Mn-oxidizing Peroxidases Research Articles

HYPSIZYGUS MARMOREUS – A NOVEL POTENT DEGRADER OF LIGNOCELLULOSE RESIDUES

Hypsizygus marmoreus is an edible and medicinal commercially cultivated mushroom, whose ligninolytic potential is still insufficiently known. Therefore, the goals of this study were to profile its Mn-oxidizing peroxidases and laccases activity and its potential to depolymerize common agro-forestry residues. Raspberry and blackberry sawdust induced the production of highly active laccases (26006.83 U L-1 and 17576.79 U L-1, respectively), while the activity peaks of Mn-dependent peroxidase (134.47 U L-1) and Mn-independent peroxidase (184.40 U L-1) were recorded on plum sawdust and wheat straw, respectively. The pretreatment of corn stalks resulted in the highest lignin removal (57.15%), while the highest lignin/cellulose removal ratio (selectivity) was noted on oak sawdust (17.69). The maximal (20%) and the minimal (8.30%) dry matter loss were obtained for corn stalks and oak sawdust, respectively. The presented results showed the exceptional potential of the ligninolytic system of H. marmoreus HAI 605 and the possibility of its application in numerous biotechnological processes.

Potential of selected fungal species to degrade wheat straw, the most abundant plant raw material in Europe

BackgroundStructural component of plant biomass, lignocellulose, is the most abundant renewable resource in nature. Lignin is the most recalcitrant natural aromatic polymer and its degradation presents great challenge. Nowadays, the special attention is given to biological delignification, the process where white-rot fungi take the crucial place owing to strong ligninolytic enzyme system. However, fungal species, even strains, differ in potential to produce high active ligninolytic enzymes and consequently to delignify plant biomass. Therefore, the goals of the study were characterization of Mn-oxidizing peroxidases and laccases of numerous mushrooms as well as determination of their potential to delignify wheat straw, the plant raw material that, according to annual yield, takes the first place in Europe and the second one in the world.ResultsDuring wheat straw fermentation, Lentinus edodes HAI 858 produced the most active Mn-dependent and Mn-independent peroxidases (1443.2 U L−1 and 1045.5 U L−1, respectively), while Pleurotus eryngii HAI 711 was the best laccase producer (7804.3 U L−1). Visualized bends on zymogram confirmed these activities and demonstrated that laccases were the dominant ligninolytic enzymes in the studied species. Ganoderma lucidum BEOFB 435 showed considerable ability to degrade lignin (58.5%) and especially hemicellulose (74.8%), while the cellulose remained almost intact (0.7%). Remarkable selectivity in lignocellulose degradation was also noted in Pleurotus pulmonarius HAI 573 where degraded amounts of lignin, hemicellulose and cellulose were in ratio of 50.4%:15.3%:3.8%.ConclusionsAccording to the presented results, it can be concluded that white-rot fungi, due to ligninolytic enzymes features and degradation potential, could be important participants in various biotechnological processes including biotransformation of lignocellulose residues/wastes in food, feed, paper and biofuels.

Lignocellulose degradation by Daedaleopsis confragosa and D. tricolor

The properties and capacities of the ligninolytic enzymes of Daedaleopsis spp. are still unknown. This is the first study on the effect of plant residues and period of cultivation on the properties of Mn-oxidizing peroxidases and laccases of D. confragosa and D. tricolor, as well as their ligninolytic potentials. Wheat straw was the optimal carbon source for synthesis of highly active Mn-dependent peroxidases (4126.9 U/L in D. confragosa and 2037.9 U/L in D. tricolor). However, laccases were the predominant enzymes, and the best inducer of their activity (up 16000.0 U/L) was cherry sawdust. Wheat straw was the most susceptible plant residue to the effect of the enzymes, and extent of lignin degradation was 43.3% after 14 days of fermentation with D. tricolor. However, D. confragosa was a more effective lignin degrader, as it converted even 21.3% wheat straw lignin on the 6th day of cultivation. The results of the study clearly showed that delignification extent depends on mushroom species and on the type of plant residue, which is extremely important for potential use in biotechnological processes.

Wheat Straw Degradation by Trametes gibbosa: The Effect of Calcium Ions

Wheat straw is the major crop residue in Europe which makes it the most promising material for bioconversion. However, only 3% of annual production is used directly while the rest is considered as waste. A key step in processing of lignocellulosics is delignification by fungi and the presence of calcium can be of a great importance as it can modulate ligninolytic enzymes activities. Trametes gibbosa BEOFB 310 was used for solid-state fermentation of wheat straw in the presence of calcium in different concentrations. Samples were extracted after 19 days of fermentation and activities of Mn-oxidizing peroxidases and laccase were determined spectrophotometrically. Quantitative procedures were used to determine contents of hemicelluloses, cellulose, and lignin. Calcium induced activity of Mn-dependent peroxidase to a concentration of 5.0 mM (7185.2 ± 791.4 U L−1), which was threefold higher than in the control. Lignocellulose loss in wheat straw was stimulated by calcium addition and the maximum delignification was detected at concentration of 5.0 mM (52.9 ± 0.9%). Delignification was positively correlated to activity of Mn-dependent peroxidase. This study showed that wheat straw supplementation with calcium can significantly improve solid-state fermentation by increasing Trametes gibbosa Mn-dependent peroxidase activity and augmenting lignin degradation.

Degradation of beech wood and wheat straw by Trametes gibbosa

The aim of this study was to optimize cultivation conditions for Mn-oxidizing peroxidases and laccase production and selective degradation of beech wood and wheat straw lignin by Trametes gibbosa. To promote lignin degradation, the effect of different carbon and nitrogen sources, type of cultivation, enzyme dynamics and inducers were studied. Solid-state cultivation was optimal for lignin degradation. Wheat straw was the optimal carbon source for peroxidase activities stimulating the synthesis of numerous isoforms. The dynamics of enzymatic activities showed that 19-day-old fermentation is optimal and that activities were directly associated with enzyme production. Alteration of nitrogen sources and inducers was applied to increase the rate of lignin degradation and decrease cellulose degradation. The presence of nitrogen in the form of (NH4)2SO4 and concentration of 10 mM significantly increased the extent and selectivity of delignification of wheat straw compared with cellulose (44.1 vs. 36.1%). The most effective and selective wheat straw degradation (52.0% of lignin vs. 31.3% of cellulose) was achieved by enriching the optimum medium, as defined above, with 1.0 mM p-anisidine. The optimum conditions for wheat straw processing to achieve delignification and production of highly active ligninolytic enzymes by T. gibbosa were (NH4)2SO4/wheat bran medium supplemented with p-anisidine. This study indicated the significant potential of optimizing external factors as a promising tool for induction and selectivity in the degradation of lignocelluloses by T. gibbosa as a pretreatment for several biotechnological processes.

Degradation of wheat straw and oak sawdust by Ganoderma applanatum

Ganoderma applanatum is a widely-distributed wood decaying species whose ligninolytic enzyme system has not been sufficiently studied. The aims of the study were to profile its Mn-oxidizing peroxidases and laccases and defining of wheat straw and oak sawdust delignification extents depending on cultivation type. Activities of these enzymes were higher in submerged than in solid-state cultivation. Oak sawdust induced the highest activities of Mn-dependent (5545.5 U L−1) and Mn-independent peroxidases (5810.0 U L−1), and wheat straw stimulated laccase activity (11007.0 U L−1). The isoelectric focusing profiles of enzymes and extent of lignocellulose degradation were affected by plant residue as well as type and period of cultivation. Submerged cultivation induced the synthesis of a higher number of enzyme isoforms and the maximum levels of lignin, hemicellulose and cellulose degradation (40.9%, 32.7% and 27.4%) were reached during this oak sawdust fermentation. However, selectivity in fiber mineralisations was the highest during solid-state fermentation of wheat straw, which is important for possible application in various biotechnological processes that require accessible cellulose, such as production of more digestible feed, paper pulp and bioethanol.

Induction of wheat straw delignification by Trametes species.

Wheat straw is the major crop residue in European countries which makes it the most promising material for bioconversion into biofuels. However, cellulose and hemicellulose are protected with lignin, so delignification is an inevitable phase in lignocellulose processing. The organisms predominantly responsible for its degradation are white-rot fungi and among them Trametes species represent promising degraders due to a well-developed ligninolytic enzyme system. Although numerous studies have confirmed that low molecular weight compounds can induce the production and activity of ligninolytic enzymes it is not clear how this reflects on the extent of delignification. The aim of the study was to assess the capacity of p-anisidine and veratryl alcohol to induce the production and activity of Mn-oxidizing peroxidases and laccases, and wheat straw delignification by six Trametes species. Significant inter- and intraspecific variations in activity and features of these enzymes were found, as well as differences in the potential of lignocellulose degradation in the presence or absence of inducers. Differences in the catalytic properties of synthesized enzyme isoforms strongly affected lignin degradation. Apart from enhanced lignin degradation, the addition of p-anisidine could significantly improve the selectivity of wheat straw ligninolysis, which was especially evident for T. hirsuta strains.

Activity of Mn-Oxidizing Peroxidases of Ganoderma lucidum Depending on Cultivation Conditions

Trunks and stumps of various deciduous species act as natural habitats for Ganoderma lucidum. The chemical composition of their cell wall affects the development of fungal ligninolytic enzyme system as well as its ability to degrade lignin from the plant cell wall. Additionally, numerous compounds structurally similar to lignin can be degraded by the G. lucidum enzyme system which could take important roles in various biotechnological processes. The laccases, which are the dominant enzymes synthesized by G. lucidum, have been studied more extensively than the Mn-oxidizing peroxidases. Therefore, this study aimed to create the dynamics profile of Mn-oxidizing peroxidases activities in four G. lucidum strains, classifying and determining their properties depending on the cultivation type and plant residue as a carbon source in the medium, as well as to establish whether intraspecific variety exists. The findings suggest that submerged cultivation appeared to be a more appropriate cultivation type for enzyme activities compared with solid-state cultivation, and oak sawdust was a better carbon source than wheat straw. Under the optimum conditions, on day 14, G. lucidum BEOFB 431 was characterized by the highest levels of both Mn-dependent and Mn-independent peroxidase activities (4795.5 and 5170.5 U/L, respectively). Strain, cultivation type, and carbon source were factors that affected the profiles of Mn-oxidizing peroxidases isoenzymes.

The effect of trace elements on wheat straw degradation by Trametes gibbosa

The aim of the study was comparative analysis of ligninolytic enzyme productions of Trametes gibbosa and its ability to degrade wheat straw, as well as effect of trace elements on those processes. Mn-oxidizing peroxidases were the main enzymes and the tested trace elements affected their activities mainly inhibitory. Stimulatory effect on Mn-independent peroxidase had Mn2+ in all concentrations, while Zn2+ concentrations of 5.0 mM and 10.0 mM led to the maximum Mn-dependent peroxidase activity. The trace elements affected laccase activity differently. Zn2+ and Fe2+ (0.5 mM and 10.0 mM, respectively) had the greatest effect on the activity, while Mn2+ and Cu2+ at the amount of 10.0 mM completely stopped the enzyme production. The tested enzymes acted simultaneously on wheat straw degradation. The presence of 1.0 mM of Mn2+ caused the highest level of dry matter loss (43.1%), which was the consequence of hemicellulose, cellulose and lignin degradation in the rate of 62.6%, 35.6% and 61.2%, respectively. Selective lignin degradation was noted especially in the presence of Fe2+ at the concentration of 0.5 mM.

Potential of Trametes species to degrade lignin

Well developed ligninolytic enzyme system in white-rot mushrooms is responsible for degradation of low degradable and lignin rich unused plant debris, which represents serious environmental ballast. The biological degradation is environmentally friendly and economically justified process. Therefore, assessment of the potential of Trametes pubescens and Trametes multicolor to degrade lignin of wheat straw and oak sawdust during solid-state cultivation was the object of the study. Significantly high laccase activity was noted on day 7 and day 10 of wheat straw fermentation by T. pubescens BEOFB 330, while at other days the obtained values were lower from 2- to 8-fold. Contrary to laccase, the optimum substrate for Mn-oxidizing peroxidases was oak sawdust. Maxima activities of Mn-dependent peroxidase and Mn-independent peroxidase were noted on day 14 of cultivation of T. multicolor HAI 426 and T. pubescens BEOFB 330, respectively. Tested organisms, except T. multicolor HAI 428, have degraded about 50% of lignin in wheat straw that was more degradable residue than oak sawdust, which decay level ranged between 1.8% and 5.9%. Mn-oxidizing peroxidases had the main role in oak sawdust lignin degradation. T. multicolor HAI 426, was the best lignin degrader in which laccase activity was not noted.

Influence of Trace Elements on Ligninolytic Enzyme Activity of Pleurotus ostreatus and P. pulmonarius

Trace elements, at tolerable concentrations, are either part of the active site or act as an activity modulator of ligninolytic enzymes of white-rot mushrooms. They are usually in plant raw materials in a non-toxic amount or non-available and extractable forms. This study evaluated the effects of Fe, Zn, and Se on the activity of laccase and Mn-oxidizing peroxidases of P. ostreatus and P. pulmonarius during solid-state fermentation of grapevine sawdust. The studied species showed different levels of tolerance to the trace elements. A stimulatory effect of the microelements on laccase activity was demonstrated in P. ostreatus, while Fe and Zn were strong inhibitors of the activity of P. pulmonarius, which was contrary to Se, independent of the cultivation period. With the exception of SeO2, activity of Mn-oxidizing peroxidases in P. ostreatus was suppressed in various levels by the elements. However, in P. pulmonarius, activity against phenol red oxidation in the presence of external Mn2+ was stimulated by the elements on day 7, while on day 10, activity inhibition by Fe and Zn and stimulation by Se was noted. The effect on activity against phenol red oxidation in the absence of external Mn2+ was the opposite.

Trametes suaveolens as ligninolytic enzyme producer

Species of the genus Trametes represent one of the most efficient lignin-degraders which can be attributed to a well developed ligninolytic enzyme system. Current trends are screening of ability of new species to produce these enzymes, as well as the optimization of conditions for their overproduction. Therefore, the aim of the study was to evaluate the potential of T. suaveolens to synthesize laccase and Mn-oxidizing peroxidases during fermentation of the selected plant raw materials. Level of enzyme activities was measured on 7, 10 and 14th day of submersion, as well as the solid-state fermentation of wheat straw and oak sawdust in the presence of NH4NO3 in previously determined optimal nitrogen concentration of 25 mM. The enzyme activity was determined spectrophotometrically using ABTS and phenol red as the substrates. The highest level of laccase activity (1087.1 U/L) was noted after 7 days of wheat straw solid-state fermentation, while during the submerged cultivation the production of the enzyme was not noted. Submerged cultivation in oak sawdust-enriched medium was the optimal for activity of Mn-dependent peroxidase (1767.7 U/L on day 14) and Mn-independent peroxidase (1113.7 U/L on day 7). Introduction of T. suaveolens to produce ligninolytic enzyme represented the base for further study, as well as the determination of relation between enzyme activity and rate of lignin degradation. It could lead to greater possibility of fungal species selection with high delignification capacity, which could take participation in sustainable production of food, feed, fibres, and energy, environmentally friendly pollution prevention, and bioremediation.

Ligninolytic enzyme production by Lenzites betulinus on selected plant raw materials

To get a better insight into the ligninolytic system of Lenzites betulinus, the effect of wheat straw and oak sawdust, as carbon sources, on production of Mn-oxidizing peroxidases and laccase, under solid-state and submerged fermentation, was studied. Obtained results revealed considerable differences related to the both factors affecting enzyme activities. Wheat straw was more favorable carbon source for Mn-oxidizing peroxidases and oak sawdust for laccase activity. Solid-state fermentation of wheat straw was optimal for Mn-dependent peroxidase activity (72.1 Ul-1). In contrary to this, submerged fermentation of the same residue gave the highest level of versatile peroxidase activity (25.4 Ul-1). The peak of laccase activity was noted during solid-state fermentation of oak sawdust (32.3 Ul-1), while this enzyme was not detected under submerged fermentation of any plant residues.

Morpho-physiological Diversity between Lingzhi or Reishi Medicinal Mushroom Ganoderma lucidium (W. Curt.:Fr.) P. Karst. and G. carnosum Pat.

Two weakly differentiated taxa, Ganoderma lucidum and G. carnosum, were compared in their sufficient morphological and physiological features. The obtained results showed that dimensions of basidiospores and pileocystidia were insignificantly different, while pore shape and dimensions have shown greater diversity with average diameter of 138.46 μm in G. carnosum and 238.34 μm in G. lucidum. Mycelial growth rate was higher in G. lucidum (8.39 mm day-1) than in G. carnosum (6.02 mm day-1). G. lucidum was also a slightly better producer of biomass and extracellular polysaccharides (28.16 g L-1 and 1.42 mg mL-1, respectively) than G. carnosum (23.68 g L-1 and 0.35 mg mL-1, respectively). However, a higher amount of synthesized intracellular polysaccharides was noted in G. carnosum than in G. lucidum (40.00 mg g-1 and 30.00 mg g-1 of dry biomass, respectively). Higher activity levels of Mn-oxidizing peroxidases were obtained in G. carnosum, while G. lucidum was a better laccase producer. In G. carnosum, corn stem/NH₄NO₃ medium with nitrogen concentration of 20 mM was the optimum for Mn-dependent peroxidase production (88.00 U L-1), while the highest versatile peroxidase activity was detected in the medium with grapevine sawdust and 10 mM of nitrogen (80.80 U L-1). Wheat straw was the best carbon source for laccase synthesis in G. lucidum (55.75 U L-1).

Interaction of Trace Elements and Ligninolytic Enzymes in Pleurotus eryngii

Considering the fact that some trace elements have influence on the growth and physiological processes in the fungi, the aim of this study was to resolve the question of whether Fe, Zn, and Se, added to the medium in the different forms, affect activity of selected Pleurotus eryngii ligninolytic enzymes participating in mandarine peels submerged fermentation. Compared with the control, Fe and Zn showed stimulatory effect on laccase activity. Reduction in the activity was emphasized with cultivation time, but it was slower in the presence of Zn than in the presence of Fe. Among studied Se forms, SeO2 was the most unfavorable Se source for laccase activity. All tested trace elements, independent on form, showed stimulatory effect on the activity of Mn-oxidizing peroxidases on the seventh day of cultivation. Based on the results, it could be concluded that the presence of the tested trace elements can advance various biotechnological processes among which is biodegradation of agricultural wastes, potential environmental pollutants.

Wheat straw conversion by enzymatic system of Ganoderma lucidum

The purpose of this study was to resolve the question of whether various nitrogen sources and concentrations affect characteristics of selected G. lucidum ligninolytic enzymes participating in wheat straw fermentation. This is the first study reporting the presence of versatile peroxidase activity in crude extract of G. lucidum culture, as well as isoforms profile of Mn-oxidizing peroxidases. NH4NO3 was the optimum nitrogen source for laccase and Mn-dependent peroxidase activity, while peptone was the optimum one for versatile peroxidase activity. with laccase activity were obtained by native PAGE and IEF separations from medium enriched with inorganic nitrogen source, and only two bands from medium containing organic source. Medium composition was not shown to affect isoenzyme patterns of Mn-oxidizing peroxidases. Four isoforms of Mn-dependent peroxidase and three of versatile peroxidase were obtained on native PAGE. By IEF separation, five isoforms of Mn-dependent peroxidase and only two of versatile peroxidase were observed. The results demonstrated that G. lucidum has potential for mineralization and transformation of various agricultural residues and should take more significant participation in large-scale biotechnological processes.

P9 Clinical features and prognosis of hereditary breast cancer

The aim of the study was comparative analysis of ligninolytic enzyme productions of Trametes gibbosa and its ability to degrade wheat straw, as well as effect of trace elements on those processes. Mn-oxidizing peroxidases were the main enzymes and the tested trace elements affected their activities mainly inhibitory. Stimulatory effect on Mn-independent peroxidase had Mn2+ in all concentrations, while Zn2+ concentrations of 5.0 mM and 10.0 mM led to the maximum Mn-dependent peroxidase activity. The trace elements affected laccase activity differently. Zn2+ and Fe2+ (0.5 mM and 10.0 mM, respectively) had the greatest effect on the activity, while Mn2+ and Cu2+ at the amount of 10.0 mM completely stopped the enzyme production. The tested enzymes acted simultaneously on wheat straw degradation. The presence of 1.0 mM of Mn2+ caused the highest level of dry matter loss (43.1%), which was the consequence of hemicellulose, cellulose and lignin degradation in the rate of 62.6%, 35.6% and 61.2%, respectively. Selective lignin degradation was noted especially in the presence of Fe2+ at the concentration of 0.5 mM.

Intraspecific Diversity within Ganoderma lucidum in the Production of Laccase and Mn-Oxidizing Peroxidases During Plant Residues Fermentation

Comparison of the potential for laccase and Mn-oxidizing peroxidases synthesis by ten strains of Ganoderma lucidum, originating from different worldwide areas, during solid-state fermentation of selected plant raw materials was the aim of this study. The great intraspecific variability in the production of analyzed enzymes as well as the dependence of the enzyme activity on plant raw materials were reported. The strain HAI 957 was the best laccase producer in the presence of corn stem, as a unique carbon source (129.46 U/L). The highest level of Mn-dependent peroxidase activity was noted after wheat straw fermentation by G. lucidum HAI 246 (78.64 U/L), while the maximal versatile peroxidase production (59.72 U/L) was observed in strain HAI 957 in the medium with oak sawdust.

Isolation and selection of novel basidiomycetes for decolorization of recalcitrant dyes

Thirty wood-rotting basidiomycetes, most of them causing white rot in wood, were isolated from fruiting bodies growing on decaying wood from the Sierra de Ayllón (Spain). The fungi were identified on the basis of their morphological characteristics and compared for their ability to decolorize Reactive Black 5 and Reactive Blue 38 (as model of azo and phthalocyanine type dyes, respectively) at 75 and 150 mg/L. Only eighteen fungal strains were able to grow on agar plates in the presence of the dyes and only three species (Calocera cornea, Lopharia spadicea, Polyporus alveolaris) decolorized efficiently both dyes at both concentrations. The ligninolytic activities, involved in decolorization dyes (laccases, lignin peroxidases, Mn-oxidizing peroxidases), were followed in glucose basal medium in the presence of enzyme inducers. The results indicate a high variability of the ligninolytic system within white-rot basidiomycetes. These fungal species and their enzymes can represent new alternatives for the study of new biological systems to degrade aromatic compounds causing environmental problems.

Effect of cultivation conditions on ligninolytic enzyme production by Ganoderma carnosum

Ganoderma carnosum has been found in Europe only, at coniferous trees and it is difficult to distinguish it morphologically from G. lucidum. Since G. carnosum has not been studied biochemically yet, the aim of this study was to analyse its ability to produce Mn-dependent peroxidase (MnP), versatile peroxidase (VP) and laccase (Lac) under conditions of solid-state fermentation of selected plant raw materials as carbon sources, in the presence of different nitrogen concentrations in the medium. Wheat straw, corn stem, oak and grapevine sawdust were the analysed plant raw materials. Nitrogen source in synthetic medium was NH4NO3 and its concentrations were: 10mM N and 20 mM N. Enzyme activity was determined spectrophotometrically, using ABTS and phenol red, as the substrates for Lac and Mn-oxidizing peroxidases, respectively. Maximum level of MnP activity (56.82 U/l) was obtained in the medium with wheat straw and nitrogen concentration of 10 mM. Best carbon source for VP production was grapevine sawdust at nitrogen concentration of 10 mM (80.80 U/l). The obtained Lac activity was very low in the medium with wheat straw (1.80 U/l), while it was not detected in the presence of other three analyzed plant raw materials. Maximum of total protein content (0.06 mgml-1) was noted in the medium where oak sawdust was carbon source and nitrogen concentration was 20 mM.