Laccases of fungi attract considerable attention due to their possible involvement in the transformation of a wide variety of phenolic compounds including the polymeric lignin and humic substances. So far, more than a 100 enzymes have been purified from fungal cultures and characterized in terms of their biochemical and catalytic properties. Most ligninolytic fungal species produce constitutively at least one laccase isoenzyme and laccases are also dominant among ligninolytic enzymes in the soil environment. The fact that they only require molecular oxygen for catalysis makes them suitable for biotechnological applications for the transformation or immobilization of xenobiotic compounds.

Fungal laccases - occurrence and properties.

This review focuses on some important and challenging aspects of soil extracellular enzyme research. We report on recent discoveries, identify key research needs and highlight the many opportunities offered by interactions with other microbial enzymologists. The biggest challenges are to understand how the chemical, physical and biological properties of soil affect enzyme production, diffusion, substrate turnover and the proportion of the product that is made available to the producer cells. Thus, the factors that regulate the synthesis and secretion of extracellular enzymes and their distribution after they are externalized are important topics, not only for soil enzymologists, but also in the broader context of microbial ecology. In addition, there are many uncertainties about the ways in which microbes and their extracellular enzymes overcome the generally destructive, inhibitory and competitive properties of the soil matrix, and the various strategies they adopt for effective substrate detection and utilization. The complexity of extracellular enzyme activities in depolymerising macromolecular organics is exemplified by lignocellulose degradation and how the many enzymes involved respond to structural diversity and changing nutrient availabilities. The impacts of climate change on microbes and their extracellular enzymes, although of profound importance, are not well understood but we suggest how they may be predicted, assessed and managed. We describe recent advances that allow for the manipulation of extracellular enzyme activities to facilitate bioremediation, carbon sequestration and plant growth promotion. We also contribute to the ongoing debate as to how to assay enzyme activities in soil and what the measurements tell us, in the context of both traditional methods and the newer techniques that are being developed and adopted. Finally, we offer our collective vision of the future of extracellular enzyme research: one that will depend on imaginative thinking as well as technological advances, and be built upon synergies between diverse disciplines.

Soil enzymes in a changing environment: Current knowledge and future directions

▪ Abstract Manganese(IV) oxides produced through microbial activity, i.e., biogenic Mn oxides or Mn biooxides, are believed to be the most abundant and highly reactive Mn oxide phases in the environment. They mediate redox reactions with organic and inorganic compounds and sequester a variety of metals. The major pathway for bacterial Mn(II) oxidation is enzymatic, and although bacteria that oxidize Mn(II) are phylogenetically diverse, they require a multicopper oxidase-like enzyme to oxidize Mn(II). The oxidation of Mn(II) to Mn(IV) occurs via a soluble or enzyme-complexed Mn(III) intermediate. The primary Mn(IV) biooxide formed is a phyllomanganate most similar to δ-MnO2 or acid birnessite. Metal sequestration by the Mn biooxides occurs predominantly at vacant layer octahedral sites.

Biogenic manganese oxides: Properties and mechanisms of formation

Nonylphenol in the environment: A critical review on occurrence, fate, toxicity and treatment in wastewaters

Geomycology: biogeochemical transformations of rocks, minerals, metals and radionuclides by fungi, bioweathering and bioremediation

Fungi possess the biochemical and ecological capacity to degrade environmental organic chemicals and to decrease the risk associated with metals, metalloids and radionuclides, either by chemical modification or by influencing chemical bioavailability. Furthermore, the ability of these fungi to form extended mycelial networks, the low specificity of their catabolic enzymes and their independence from using pollutants as a growth substrate make these fungi well suited for bioremediation processes. However, despite dominating the living biomass in soil and being abundant in aqueous systems, fungi have not been exploited for the bioremediation of such environments. In this Review, we describe the metabolic and ecological features that make fungi suited for use in bioremediation and waste treatment processes, and discuss their potential for applications on the basis of these strengths.

Untapped potential: exploiting fungi in bioremediation of hazardous chemicals

Synthetic polymers, commonly named plastics, are among the most widespread anthropogenic pollutants of marine, limnic and terrestrial ecosystems. Disruptive effects of plastics are known to threaten wildlife and exert effects on natural food webs, but signs for and knowledge on plastic biodegradation are limited. Microorganisms are the most promising candidates for an eventual bioremediation of environmental plastics. Laboratory studies have reported various effects of microorganisms on many types of polymers, usually by enzymatic hydrolysis or oxidation. However, most common plastics have proved to be highly recalcitrant even under conditions known to favour microbial degradation. Knowledge on environmental degradation is yet scarcer. With this review, we provide a comprehensive overview of the current knowledge on microbiological degradation of several of the most common plastic types. Furthermore, we illustrate the analytical challenges concerning the evaluation of plastic biodegradation as well as constraints likely standing against the evolution of effective biodegradation pathways.

Prospects for microbiological solutions to environmental pollution with plastics

Research on freshwater fungi has concentrated on their role in plant litter decomposition in streams. Higher fungi dominate over bacteria in terms of biomass, production and enzymatic substrate degradation. Microscopy-based studies suggest the prevalence of aquatic hyphomycetes, characterized by tetraradiate or sigmoid spores. Molecular studies have consistently demonstrated the presence of other fungal groups, whose contributions to decomposition are largely unknown. Molecular methods will allow quantification of these and other microorganisms. The ability of aquatic hyphomycetes to withstand or mitigate anthropogenic stresses is becoming increasingly important. Metal avoidance and tolerance in freshwater fungi implicate a sophisticated network of mechanisms involving external and intracellular detoxification. Examining adaptive responses under metal stress will unravel the dynamics of biochemical processes and their ecological consequences. Freshwater fungi can metabolize organic xenobiotics. For many such compounds, terrestrial fungal activity is characterized by cometabolic biotransformations involving initial attack by intracellular and extracellular oxidative enzymes, further metabolization of the primary oxidation products via conjugate formation and a considerable versatility as to the range of metabolized pollutants. The same capabilities occur in freshwater fungi. This suggests a largely ignored role of these organisms in attenuating pollutant loads in freshwaters and their potential use in environmental biotechnology.

Fungi in freshwaters: ecology, physiology and biochemical potential

Laccases (EC 1.10.3.2) are extracellular multicopper oxidases produced by different kinds of fungi (39), which oxidize lignin and many organic xenobiotics (7, 23, 27, 45). These enzymes couple four one-electron substrate oxidations to the four-electron reduction of dioxygen to water, without formation of free reduced oxygen species (7, 45).

Manganese peroxidases (MnP; EC 1.11.1.13) are part of the ligninolytic system of white rot and litter-decaying basidiomycetes. During the catalytic cycle, the active center is oxidized by H2O2. Reduction to the resting enzyme is achieved by two successive one-electron transfers, thereby oxidizing Mn2+ to Mn3+, respectively. This is facilitated by fungal organic acids such as oxalate or malonate upon chelation of the highly reactive Mn3+ state (4, 20-22, 40, 43). MnP catalyzes the oxidation of lignin, humic substances, and many organopollutants (16, 20, 29).


Extracellular H2O2 is required as a substrate for ligninolytic peroxidases. Extracellular enzymes like aryl alcohol oxidase (31) and glyoxal oxidase (19) produce H2O2. This compound is also formed upon oxidation of hydroquinones by ligninolytic enzymes and autoxidation of the resulting semiquinones concomitantly reducing O2 to superoxide anion radical (13, 27). Mn2+ reduces superoxide to H2O2 and is thereby oxidized to Mn3+ (2, 27). Furthermore, superoxide may dismutate to H2O2 and O2. Oxidation of oxalate, glyoxylate, and malonate by Mn3+ was also considered to be a source of H2O2 (16, 42). For oxalate, the following reactions are well established (41, 42): 




(1)







(2)







(3)







(4)


For abiotic decomposition of malonate, the following reactions were proposed (16): 




(5)







(6)







(7)







(8)


Superoxide and oxalate derived from reaction 7 subsequently can contribute to reactions 1 to 4. Autocatalytic generation of traces of Mn3+, which leads to H2O2 (16) and the release of small H2O2 concentrations from fungal mycelia (42), was considered to initiate MnP reactions, thereby enhancing the Mn3+ concentration and facilitating H2O2 production in the aforementioned follow-up reactions.


In a previous paper, we have demonstrated that a purified laccase from the white rot fungus Trametes versicolor oxidized Mn2+ to Mn3+ in the presence of the Mn3+ chelator pyrophosphate (15). Mn2+ oxidation involved concomitant reduction of laccase type 1 copper, thus providing evidence that Mn2+ oxidation occurs via one-electron transfer to type 1 copper as usual for substrate oxidation by blue laccases (7, 45). A Phellinus ribis laccase devoid of type 1 copper did not oxidize Mn2+ in the presence of pyrophosphate (25). The litter-decaying basidiomycete Stropharia rugosoannulata degrades chlorophenols (36), the fluoroquinolone antibacterial drug ciprofloxacin (44), 2,4,6-trinitrotoluene (33), and synthetic lignin (38) to CO2 and H2O. Here, we show that a catalytic system consisting of purified laccase from S. rugosoannulata, Mn2+, and organic Mn chelators such as oxalate and malonate generates H2O2. We further assessed key aspects of the catalytic mechanism underlying this new reaction and the impact on MnP produced along with laccase by S. rugosoannulata under certain culture conditions. This study was attempted to provide evidence for a physiological role of laccase-catalyzed Mn2+ oxidation and establishes a novel type of laccase-MnP cooperation with potential significance for lignocellulose breakdown and degradation of xenobiotics.

Laccase-catalyzed oxidation of Mn(2+) in the presence of natural Mn(3+) chelators as a novel source of extracellular H(2)O(2) production and its impact on manganese peroxidase.

Because the endocrine disrupting effects of nonylphenol (NP) and octylphenol became evident, the degradation of long-chain alkylphenols (AP) by microorganisms was intensively studied. Most NP-degrading bacteria belong to the sphingomonads and closely related genera, while NP metabolism is not restricted to defined fungal taxa. Growth on NP and its mineralization was demonstrated for bacterial isolates, whereas ultimate degradation by fungi still remains unclear. While both bacterial and fungal degradation of short-chain AP, such as cresols, and the bacterial degradation of long-chain branched AP involves aromatic ring hydroxylation, alkyl chain oxidation and the formation of phenolic polymers seem to be preferential elimination pathways of long-chain branched AP in fungi, whereby both intracellular and extracellular oxidative enzymes may be involved. The degradation of NP by sphingomonads does not proceed via the common degradation mechanisms reported for short-chain AP, rather, via an unusual ipso-substitution mechanism. This fact underlies the peculiarity of long-chain AP such as NP isomers, which possess highly branched alkyl groups mostly containing a quaternary α-carbon. In addition to physicochemical parameters influencing degradation rates, this structural characteristic confers to branched isomers of NP a biodegradability different to that of the widely used linear isomer of NP. Potential biotechnological applications for the removal of AP from contaminated media and the difficulties of analysis and application inherent to the hydrophobic NP, in particular, are also discussed. The combination of bacteria and fungi, attacking NP at both the phenolic and alkylic moiety, represents a promising perspective.

Dietmar Schlosser

Papers

Untapped potential: exploiting fungi in bioremediation of hazardous chemicals

Prospects for microbiological solutions to environmental pollution with plastics

Fungi in freshwaters: ecology, physiology and biochemical potential

Laccase-catalyzed oxidation of Mn(2+) in the presence of natural Mn(3+) chelators as a novel source of extracellular H(2)O(2) production and its impact on manganese peroxidase.

Microbial degradation of nonylphenol and other alkylphenols—our evolving view