of a role in modulating growth rate or in defence, either by the deposition of lignin, suberin or cutin, or by increased cross-linking of various cell-wall polymers, or both (Chapter 7).

    In order to analyse the developmentally-controlled regulation of PRXa synthesis, it was attempted to obtain a cDNA probe for PRXa mRNA. Using a Poly(A) +-RNA fraction isolated from leaves, a possible precursor of PRXa was detected upon immunoprecipitation of invitro translation products, and an extended product of about 110 nucleotides was detected after reverse transcription primed with a mixed oligonucleotide probe based on a conserved amino-acid sequence in other plant peroxidases. The incorporation of 35S-methionine and 32P-CTP, respectively, was very low, suggesting that peroxidase mRNA was present at a low concentration only. A cDNA library in the bacteriophage λgt11 was constructed and screened with the antiserum or the oligonucleotide probe, but no positive clones were detected. Similarly, no peroxidase-specific clones were found upon screening a corrola- specific cDNA library with the antiserum (Chapter 8).

    In Chapter 9 possible functions of PRXa are discussed. Based on its extracellular localization, its organ and tissue specificity, and its catalytic properties, a function in the rigidification of cell walls in response to internal and external signals seems likely. Petunia PRXa strongly resembles a supposedly lignin-forming peroxidase in tobacco, and seems to be different from a suberin- forming peroxidase induced in potato tubers and tomato fruits upon wounding. Accordingly, in contrast to the total peroxidase activity, PM activity does not increase in floating leaf discs, indicating that it is not induced upon wounding. The identification of genetically defined peroxidase isoenzymes in members of the Solanaceae as being involved in specific physiological processes will provide insight into the function and the regulation of the peroxidase system in this plant family.

    Peroxydase en katalase in planten
    Braber, J.M. - \ 1979
    Wageningen : Centrum voor Agrobiologisch Onderzoek (CABO-verslag no. 22) - 28
    katalase - fermentatie - peroxidasen - plantenfysiologie - catalase - fermentation - peroxidases - plant physiology
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