Effect of Culture Conditions on the Production of
            <scp>d</scp>
            -Galactose Oxidase by
            <i>Dactylium dendroides</i>

Markus, Z.; Miller, G.; Avigad, Gad

doi:10.1128/am.13.5.686-693.1965

Cited by 15 publications

(3 citation statements)

References 15 publications

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“…Culture growth. As reported earlier (22,36,37), cultures of D. dendroides are easily maintained on a minimal medium in which thiamine pyrophosphate is the only added vitamin (22,36). Growth remained normal at all copper con-centrations used in culturing the fungus (10 nM to 1 mM) except for the changes in fungal cell coloration.…”

Section: Resultssupporting

confidence: 52%

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Biosynthesis and Cellular Distribution of the Two Superoxide Dismutases of Dactylium dendroides

Shatzman

Kosman

1979

J Bacteriol

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The synthesis and subcellular localization of the two superoxide dismutases of Dactylium dendroides were studied in relation to changes in copper and manganese availability. Cultures grew normally at all medium copper concentrations used (10 nM to 1 mM). In the presence of high (10 μM) copper, manganese was poorly absorbed in comparison to the other metals in the medium. However, cells grown at 10 nM copper exhibited a 3.5-fold increase in manganese content, while the concentration of the other metals remained constant. Cultures grown at 10 nM copper or more had 80% Cu/Zn enzyme and 20% mangani enzyme; the former was entirely in the cytosol, and the latter was mitochondrial. Removal of copper from the medium resulted in decreased Cu/Zn superoxide dismutase synthesis with a concomitant increase in the mangani enzyme such that total cellular superoxide dismutase activity remained constant. The mangani enzyme in excess of the 20% was present in the non-mitochondrial fraction. The mitochondria, therefore, show no variability with respect to superoxide dismutase content, whereas the soluble fraction varies from 100 to 13% Cu/Zn superoxide dismutase. Copper-starved cells that were synthesizing predominantly mangani superoxide dismutase could be switched over to mostly Cu/Zn superoxide dismutase synthesis by supplementing the medium with copper during growth. Immunoprecipitation experiments suggest that the decrease in Cu/Zn activity at low copper concentration is a result of decreased synthesis of that protein rather than the production of an inactive apoprotein.

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Section: Resultssupporting

confidence: 52%

“…Growth of organism and media preparation. The fungal isolate (NRRL 2903) and growth conditions are those previously described (22,36). Cells were cultured in an artificial medium that had been treated with Chelex 100 (Bio-Rad) as described earlier (37).…”

Section: Methodsmentioning

confidence: 99%

Biosynthesis and Cellular Distribution of the Two Superoxide Dismutases of Dactylium dendroides

Shatzman

Kosman

1979

J Bacteriol

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“…Based on the few studies reporting the detection in some Ascomycota fungal secretomes of a range of coppercontaining enzymes from the AA class including the FgrGalOx and other AA5 (45-47), we identified copper, manganese and ethanol as possible inducers of AA5. Copper is known to be crucial to activate the FgrGalOx and was used previously in combination with manganese to recover active FgrGalOx from fungal cultures (47). Ethanol is often used as an inducer of laccases (12) (other copper-containing AAs), and we noticed the presence of CROs in some secretomes induced with ethanol (45).…”

Section: Selection Of Fungal Strains and Time-course Secretomic Studymentioning

confidence: 66%

Identification of Copper-Containing Oxidoreductases in the Secretomes of Three Colletotrichum Species with a Focus on Copper Radical Oxidases for the Biocatalytic Production of Fatty Aldehydes

Ribeaucourt

Saker

Navarro

et al. 2021

Appl Environ Microbiol

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Copper Radical Alcohol Oxidases (CRO-AlcOx), which have been recently discovered among fungal phytopathogens are attractive for the production of fragrant fatty aldehydes. With the initial objective to investigate the secretion of CRO-AlcOx by natural fungal strains, we undertook time-course analyses of the secretomes of three Colletotrichum species ( C. graminicola , C. tabacum and C. destructivum) using proteomics. The addition of a copper-manganese-ethanol mixture in absence of any plant-biomass mimicking compounds to Colletotrichum cultures unexpectedly induced the secretion of up to 400 proteins, 29-52% of which were carbohydrate-active enzymes (CAZymes), including a wide diversity of copper-containing oxidoreductases from the auxiliary activities (AA) class (AA1, AA3, AA5, AA7, AA9, AA11-AA13, AA16). Under these specific conditions, while a CRO-glyoxal oxidase from the AA5_1 subfamily was among the most abundantly secreted proteins, the targeted AA5_2 CRO-AlcOx were secreted at lower levels, suggesting heterologous expression as a more promising strategy for CRO-AlcOx production and utilization. C. tabacum and C. destructivum CRO-AlcOx were thus expressed in Pichia pastoris and their preference toward both aromatic and aliphatic primary alcohols was assessed. The CRO-AlcOx from C. destructivum was further investigated in applied settings, revealing a full conversion of C6 and C8 alcohols into their corresponding fragrant aldehydes. IMPORTANCE In the context of the industrial shift toward greener processes, the biocatalytic production of aldehydes is of utmost interest owing to their importance for their use as flavors and fragrances ingredients. CRO-AlcOx have the potential to become platform enzymes for the oxidation of alcohols to aldehydes. However, the secretion of CRO-AlcOx by natural fungal strains has never been explored, while the use of crude fungal secretomes is an appealing approach for industrial application to alleviate various costs pertaining to biocatalysts production. While investigating this primary objective, the secretomics studies revealed unexpected results showing that under the oxidative-stressful conditions we probed, Colletotrichum species can secrete a broad diversity of copper-containing enzymes (laccases, sugar oxidoreductases, LPMOs) usually assigned to “plant-cell wall degradation”, despite the absence of any plant-biomass mimicking compound, and only little amount of CRO-AlcOx were secreted, pointing out at recombinant expression as the most promising path for their biocatalytic application.

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Inhibition of Galactose Oxidase by Extracts Obtained from the Mycelia Which Produce the Enzyme

Avigad

Markus

1965

Israel Journal of Chemistry

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Galactose oxidase (o -galactose: O 2 oxidoreductase, EC 1.1.3.9) is found as an extracellular enzyme in Dactylium dendroides cultures [1,2]. The enzyme is a copper protein and its activity can be abolished by the removal of Cu++ and reconstituted by the addition of the metal to the apoenzyme [3].In the course of a study on the effect of physiological conditions on galactose oxidase production by D. dendroides [4,5], it has been observed that incubation of the enzyme with autolyzing mycelia or mycelial extracts resulted in a slow decrease in enzyme activity. Further observations indicated that this inhibition is not due to proteolytic activity but rather to the presence of a galactose oxidase inhibitor. The present communication describes some of the properties of this inhibitory system.Washed mycelium of glucose grown [5] D. dendroides was dispersed in 5 times its wt. of 1 mM EDTA with 0.1 times its wt. of A-541 Alumina (80-200 mesh; Fisher Scientific Co ,; New York). The suspension was treated for 45 minutes in the Raytheon 10 KC sonic oscillator. Samples from the supernatant obtained by 20 minutes centrifugation at 8,000 X g were tested for galactose oxidase inhibition (Figure 1). The initial lag observed in the course of this inhibition was markedly reduced by 10 minutes boiling of the crude extract, a treatment which did not destroy inhibitory activity. Boiled extracts were then acidified to pH 4.5-5.0 by addition of 0.1 N HCI and the precipitate was removed by 5 minutes centrifugation at 3,000 X g. After neutralization by 0.1 M NaOH the supernatant was kept at -20 0 and used as an inhibitor I stock solution' .The inhibitor was not retained effectively by DEAE, carboxymethyl or phosphorylated cellulose columns. However, about 600/0 of contaminating protein (material reacting with the phenol reagent [6] could be removed from the stock solution by transfer through a DEAE-cellulose column by use of 1 mM EDTA in 1 mM phosphate buffer at pH 7.0 for elution of the inhibitor. Gel filtration on Sephadex G-25; G-75 (Pharmacia Inc ,; Uppsala , Sweden) or Bio-Gel P2 and P4 (BioRad Laboratories, Richmond, Cal.if.) columns ( Figure 2) indicated that the inhibitor is of small molecular weight. It was readily dialyzable only after the heat and acid treatment. This observation could be indicative of a tight association between the small •

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Effect of Culture Conditions on the Production of d -Galactose Oxidase by Dactylium dendroides

Cited by 15 publications

References 15 publications

Biosynthesis and Cellular Distribution of the Two Superoxide Dismutases of Dactylium dendroides

Biosynthesis and Cellular Distribution of the Two Superoxide Dismutases of Dactylium dendroides

Identification of Copper-Containing Oxidoreductases in the Secretomes of Three Colletotrichum Species with a Focus on Copper Radical Oxidases for the Biocatalytic Production of Fatty Aldehydes

Inhibition of Galactose Oxidase by Extracts Obtained from the Mycelia Which Produce the Enzyme

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