Non-phenolic Substrates Research Articles

Mechanism of non-phenolic substrate oxidation by the fungal laccase Type 1 copper site from Trametes versicolor: the case of benzo[a]pyrene and anthracene.

Laccases (EC 1.10.3.2) are multicopper oxidases with the capability to oxidize diverse phenolic and non-phenolic substrates. While the molecular mechanism of their activity towards phenolic substrates is well-established, their reactivity towards non-phenolic substrates, such as polycyclic aromatic hydrocarbons (PAHs), remains unclear. To elucidate the oxidation mechanism of PAHs, particularly the activation mechanism of the sp2 aromatic C-H bond, we conducted a density functional theory investigation on the oxidation of two PAHs (anthracene and benzo[a]pyrene) using an extensive model of the T1 copper catalytic site of the fungal laccase from Trametes versicolor.

Biochemical Characterization of Laccase from Spirulina CPCC-695 and Their Role in Estrone Degradation.

The addition of exogenous endocrine disrupting compounds (EDCs) like estrone, in the food chain through the aquatic system, disrupts steroid biosynthesis and metabolism by altering either the genomic or non-genomic pathway that eventually results in various diseases. Thus, bioremediation of these compounds is urgently required to prevent their addition and persistence in the environment. Enzymatic degradation has proven to be a knight in shining armour as it is safe and generates no toxic products. The multicopper oxidases (E.C. 1.10.3.2 benzenediol: oxygen oxidoreductase), laccase with the potential to degrade both phenolic and non-phenolic substrates has recently gained attention. In this study, the laccase was purified, characterized, and used to study estrone degradation. The culture filtrate (crude laccase) was concentrated and precipitated using cold-acetone and dialyzed against tris buffer (50mM) giving a four-fold partially purified form, with 45.56% yield and 204.14 U/mg as specific activity and a single peak at 250-300nm. The partially purified laccase was approximately 80kDa as estimated by SDS-PAGE preferred ABTS as substrate. Both crude and partially purified laccase showed maximum activity at pH 3.0, 40°C, and 4mM ABTS. Kinetic constants (Km, Vmax) of crude and partially purified laccase were found to be 0.83mM; 494.31mM/min, and 0.58mM; 480.54mM/min respectively. Iron sulphate and sodium azide inhibited laccase maximally. Crude and partially purified laccase degradation efficiency was 87.55 and 91.35% respectively. Spirulina CPCC-695 laccase with efficient estrone degradation ability renders them promising candidates for EDCs bioremediation.

Heterologous expression, purification, and characterization of thermo- and alkali-tolerant laccase-like multicopper oxidase from Bacillus mojavensis TH309 and determination of its antibiotic removal potential.

Laccases or laccase-like multicopper oxidases have great potential in bioremediation to oxidase phenolic or non-phenolic substrates. However, their inability to maintain stability in harsh environmental conditions and against non-substrate compounds is one of the main reasons for their limited use. The gene (mco) encoding multicopper oxidase from Bacillus mojavensis TH309 were cloned into pET14b( +), expressed in Escherichia coli, and purified as histidine tagged enzyme (BmLMCO). The molecular weight of the enzyme was about 60kDa. The enzyme exhibited laccase-like activity toward 2,6-dimethoxyphenol (2,6-DMP), syringaldazine (SGZ), and 2,2'-azino-bis (3-ethylbenzothiazoline-6-sulphonic acid) (ABTS). The highest enzyme activity was recorded at 80°C and pH 8. BmLMCO showed a half-life of ~ 305, 99, 50, 46, 36, and 20min at 40, 50, 60, 70, 80, and 90°C, respectively. It retained more than 60% of its activity after pre-incubation in the range of pH 5-12 for 60min. The enzyme activity significantly increased in the presence of 1mM of Cu2+. Moreover, BmLMCO tolerated various chemicals and showed excellent compatibility with organic solvents. The Michaelis constant (Km) and the maximum velocity (Vmax) values of BmLMCO were 0.98mM and 93.45µmol/min, respectively, with 2,6-DMP as the substrate. BmLMCO reduced the antibacterial activity of cefprozil, gentamycin, and erythromycin by 72.3 ± 1.5%, 79.6 ± 6.4%, and 19.7 ± 4.1%, respectively. This is the first revealing shows the recombinant production of laccase-like multicopper oxidase from any B. mojavensis strains, its biochemical properties, and potential for use in bioremediation.

Investigation of Abortiporus biennis lignocellulolytic toolbox, and the role of laccases in polystyrene degradation

White-rot basidiomycetes are the only microorganisms able to produce both hydrolytic (cellulases and hemicellulases) and oxidative (ligninolytic) enzymes for degrading all lignocellulose constituents. Their enzymatic machinery makes them ideal for the discovery of novel enzymes with desirable properties. In the present work, Abortiporus biennis, a white-rot fungus, was studied in regard to its lignocellulolytic potential. Secretomics and biochemical analyses were employed to study the strain's enzymatic arsenal, after growth in corn stover cultures and xylose-based defined media. The results revealed the presence of all the necessary enzymatic activities for complete breakdown of biomass, while the prominent role of oxidative enzymes in the lignocellulolytic strategy of the strain became evident. Two novel laccases, AbiLac1 and AbiLac2, were isolated from the culture supernatant with ion-exchange chromatography. Characterization of purified laccases revealed their ability to oxidize a wide variety of phenolic and non-phenolic substrates. AbiLac1 was found to oxidize polystyrene powder, showing high depolymerization potential, based on radical chain scission mechanism as evidenced by molecular weight decrease. The results of the present study demonstrate the biotechnological potential of the unexplored enzymatic machinery of white-rot basidiomycetes, including the design of improved lignocellulolytic cocktails, as well as the degradation and/or valorization of plastic waste materials.

Time-dependent electrochemical characteristics of a phenolic and non-phenolic compound in the presence of laccase/ABTS system.

The laccase/ABTS system has found several industrial applications ranging from biodeterioration to biodegradation and bioremediation. However, the capability of the laccase/ABTS system varies depending upon the type of substrate used. Voltammetric studies involving two widely used substrates, i.e., veratryl alcohol (VA) and alkali lignin (AL), were performed to gain new insight into the electrochemical behavior of the reactions. The individual electrochemical reactions established the differential nature of the two compounds over a concentration range, along with the mediator ABTS producing a distinguishing effect on their oxidative reactions, which was further studied over a 12hour period. It was followed by the reaction of both the compounds against the laccase/ABTS system that helped verify the role of the enzyme and the mediator in the electron transfer process and elucidate the mediated oxidations carried out by laccase against the phenolic and non-phenolic substrate through the process of cyclic voltammetry.

Biotechnological Applications of Manganese Peroxidases for Sustainable Management

Microbial manganese peroxidases (MnPs) are ligninolytic enzymes primarily responsible for degrading lignin, but they are also capable of degrading dyes, organic pollutants, and emerging contaminants in wastewater. They have been isolated and characterized from many white-rot fungi and few bacteria. For microbial MnPs, the optimum pH range is between 3.5 and 9.0; the optimum temperature range is between 25°C and 70°C. Their molecular weights range between 25 kDa and 68 kDa. By oxidizing Mn2+ to Mn3+, they can oxidize a wide range of phenolic and non-phenolic substrates. MnPs can be used in a wide range of industrial applications, including delignification of pulp, wastewater treatment, biofuel production, dye removal, biopulping, biobleaching, and juice extract clarification. The purpose of this review is to summarize biotechnological applications of manganese peroxidases.

Laccase-catalyzed conjugation of BSA mediated by gallic acid: Preparation, characterization, and antioxidant activity

Laccase is one of the enzymes that catalyze the oxidation of phenolic and non-phenolic substrates and show encouraging potential as a biocatalyst in the synthesis of bioactive compounds. It is known that phenolic acids have an antioxidant effect. Bovine serum albumin (BSA) shows gelling activity, and nutraceutical binding ability but it does not show antioxidant activity. In this study, BSA which has no antioxidant activity using laccase, started to show antioxidant activity with gallic acid (GA) conjugation. The synthesized conjugates were analyzed by polyacrylamide gel electrophoresis (PAGE), ultraviolet–visible spectrophotometry (UV-Vis), and Fourier-transform infrared spectroscopy (FTIR). Radical scavenging capacity for antioxidant activity was measured. GA-functionalized-BSA displayed greatly improved 2,2'-azinobis-(3- ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 1,1-diphenyl-2- picryl-hydrazyl (DPPH) radical scavenging capacities, compared with the untreated BSA. Protein–flavonoid conjugates can improve the natural properties of proteins, being promising products to be used in medical, food and polymer fields where antioxidant ability is an essential feature.

Eminent Industrial and Biotechnological Applications of Laccases from Bacterial Source: a Current Overview.

Laccases are blue multicopper oxidases that oxidize a wide range of phenolic as well as non-phenolic substrates in the presence or absence of mediators. They occur in various species of bacteria, fungi, insects, and plants; bacterial laccases show high substrate specificity. Bacteria produce these enzymes either extracellularly or intracellularly and exhibit stability to a wide range of pH and temperature. Therefore, they are suitable for various industrial processes such as food, textile, and paper and pulp industry. They are also valuable for producing biofuels, pharmaceuticals, biosensors, and degradation of various environmental pollutants and xenobiotics compounds. Since bacterial laccases are more versatile in the sense of nutritional needs and ecological factors, their use can provide a promising solution to various problems related to industry and the field of biotechnology. However, there is a need for a thorough understanding of the chemistry and activity of bacterial laccases to enable their full potential use in bioremediation and biofuel production.

A novel Streptomyces strain isolated by functional bioprospecting for laccases

In discovering novel laccase enzymes with interesting catalytic and stability properties, there is still scope for improvement in exploring new sites in Algeria. The bio-prospecting approach allowed the isolation of a novel strain, Streptomyces sp. HBD30 from the Agrioun River, northern Algeria. Following a functional approach, two laccases (LacI and LacII) were purified and characterised, showing appealing properties as thermostable enzymes, and oxidised typical phenolic and non-phenolic substrates with a higher affinity towards syringaldazine, followed by 2,6-dimethoxyphenol and 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid). Further analyses revealed that the Streptomyces sp. HDB30 genome contained an array of genes coding for carbohydrate active enzymes (CAZome) and lignin-degrading enzymes to degrade lignocellulose. In addition to enlarging the repertoire of thermostable laccases, this study uncovered the metabolic potential of a new Streptomyces strain, which is useful for a wide range of applications.

Production of Recombinant Laccase From Coprinopsis cinerea and Its Effect in Mediator Promoted Lignin Oxidation at Neutral pH.

Laccases are multi-copper oxidases that use molecular oxygen as the electron acceptor to oxidize phenolic and indirectly also non-phenolic substrates by mechanisms involving radicals. Due to their eco-friendliness and broad substrate specificity, laccases span a wide range of biotechnological applications. We have heterologously expressed a laccase from the coprophilic basidiomycete Coprinopsis cinerea (CcLcc9) in the methylotrophic yeast Pichia pastoris. The recombinant CcLcc9 (rCcLcc9) oxidized 2,6-dimethoxyphenol in the neutral pH range, and showed thermostability up to 70°C. The rCcLcc9 efficiently oxidized veratryl alcohol to veratraldehyde in the presence of low molecular weight mediators syringyl nitrile, methyl syringate and violuric acid, which are syringyl-type plant phenolics that have shown potential as natural co-oxidants for lignocellulosic materials. In addition, rCcLcc9 is able to depolymerize biorefinery hardwood lignin in the presence of methyl syringate and syringyl nitrile as indicated by gel permeation chromatography, and infrared spectral and nucleic magnetic resonance analyses. Furthermore, we showed that several added-value aromatic compounds, such as vanillin, vanillic acid, syringaldehyde, syringic acid and p-hydroxybenzoic acid, were formed during sequential biocatalytic chemical degradation of biorefinery lignin, indicating that rCcLcc9 harbors a great potential for sustainable processes of circular economy and modern biorefineries.

Rational engineering of the lccβ T. versicolor laccase for the mediator-less oxidation of large polycyclic aromatic hydrocarbons

Laccases are among the most sought-after biocatalyst for many green applications, from biosensors to pollution remedial, because they simply need oxygen from the air to oxidize and degrade a broad range of substrates. However, natural laccases cannot process large and toxic polycyclic aromatic hydrocarbons (PAHs) except in the presence of small molecules, called mediators, which facilitate the reaction but are inconvenient for practical on-field applications. Here we exploited structure-based protein engineering to generate rationally modified fungal laccases with increased ability to process bulky PAHs even in a mediator-less reaction. Computational simulations were used to estimate the impact of mutations in the enzymatic binding pocket on the ability to bind and oxidize a selected set of organic compounds. The most promising mutants were produced and their activity was evaluated by biochemical assays with phenolic and non-phenolic substrates. Mutant laccases engineered with a larger binding pocket showed enhanced activity (up to ~ 300% at pH 3.0) in a wider range of pH values (3.0–8.0) in comparison to the wild type enzyme. In contrast to the natural laccase, these mutants efficiently degraded bulky and harmful triphenylmethane dyes such as Ethyl Green (up to 91.64% after 24 h), even in the absence of mediators, with positive implications for the use of such modified laccases in many green chemistry processes (e.g. wastewater treatment).

Optimization and scale-up of laccase production by Bacillus sp. BAB-4151 isolated from the waste of the soap industry

Laccase is also known as the blue copper oxidase enzyme. Owing to its ability to oxidizes a wide variety of phenolic and non-phenolic substrates, this enzyme has many biotechnological applications. It is produced by plants, fungi, and a wide variety of bacteria. However, bacterial laccases have been studied more due to their merits over other sources. Production of bacterial laccase at the commercial level demands high yielding cultures and optimization of physicochemical parameters for more yields in a shorter period. Hence, the present research was aimed to isolate the potent laccase producing bacteria and to optimize cultural conditions for higher production of laccase. In this study, six laccase positive bacterial cultures were isolated from the waste of the soap industry in Kadi, Gujarat, India. One isolate later identified as Bacillus sp. BAB-4151 produced copious amounts of laccase. The production process of laccase was optimized at shake flask level by using one variable at a time (OVAT) method and scaled-up to 5 L capacity bioreactor. The isolate produced a higher yield of laccase under diverse physicochemical and nutritional conditions. The OVAT approach resulted in 2.9 folds improvement in the yield of laccase, which further increased by 1.4 fold during scale-up at a laboratory-scale bioreactor. The ability of the organism to produce laccase over of range of physico-chemical conditions and to retain the productivity during scale up of the process at bioreactor level suggests that the organism can be of great commercial significance.

Laccases in Food Industry: Bioprocessing, Potential Industrial and Biotechnological Applications.

Laccase is a multi-copper oxidase that catalyzes the oxidation of one electron of a wide range of phenolic compounds. The enzyme is considered eco-friendly because it requires molecular oxygen as co-substrate for the catalysis and it yields water as the sole by-product. Laccase is commonly produced by fungi but also by some bacteria, insects and plants. Due it is capable of using a wide variety of phenolic and non-phenolic substrates, laccase has potential applications in the food, pharmaceutical and environmental industries; in addition, it has been used since many years in the bleaching of paper pulp. Fungal laccases are mainly extracellular enzyme that can be recovered from the residual compost of industrial production of edible mushrooms as Agaricus bisporus and Pleurotus ostreatus. It has also been isolated from microorganisms present in wastewater. The great potential of laccase lies in its ability to oxidize lignin, one component of lignocellulosic materials, this feature can be widely exploited on the pretreatment for agro-food wastes valorization. Laccase is one of the enzymes that fits very well in the circular economy concept, this concept has more benefits over linear economy; based on “reduce-reuse-recycle” theory. Currently, biorefinery processes are booming due to the need to generate clean biofuels that do not come from oil. In that sense, laccase is capable of degrading lignocellulosic materials that serve as raw material in these processes, so the enzyme’s potential is evident. This review will critically describe the production sources of laccase as by-product from food industry, bioprocessing of food industry by-products using laccase, and its application in food industry.

Biomedical and Pharmaceutical-Related Applications of Laccases.

The oxidation of a vast range of phenolic and non-phenolic substrates has been catalyzed by laccases. Given a wide range of substrates, laccases can be applied in different biotechnological applications. The present review was conducted to provide a broad context in pharmaceutical- and biomedical- related applications of laccases for academic and industrial researchers. First, an overview of biological roles of laccases was presented. Furthermore, laccase-mediated strategies for imparting antimicrobial and antioxidant properties to different surfaces were discussed. In this review, laccase-mediated mechanisms for endowing antimicrobial properties were divided into laccase-mediated bio-grafting of phenolic compounds on lignocellulosic fiber, chitosan and catheters, and laccase-catalyzed iodination. Accordingly, a special emphasis was placed on laccase-mediated functionalization for creating antimicrobials, particularly chitosan-based wound dressings. Additionally, oxidative bio-grafting and oxidative polymerization were described as the two main laccase-catalyzed reactions for imparting antioxidant properties. Recent laccase-related studies were also summarized regarding the synthesis of antibacterial and antiproliferative agents and the degradation of pharmaceuticals and personal care products.

Phenolic and non-phenolic substrates oxidation by laccase at variable oxygen concentrations: Selection of bisubstrate kinetic models from polarographic data

Using laccase as a case study, an alternative methodology for discrimination of bisubstrate mechanism types, useful for Bi-Bi reactions when water is one of the products, is presented. A purified enzyme from fungal cultures of Ganoderma resinaceum and two reducing substrates: 2,2´-azino-bis(3-ethylbenzthiazoline-6-sulphonic acid) (ABTS) and 2,6-dimethoxyphenol (DMP) are used. Kinetic assays of ABTS or DMP oxidation using variable initial concentrations of dioxygen are performed in a closed system and depletion rates of dioxygen are measured using polarography. Seven bisubstrate models based on Michaelis-Menten Bi-Bi mechanisms are applied to the experimental data. Akaike information criterion, a penalized likelihood analysis, and Akaike weights are used for model selection. Results show that the more likely bisubstrate models for ABTS and DMP oxidation are ping-pong and Theorell-Chance, respectively. However, the catalytic efficiency of oxygen reduction to water exhibits similar values, and is compatible with a process relatively independent of reducing substrates and the bisubstrate mechanism type. Finally, Michaelis-Menten constants towards dioxygen, 233 μM with ABTS and 700 μM with DMP, indicate that the soluble electron acceptor available for laccase-catalyzed reactions, is far removed from saturation.

Cohnella sp. A01 laccase: thermostable, detergent resistant, anti-environmental and industrial pollutants enzyme

Laccase (EC 1.10.3.2; benzenediol; oxygen oxidoreductases) is a multi-copper oxidase that catalyzes the oxidation of phenols, polyphenols, aromatic amines, and different non-phenolic substrates with concomitant reduction of O2 to H2O. Enzymatic oxidation techniques have the potential of implementation in different areas of industrial fields. In this study, the Cohnella sp. A01 laccase gene was cloned into pET-26 (b+) vector and was transformed to E. coli BL21. Then it was purified using His tag affinity (Ni sepharose resin) chromatography. The estimated molecular weight was approximately 60 kDa using SDS-PAGE. The highest enzyme activity and best pH for 2,6-dimethoxyphenol (DMP) oxidation were recorded as 8 at 90 °C respectively. The calculated half-life and kinetic values including Km, Vmax, turn over number (kcat), and catalytic efficiency (kcat/Km) of the enzyme were 106 min at 90 °C and 686 μM, 10.69 U/ml, 20.3 S−, and 0.029 s−1 μM−1, respectively. The DMP was available as the substrate in all the calculations. Enzyme activity enhanced in the presence of Cu2+, NaCl, SDS, n-hexane, Triton X-100, tween 20, and tween 80, significantly. The binding residues were predicted and mapped upon the modeled tertiary structure of identified laccase. The remaining activity and structural properties of Cohnella sp. A01 laccase in extreme conditions such as high temperatures and presence of metals, detergents, and organic solvents suggest the potential of this enzyme in biotechnological and industrial applications. This process has been patented in Iranian Intellectual Property Centre under License No: 91325.

A novel starch-binding laccase from the wheat pathogen Zymoseptoria tritici highlights the functional diversity of ascomycete laccases

BackgroundLaccases are multicopper oxidases, which are assigned into auxiliary activity family 1 (AA1) in the CAZy database. These enzymes, catalyzing the oxidation of phenolic and nonphenolic substrates coupled to reduction of O2 to H2O, are increasingly attractive as eco-friendly oxidation biocatalysts. Basidiomycota laccases are well characterized due to their potential in de-lignification of lignocellulose. By contrast, insight into the biochemical diversity of Ascomycota counterparts from saprophytes and plant pathogens is scarce.ResultsHere, we report the properties of the laccase from the major wheat pathogen Zymoseptoria tritici (ZtrLac1A), distinguished from common plant fungal pathogens by an apoplastic infection strategy. We demonstrate that ZtrLac1A is appended to a functional starch-binding module and displays an activity signature disfavoring relatively apolar phenolic redox mediators as compared to the related biochemically characterized laccases. By contrast, the redox potential of ZtrLac1A (370 mV vs. SHE) is similar to ascomycetes counterparts. The atypical specificity is consistent with distinctive sequence substitutions and insertions in loops flanking the T1 site and the enzyme C-terminus compared to characterized laccases.ConclusionsZtrLac1A is the first reported modular laccase appended to a functional starch-specific carbohydrate binding module of family 20 (CBM20). The distinct specificity profile of ZtrLac1A correlates to structural differences in the active site region compared to previously described ascomycetes homologues. These differences are also highlighted by the clustering of the sequence of ZtrLac1A in a distinct clade populated predominantly by plant pathogens in the phylogenetic tree of AA1 laccases. The possible role of these laccases in vivo merits further investigations. These findings expand our toolbox of laccases for green oxidation and highlight the binding functionality of CBM-appended laccases as versatile immobilization tags.

Enhancement of laccase activity by pre-incubation with organic solvents

Laccases that are tolerant to organic solvents are powerful bio-catalysts with broad applications in biotechnology. Most of these uses must be accomplished at high concentration of organic solvents, during which proteins undergo unfolding, thereby losing enzyme activity. Here we show that organic-solvent pre-incubation provides effective and reversible 1.5- to 4.0-fold enhancement of enzyme activity of fungal laccases. Several organic solvents, including acetone, methanol, ethanol, DMSO, and DMF had an enhancement effect among all laccases studied. The enhancement was not substrate-specific and could be observed by using both phenolic and non-phenolic substrates. Laccase preincubated with organic solvents was sensitive to high temperature but remained stable at 25 °C, for an advantage for long-term storage. The acetone-pre-incubated 3-D structure of DLac, a high-efficiency fungal laccase, was determined and confirmed that the DLac protein structure remains intact and stable at a high concentration of organic solvent. Moreover, the turnover rates of fungal laccases were improved after organic-solvent pre-incubation, with DLac showing the highest enhancement among the fungal laccases examined. Our investigation sheds light on improving fungal laccase usage under extreme conditions and extends opportunities for bioremediation, decolorization, and organic synthesis.

Recovery of laccase-producing gammaproteobacteria from wastewater

Wastewater environment is a rich source of microorganisms with the capability for the degradation of malicious aromatic pollutants. Although wastewater could be regarded as both a resource and a problem, we intended to elucidate its beneficial aspect in this study sourcing for laccase-producing proteobacteria. Different wastewater samples, from selected wastewater treatment plants (WWTPs), were selectively enriched with some model compounds (vanillin, lignin and potassium hydrogen phthalate) to screen out bacterial isolates that possess excellent degradation potentials. Thereafter, positive isolates were screened for the production of laccase and degradation on phenolic (guaiacol, α-naphthol and syringaldazine) and non-phenolic (ABTS; 2,2 azino-bis -(3-ethylbenzothiazoline 6 sulphonic acid) and PFC; potassium ferrocyanoferrate) substrates characteristic of laccase oxidation. Remarkable laccase producers were identified based on their 16 S rRNA sequences and their secreted enzymes were subjected to substrate specificity test, employing laccase substrates; ABTS, PFC, guaiacol, α-naphthol, 2,6-dimethoxyphenol and pyrogallol. Results showed that wastewater and selective enrichment, in tandem, produced the gammaproteobacteria Pseudomonas aeruginosa DEJ16, Pseudomonas mendocina AEN16 and Stenotrophomonas maltophila BIJ16, which preferably oxidized the non-phenolic substrates. Units of extracellular laccase activity ranging between cca. 490 and cca. 600 U/mL were recorded for ABTS whereas outputs recorded from PFC catalysis ranged from cca. 320 to cca. 430 U/mL. Stenotrophomonas maltophila BIJ16 presented an unparalleled high laccase activity and had a responsive substrate specificity to aromatic and inorganic substrates, thereby suggesting its employment for in situ biodegradation studies. In conclusion, wastewater serves as an ideal milieu for the isolation of laccase producing bacteria.

Development of an ISFET-based enzyme assay forp-diphenol oxidase from locally isolated fungusSchizophyllum communeMF-O5

p-Diphenol oxidases (PDO) are copper-containing enzymes that catalyze the oxidation of a wide range of non-phenolic substrates and phenolic compounds utilizing oxygen as the acceptor of electrons released during enzymatic reactions.