Extracellular Oxidases Research Articles

Reactive Oxygen Species Triggered Oxidative Degradation of Polystyrene in the Gut of Superworms (Zophobas atratus Larvae).

Oxidative decomposition of polystyrene (PS) by insects has been previously demonstrated, yet little is known about the oxidation mechanism and its effect on the metabolism of plastics within the insect gut. Here, we demonstrate the generation of reactive oxygen species (ROS) in the gut of superworms (Zophobas atratus larvae) under different feeding trails, which in turn induced the oxidative decomposition of ingested PS. The ROS were commonly generated in the larva gut, and PS consumption resulted in a significant increase of ROS with a maximum ·OH of 51.2 μmol/kg, which was five times higher than in the bran feeding group. Importantly, scavenging of ROS significantly decreased the oxidative depolymerization of PS, indicating a vital role of ROS in effective PS degradation in the gut of superworms. Further investigation suggested that the oxidative depolymerization of PS was caused by the combinatorial effect of ROS and extracellular oxidases of gut microbes. These results demonstrate that ROS were extensively produced within the intestinal microenvironment of insect larvae, which greatly favored the digestion of ingested bio-refractory polymers. This work provides new insights into the underlying biochemical mechanisms of plastic degradation in the gut.

Iodide uptake by forest soils is principally related to the activity of extracellular oxidases.

129I is a nuclear fission decay product of concern because of its long half-life (16Ma) and propensity to bioaccumulate. Microorganisms impact iodine mobility in soil systems by promoting iodination (covalent binding) of soil organic matter through processes that are not fully understood. Here, we examined iodide uptake by soils collected at two depths (0-10 and 10-20cm) from 5 deciduous and coniferous forests in Japan and the United States. Autoclaved soils, and soils amended with an enzyme inhibitor (sodium azide) or an antibacterial agent (bronopol), bound significantly less 125I tracer (93%, 81%, 61% decrease, respectively) than the untreated control soils, confirming a microbial role in soil iodide uptake. Correlation analyses identified the strongest significant correlation between 125I uptake and three explanatory variables, actinobacteria soil biomass (p = 6.04E-04, 1.35E-02 for Kendall-Tau and regression analysis, respectively), soil nitrogen content (p = 4.86E-04, 4.24E-03), and soil oxidase enzyme activity at pH 7.0 using the substrate L-DOPA (p = 2.83E-03, 4.33E-04) and at pH 5.5 using the ABTS (p = 5.09E-03, 3.14E-03). Together, the results suggest that extracellular oxidases, primarily of bacterial origin, are the primary catalyst for soil iodination in aerobic, surface soils of deciduous and coniferous forests, and that soil N content may be indicative of the availability of binding sites for reactive iodine species.

1H-13C heteronuclear single quantum coherence NMR evidence for iodination of natural organic matter influencing organo-iodine mobility in the environment

The complex biogeochemical behavior of iodine (I) isotopes and their interaction with natural organic matter (NOM) pose a challenge for transport models. Here, we present results from iodination experiments with humic acid (HA) and fulvic acid (FA) using 1H-13C heteronuclear single quantum coherence (HSQC) nuclear magnetic resonance (NMR) spectroscopy. Even though not a quantitative approach, 1H-13C HSQC NMR corroborated that iodination of NOM occurs primarily through aromatic electrophilic substitution of proton by I, and also revealed how iodination chemically alters HA and FA in a manner that potentially affects the mobility of iodinated NOM in the environment. Three types of iodination experiments were conducted with HA and FA: a) non-enzymatic iodination by IO3− (pH 3) and I− (pH 4 and 7), b) addition of lactoperoxidase to promote I−-iodination in the presence of the co-substrate, H2O2 (pH 7), and c) addition of laccase for facilitating I−-iodination in the presence of O2, with or without a mediator (pH 4). When mediators or H2O2 were present, extracellular oxidases and peroxidases enhanced I− incorporation into NOM by between 54% and 3400%. Iodination of HA, which was less than that of FA, enhanced HA's stability (inferred from increases in aliphatic compounds, decreases in carbohydrate moieties, and thus increased molecular hydrophobicity) yet reduced HA's tendency to incorporate more iodine. As such, HA is expected to act more as a sink for iodine in the environment. In contrast, iodination of FA appeared to generate additional iodine binding sites, which resulted in greater iodine uptake capability and enhanced mobility (inferred from decreases in aliphatic compounds, increases in carbohydrates, and thus decreases in molecular hydrophobicity). These results indicate that certain NOM moieties may enhance while others may inhibit radioiodine mobility in the aqueous environment.

Soil extracellular oxidases mediated nitrogen fertilization effects on soil organic carbon sequestration in bioenergy croplands

AbstractNitrogen (N) fertilization significantly affects soil extracellular oxidases, agents responsible for decomposition of slow turnover and recalcitrant soil organic carbon (SOC; e.g., lignin), and consequently influences soil carbon sequestration capacity. However, it remains unclear how soil oxidases mediate SOC sequestration under N fertilization, and whether these effects co‐vary with plant type (e.g., bioenergy crop species). Using a spatially explicit design and intensive soil sampling strategy under three fertilization treatments in switchgrass (SG: Panicum virgatum L.) and gamagrass (GG: Tripsacum dactyloides L.) croplands, we quantified the activities of polyphenolic oxidase (PHO), peroxidase (PER), and their sum associated with recalcitrant C acquisition (OX). The fertilization treatments included no N fertilizer input (NN), low N input (LN: 84 kg N ha−1 year−1 in urea), and high N input (HN: 168 kg N ha−1 year−1 in urea). Besides correlations between soil oxidases and SOC (formerly published), both descriptive and geostatistical approaches were applied to evaluate the effects of N fertilization and crop type on soil oxidases activities and their spatial distributions. Results showed significantly negative correlations between soil oxidase activities and SOC across all treatments. The negative relationship of soil oxidases and SOC was also evident under N fertilization. First, LN significantly depressed oxidases in both mean activities and spatial heterogeneity, which corresponded to increased SOC in SG (though by 5.4%). LN slightly influenced oxidases activities and their spatial heterogeneity, consistent with insignificant changes of SOC in GG. Second, HN showed trends of decrease in soil oxidase activities, which aligned with the significantly enhanced SOC in both croplands. Overall, this study demonstrated that soil oxidase activities acted as sensitive and negative mediators of SOC sequestration in bioenergy croplands and optimizing fertilizer use particularly in switchgrass cropland can improve for both carbon sequestration and environmental benefit.

Decolorization of the synthetic dye Remazol Brilliant Blue Reactive (RBBR) by Ganoderma lucidum on bio-adsorbent of the solid bleached sulfate paperboard coated with polyethylene terephthalate

White rot fungi can secrete many types of extracellular oxidases that are involved in the degradation of lignin in natural lignocellulosic substrates. Solid bleached sulfate (SBS) paperboard coated with polyethylene terephthalate (PET) is a type of cellulose and hemicellulose based waste from the packaging industry for ready-to-eat foods. In this context, the objective of this study was to evaluate the discoloration of the Remazol Brilliant Blue Reactive (RBBR) dye by bioabsorbent, consisting of ligninolytic macrofungal mycelium supported on PET-coated SBS paper. This work involved three steps: first, to determine the residual concentration of Remazol Reactive Bright Blue (RBBR) dye in the effluent produced in a dyeing process, according to the dye concentrations most applied in the textile industry. In the second step, the solution with medium concentration of the RBBR dye was then treated with the bioabsorbent, composed of ligninolytic macrofungus mycelium supported on PET-coated SBS paper. In the third stage, the technical feasibility of reusing the bioabsorbent was evaluated. The effluent generated from dyeing baths and washes was collected and analyzed, providing a final effluent with an average residual concentration of 57.3 mg/L of dye. The results showed that higher concentrations of carbon and nitrogen gave rise to the bio-adsorbent with higher values of total proteins and enzymatic activity. The bio-adsorbent use and reuse tests showed 94% color removal for the RBBR dye over 30 days of reuse. The use of the bio-adsorbent for the decolorization of dyes and textile effluents is a potential efficient and low cost.

Creation of Bifunctional Indicating Complex Based on Nanodiamonds and Extracellular Oxidases of Luminous Fungus Neonothopanus nambi.

A bifunctional indicating complex was created by immobilization of extracellular oxidases (glucose oxidase and peroxidases) of luminous fungus Neonothopanus nambi onto modified nanodiamonds (MNDs) synthesized by detonation. It was found that the enzymes firmly adsorb onto MND particles and exhibit their catalytic activity. Model in vitro experiments showed that the created MND-enzymes complex is suitable for repeated use for analyte (glucose and phenol) testing and retains its activity after storage at 4°C in deionized water for 1 month. The data obtained offer the prospects for developing a new class of reusable multifunctional indicating and diagnostic test systems on the basis of MNDs and higher fungal enzymes for medical and ecological analytics.

Trametes versicolor 의한 triphenyl methane계 염료의 분해

구름버섯(Trametes versicolor)은 phenolic compound인 CV와 MG를 효과적으로 탈색할 수 있었으며 고체와 액체배양 상태 모두에서 CV보다 MG를 더 효과적으로 탈색시켰다. 구름버섯에 의한 두 색소의 탈색 과정에서 phenolic compounds를 분해하는 것으로 알려진 세 가지 효소 중 laccase의 활성이 가장 높았다. MnP 역시 적은 수치지만 활성을 나타냈으며 LiP의 활성은 나타나지 않았다. 따라서 구름버섯에 의한 합성염료의 분해과정에서 laccase가 주로 사용되고 MnP는 탈색과정에서 보조적인 작용을 하는 것으로 추정된다. 그러나 CV의 경우 MnP가 활발하게 염료분해에 관여하는 것으로 판단된다. 또한 MG가 대부분 탈색되었을 때의 laccase 활성(0.16 U/mg)이 CV가 대부분 탈색되었을 때의 활성(0.23 U/mg)보다 현저하게 낮은 것으로 보아 구름버섯이 CV를 탈색시키는데 더 높은 활성의 laccase가 필요로 하는 것이 밝혀졌다. 본 실험에서 한국산 구름버섯 종의 CV와 MG 탈색능력이 확인되었으며 앞으로 한국산 구름버섯을 이용한 triphenyl methane계에 속하는 합성염료의 분해에 관한 친환경적 처리기술 개발에 도움이 될 것으로 기대된다. White rot fungi produce lignin-degrading enzymes such as laccase, manganese peroxidase and lignin peroxidase. These extracellular oxidases efficiently degrade recalcitrant synthetic dyestuffs with diverse chemical structures. Here, we examined the activities of lignin-degrading enzymes in Trametes versicolor using triphenyl methane dyes, crystal violet (CV) and malachite green (MG). Both dyes were decolorized by T. versicolor in solid and liquid culture conditions. T. versicolor decolorized MG more quickly than CV in both conditions. Among three ligninolytic enzymes, laccase was most abundantly found in the decolorization processes of CV and MG. However, higher activity of laccase was needed to degrade CV than MG. The much less activity of MnP was also detected. But the increase of MnP activity was well corresponded to the decolorization efficiency of CV, suggesting the involvement of MnP in CV degrading process. However, its role in the degradation process of MG is supposed to be subsidiary to laccase.

Degradation of textile dyes by white rot basidiomycetes.

ABSTRACT: Dyes released by the textile industries pose a threat to environmental quality. Ligninolytic white-rot basidiomycetes can effectively degrade colored effluents and conventional dyes. White-rot fungi produce various isoforms of extracellular oxidases including laccase, Mn peroxidase and lignin peroxidase (LiP), which are involved in the degradation of lignin in their natural lignocellulosic substrates. The textile industry, by far the most avid user of synthetic dyes, is in need of eco-efficient solutions for its colored effluents. White rot basidiomycetous fungi comprise the only group of organisms known to completely degrade lignin. Ligninolytic enzymes have potential applications in a large number of fields, including the chemical, fuel, food, agricultural, paper, textile, cosmetic industrial sectors and more. This ligninolytic system of white-rot fungi is also directly involved in the degradation of various xenobiotic compounds apart from textile dyes. Their capacities to remove xenobiotic substances make them a useful tool for bioremediation purposes. This paper reviews involvement of ligninolytic enzymes of white rot basidiomycetes in the degradation of textiles dyes and xenobiotic compounds for their industrial and biotechnological applications.

Extracellular oxidases of Cerrena sp. complementarily functioning in artificial dye decolorization including laccase, manganese peroxidase, and novel versatile peroxidases

Abstract Extracellular ligninolytic oxidoreductases produced by Cerrena sp. strain Ra, a soil-isolated basidiomycete with high artificial dye-decolorizing activity, were purified and characterized. One thermostable laccase, one typical manganese peroxidase, and two versatile peroxidases (VPI and VPII) were found in the culture of this fungus. The different characteristics of each enzyme enable the strain express wide-range of oxidizing activity under various conditions. VPI decolorizing activity was observed toward various kinds of dye compounds, and the activity and specificity varied depending on the oxidizing mediators added to the reaction mixture. Optimized VPI/mediator-coupling decolorizing will be widely used for industrial wastewater treatment.

Decomposition of organic matter by the ericoid mycorrhizal endophytes of Formosan rhododendron (Rhododendron formosanum Hemsl.)

Ericoid mycorrhizas are associated with a number of host plants in the Ericaceae in high-elevation regions of Taiwan. The ability of these microorganisms to thrive in harsh environmental conditions in the regions implies their capability of decomposing plant organic matter (raw humus). The objective of this study was to investigate the decomposition characteristics of three ericoid mycorrhizal endophytes isolated from the roots of Formosan rhododendron (Rhododendron formosanum Hemsl.). Molecular analysis indicated that strains Rf9 and Rf32 belong to the genus Cryptosporiopsis while strain Rf28 is a member of the genus Phialocephala. Mycorrhizal synthesis experiment showed that the roots of synthesized seedlings produced hyphal coils, a characteristic of ericoid mycorrhiza. Decomposition ability analysis revealed that strains Rf28 and Rf32 had the highest rates of decomposition of organic matter (up to 10.4% after 70days) while the value for strain Rf9 was about 6.8%. Consistently, these strains secreted extracellular oxidases when cultured on tannic acid medium. Enzyme assay revealed that strains Rf28 and Rf32 secreted peroxidase, laccase, tyrosinase, and cellulase, but strain Rf9 secreted mainly peroxidase and tyrosinase. Apparently, the differences in secreted hydrolytic enzymes among the three endophytes are related to their ability to decompose organic matter. In the mycorrhizal synthesis experiment, all inoculated seedlings survived in the organic matter substrate for 70days and exhibited a stronger vigor than the control. This study demonstrated that these three isolated endophytes, Rf9, Rf28, and Rf32, are ericoid mycorrhizal fungi, capable of forming ericoid mycorrhiza with Formosan rhododendron. Meanwhile, all three endophytes can secrete hydrolytic enzymes to decompose organic matter for growth, presumably a prerequisite for the adaptation of Formosan rhododendron to the harsh environments of high elevation.

Transcriptome and Secretome Analyses ofPhanerochaete chrysosporiumReveal Complex Patterns of Gene Expression

The wood decay basidiomycete Phanerochaete chrysosporium was grown under standard ligninolytic or cellulolytic conditions and subjected to whole-genome expression microarray analysis and liquid chromatography-tandem mass spectrometry of extracellular proteins. A total of 545 genes were flagged on the basis of significant changes in transcript accumulation and/or peptide sequences of the secreted proteins. Under nitrogen or carbon limitation, lignin and manganese peroxidase expression increased relative to nutrient replete medium. Various extracellular oxidases were also secreted in these media, supporting a physiological connection based on peroxide generation. Numerous genes presumed to be involved in mobilizing and recycling nitrogen were expressed under nitrogen limitation, and among these were several secreted glutamic acid proteases not previously observed. In medium containing microcrystalline cellulose as the sole carbon source, numerous genes encoding carbohydrate-active enzymes were upregulated. Among these were six members of the glycoside hydrolase family 61, as well as several polysaccharide lyases and carbohydrate esterases. Presenting a daunting challenge for future research, more than 190 upregulated genes are predicted to encode proteins of unknown function. Of these hypothetical proteins, approximately one-third featured predicted secretion signals, and 54 encoded proteins detected in extracellular filtrates. Our results affirm the importance of certain oxidative enzymes and, underscoring the complexity of lignocellulose degradation, also support an important role for many new proteins of unknown function.

Light and Oxygen Are Not Required for Harpin-induced Cell Death

Nicotiana sylvestris leaves challenged by the bacterial elicitor harpin N(Ea) were used as a model system in which to determine the respective roles of light, oxygen, photosynthesis, and respiration in the programmed cell death response in plants. The appearance of cell death markers, such as membrane damage, nuclear fragmentation, and induction of the stress-responsive element Tnt1, was observed in all conditions. However, the cell death process was delayed in the dark compared with the light, despite a similar accumulation of superoxide and hydrogen peroxide in the chloroplasts. In contrast, harpin-induced cell death was accelerated under very low oxygen (<0.1% O(2)) compared with air. Oxygen deprivation impaired accumulation of chloroplastic reactive oxygen species (ROS) and the induction of cytosolic antioxidant genes in both the light and the dark. It also attenuates the collapse of photosynthetic capacity and the respiratory burst driven by mitochondrial alternative oxidase activity observed in air. Since alternative oxidase is known to limit overreduction of the respiratory chain, these results strongly suggest that mitochondrial ROS accumulate in leaves elicited under low oxygen. We conclude that the harpin-induced cell death does not require ROS accumulation in the apoplast or in the chloroplasts but that mitochondrial ROS could be important in the orchestration of the cell suicide program.

Extracellular Oxidases of the Lignin-Degrading Fungus Panus tigrinus

Two extracellular oxidases (laccases) were isolated from the extracellular fluid of the fungus Panus (Lentinus) tigrinus cultivated in low-nitrogen medium supplemented with birch sawdust. The enzymes were purified by successive chromatography on columns with TEAE-cellulose and DEAE-Toyopearl 650M. Both oxidases catalyze oxidation of pyrocatechol and ABTS. Moreover, oxidase 1 also catalyzes oxidation of guaiacol, o-phenylenediamine, and syringaldazine. The enzymes have identical pH (7.0) and temperature (60-65 degrees C) optimums. Absorption spectra of the oxidases differ from the spectra of typical "blue" laccases and are similar to the spectrum of yellow oxidase.

Purification and characterization of intracellular and extracellular, thermostable and alkali-tolerant alcohol oxidases produced by a thermophilic fungus, Thermoascus aurantiacus NBRC 31693

Intracellular and extracellular alcohol oxidases (AO int and AO ext) were purified from the liquid and solid cultures of a thermophilic fungus, Thermoascus aurantiacus NBRC 31693, as electrophoretically and isoelectrophoretically homogeneous proteins, respectively. Both enzymes contained a flavin adenine dinucleotide (FAD) cofactor and were stained with Schiff's reagent. The molecular weight of AO int was estimated to be about 320 kDa and its subunit was 75 kDa. The molecular weight of AO ext was about 560 kDa, and it was composed of two types of subunits (75 kDa and 59 kDa). The pIs of AO int and AO ext were 5.88 and 6.08, respectively. AO int and AO ext were stable up to 60 degrees C and 55 degrees C, respectively. The enzymes were stable over a wide range of pH from 6 to 11. AO int oxidized short straight-chain alcohols (K(m) for methanol, 13.5 mM and K(m) for ethanol, 15.8 mM). On the other hand, AO ext could oxidize secondary alcohols and aromatic alcohols (veratryl alcohol and benzyl alcohol) in addition to straight-chain alcohols (K(m) for methanol, 0.5 mM and K(m) for ethanol, 10.2 mM).

White-rot fungi and their enzymes for the treatment of industrial dye effluents

White-rot fungi produce various isoforms of extracellular oxidases including laccase, Mn peroxidase and lignin peroxidase (LiP), which are involved in the degradation of lignin in their natural lignocellulosic substrates. This ligninolytic system of white-rot fungi (WRF) is directly involved in the degradation of various xenobiotic compounds and dyes. This review summarizes the state of the art in the research and prospective use of WRF and their enzymes (lignin-modifying enzymes, LME) for the treatment of industrial effluents, particularly dye containing effluents. The textile industry, by far the most avid user of synthetic dyes, is in need of ecoefficient solutions for its colored effluents. The decolorization and detoxification potential of WRF can be harnessed thanks to emerging knowledge of the physiology of these organisms as well as of the biocatalysis and stability characteristics of their enzymes. This knowledge will need to be transformed into reliable and robust waste treatment processes.

Production of cholesterol oxidase by a newly isolated Rhodococcus sp.

Fifteen strains of microorganisms with ability to degrade cholesterol were isolated. Among them a Gram-positive, non-motile, non-sporing bacterium with meso-DAP in the cell wall and with a rod-coccus cycle showed the highest ability for cholesterol degradation. It was identified as Rhodococcus sp. strain 2C and was deposited by code 1633 in Persian type culture collection (PTCC). This strain was able to produce high levels of both extracellular and cell-bound cholesterol oxidases in media containing cholesterol as a sole carbon source. The effects of medium composition and physical parameters on cholesterol oxidase production were studied. The optimized medium was found to contain cholesterol 0.15% (w/v), yeast extract 0.3% (w/v), diammonium hydrogen phosphate 0.1% (w/v), Tween 80 (0.05%). The optimum pH and temperature for cholesterol oxidase production in optimized medium were found to be 8–30 °C respectively. Triton X-100 showed the greatest effect in releasing the cell-bound enzyme. The first and most probably the main metabolite of cholesterol degradation was purified and identified as 4-cholestene-3-one.

Fungal attack on coal II. Solubilization of low-rank coal by filamentous fungi

The solubilization of low-rank coal by fungi is not accompanied by its depolymerization, which was proven by several screening programs. A total number of 728 fungal strains were yet tested for solubilizing a German low-rank coal (lignite). The coal were used oxidatively pretreated (3% H 2O 2) and natively. During the first screening, the nitrogen content of the media were reduced to select fungi, which solubilize coal under nitrogen-limited conditions. Among 480 micromycetous fungi tested, at least ten strains were able to solubilize native low-rank coal pieces on a nitrogen-reduced agar-medium. They all belong to the conidiospores forming deuteromycetes (molds). Under ‘solubilizing’ conditions, only two of these ten strains showed weak extracellular activities (oxidases, peroxidases) toward 2,2′-azinobis(3-ethylbenzthiazoline-6-sulphonate) [ABTS]. In a second screening, 256 wood and litter decaying basidiomycetes, above all white-rot fungi, were tested for their ability to solubilize low-rank coal under ligninolytic conditions: not any strain was able to form black coal droplets or conspicuous diffusion areas. By using a nitrogen-rich medium, a few basidiomycetes showed also coal solubilizing activities, however, their ligninolytic enzymes were inhibited. The ‘typical’ solubilization of low-rank coal (formation of black liquids from solid coal particles) by molds depends mainly on the nitrogen content of the medium (resulting in higher pH values) as well as on the oxidation grade of coal (e.g. H 2O 2-pretreatment). Extracellular oxidases and peroxidases, both ligninolytic and nonligninolytic ones, seem to play a minor, (or not any), role for this process.

Extracellular oxidases and the transformation of solubilised low-rank coal by wood-rot fungi

The involvement of extracellular oxidases in biotransformation of low-rank coal was assessed by correlating the ability of nine white-rot and brown-rot fungi to alter macromolecular material in alkali-solubilised brown coal with the spectrum of oxidases they produce when grown on low-nitrogen medium. The coal fraction used was that soluble at 3.0?pH?6.0 (SWC6 coal). In 15-ml cultures, Gloeophyllum trabeum, Lentinus lepideus and Trametes versicolor produced little or no lignin peroxidase, manganese (Mn) peroxidase or laccase activity and caused no change to SWC6 coal. Ganoderma applanatum and Pycnoporus cinnabarinus also produced no detectable lignin or Mn peroxidases or laccase yet increased the absorbance at 400 nm of SWC6 coal. G. applanatum, which produced veratryl alcohol oxidase, also increased the modal apparent molecular mass. SWC6 coal exposed to Merulius tremellosus and Perenniporia tephropora, which secreted Mn peroxidases and laccase and Phanerochaete chrysosporium, which produced Mn and lignin peroxidases was polymerised but had unchanged or decreased absorbance. In the case of both P. chrysosporium and M. tremellosus, polymerisation of SWC6 coal was most extensive, leading to the formation of a complex insoluble in 100 mM NaOH. Rigidoporus ulmarius, which produced only laccase, both polymerised and reduced the A 400 of SWC6 coal. P. chrysosporium, M. tremellosus and P. tephropora grown in 10-ml cultures produced a spectrum of oxidases similar to that in 15-ml cultures but, in each case, caused more extensive loss of A 400, and P. chrysosporium depolymerised SWC6 coal. It is concluded that the extracellular oxidases of white-rot fungi can transform low-rank coal macromolecules and that increased oxygen availability in the shallower 10-ml cultures favours catabolism over polymerisation.

Involvement of a new enzyme, glyoxal oxidase, in extracellular H2O2 production by Phanerochaete chrysosporium.

The importance of extracellular H2O2 in lignin degradation has become increasingly apparent with the recent discovery of H2O2-requiring ligninases produced by white-rot fungi. Here we describe a new H2O2-producing activity of Phanerochaete chrysosporium that involves extracellular oxidases able to use simple aldehyde, alpha-hydroxycarbonyl, or alpha-dicarbonyl compounds as substrates. The activity is expressed during secondary metabolism, when the ligninases are also expressed. Analytical isoelectric focusing of the extracellular proteins, followed by activity staining, indicated that minor proteins with broad substrate specificities are responsible for the oxidase activity. Two of the oxidase substrates, glyoxal and methylglyoxal, were also identified, as their quinoxaline derivatives, in the culture fluid as secondary metabolites. The significance of these findings is discussed with respect to lignin degradation and other proposed systems for H2O2 production in P. chrysosporium.

Extracellular enzymes of some additional fungi associated with mushroom culture

Twenty-four fungus isolates from the compost utilized in commercially growing Agaricus brunnescens were tested for their ability to produce extracellular enzymes involved in the degradation of cellulose, lignin and xylan, the major components of the straw of the compost. All 24 isolates were able to degrade carboxymethyl cellulose. Most were classified as weak or moderate producers of exo-β-glucanase. Twenty of the 24 were also able to hydrolyze filter paper, a crystalline cellulose. Nineteen of the 24 were able to hydrolyze xylan, a hemicellulose. The production of extracellular polyphenol oxidases was detected utilizing two tests; the blueing of alcoholic gum guaiacol, which indicates tyrosinase production, and the browning of malt extract-gallic acid agar, which indicates laccase production. Twenty produced tyrosinase, but only eight produced laccase. Agaricus brunnescens was also included in all of the tests. It produced exo-β-glucanase, hemicellulase, tyrosinase and lactase.