Philip J. Kersten

Bio: Philip J. Kersten is an academic researcher from United States Department of Agriculture. The author has contributed to research in topics: Phanerochaete & Chrysosporium. The author has an hindex of 26, co-authored 39 publications receiving 3082 citations. Previous affiliations of Philip J. Kersten include United States Forest Service & University of Wisconsin-Madison.

Comparison of lignin peroxidase, horseradish peroxidase and laccase in the oxidation of methoxybenzenes.

Abstract: Lignin peroxidase oxidizes non-phenolic substrates by one electron to give aryl-cation-radical intermediates, which react further to give a variety of products. The present study investigated the possibility that other peroxidative and oxidative enzymes known to catalyse one-electron oxidations may also oxidize non-phenolics to cation-radical intermediates and that this ability is related to the redox potential of the substrate. Lignin peroxidase from the fungus Phanerochaete chrysosporium, horseradish peroxidase (HRP) and laccase from the fungus Trametes versicolor were chosen for investigation with methoxybenzenes as a homologous series of substrates. The twelve methoxybenzene congeners have known half-wave potentials that differ by as much as approximately 1 V. Lignin peroxidase oxidized the ten with the lowest half-wave potentials, whereas HRP oxidized the four lowest and laccase oxidized only 1,2,4,5-tetramethoxybenzene, the lowest. E.s.r. spectroscopy showed that this congener is oxidized to its cation radical by all three enzymes. Oxidation in each case gave the same products: 2,5-dimethoxy-p-benzoquinone and 4,5-dimethoxy-o-benzoquinone, in a 4:1 ratio, plus 2 mol of methanol for each 1 mol of substrate. Using HRP-catalysed oxidation, we showed that the quinone oxygen atoms are derived from water. We conclude that the three enzymes affect their substrates similarly, and that whether an aromatic compound is a substrate depends in large part on its redox potential. Furthermore, oxidized lignin peroxidase is clearly a stronger oxidant than oxidized HRP or laccase. Determination of the enzyme kinetic parameters for the methoxybenzene oxidations demonstrated further differences among the enzymes.

Comparative Transcriptome and Secretome Analysis of Wood Decay Fungi Postia placenta and Phanerochaete chrysosporium

Abstract: Cellulose degradation by brown rot fungi, such as Postia placenta, is poorly understood relative to the phylogenetically related white rot basidiomycete, Phanerochaete chrysosporium To elucidate the number, structure, and regulation of genes involved in lignocellulosic cell wall attack, secretome and transcriptome analyses were performed on both wood decay fungi cultured for 5 days in media containing ball-milled aspen or glucose as the sole carbon source Using liquid chromatography-tandem mass spectrometry (LC-MS/MS), a total of 67 and 79 proteins were identified in the extracellular fluids of P placenta and P chrysosporium cultures, respectively Viewed together with transcript profiles, P chrysosporium employs an array of extracellular glycosyl hydrolases to simultaneously attack cellulose and hemicelluloses In contrast, under these same conditions, P placenta secretes an array of hemicellulases but few potential cellulases The two species display distinct expression patterns for oxidoreductase-encoding genes In P placenta, these patterns are consistent with an extracellular Fenton system and include the upregulation of genes involved in iron acquisition, in the synthesis of low-molecular-weight quinones, and possibly in redox cycling reactions

Glyoxal oxidase of Phanerochaete chrysosporium: its characterization and activation by lignin peroxidase.

Abstract: Glyoxal oxidase (GLOX) is an extracellular H2O2-generating enzyme produced by ligninolytic cultures of Phanerochaete chrysosporium. The production, purification, and partial characterization of GLOX from agitated cultures are described here. High-oxygen levels are critical for GLOX production as for lignin peroxidase. GLOX purified by anion-exchange chromatography appears homogeneous by NaDod-SO4/PAGE (molecular mass = 68 kDa). However, analysis by isoelectric focusing indicates two major bands (pI 4.7 and 4.9) that stain as glycoproteins as well as for H2O2-producing activity in the presence of methylglyoxal. Purified GLOX shows a marked stimulation in activity when incubated with Cu2+; full activation takes more than 1 hr with 1 mM CuSO4 at pH 6. The steady-state kinetic parameters for the GLOX oxidation of methylglyoxal, glyceraldehyde, dihydroxyacetone, glycolaldehyde, acetaldehyde, glyoxal, glyoxylic acid, and formaldehyde, were determined by using a lignin peroxidase coupled-assay at pH 4.5. Of these substrates, the best is the extracellular metabolite methylglyoxal with a Km of 0.64 mM an apparent rate of catalysis, kcat, of 198 s1 under air-saturated conditions. The Km for oxygen is greater than the concentration of oxygen possible at ambient pressure--i.e., >1.3 mM at 25 degrees C. Importantly, oxygen-uptake experiments show that purified GLOX is inactive unless coupled to the peroxidase reaction. With this coupled reaction, for each mol of methylglyoxal, veratryl alcohol (a lignin peroxidase substrate), and oxygen consumed, 1 mol each of pyruvate and veratraldehyde is produced. The importance of these results is discussed in relation to the physiology of lignin biodegradation and possible extracellular regulatory mechanisms for the control of oxidase and peroxidase activities.

Common and uncommon cytochrome P450 reactions related to metabolism and chemical toxicity.

Abstract: Cytochrome P450 (P450) enzymes catalyze a variety of reactions and convert chemicals to potentially reactive products as well as make compounds less toxic. Most of the P450 reactions are oxidations. The majority of these can be rationalized in the context of an FeO3+ intermediate and odd electron abstraction/rebound mechanisms; however, other iron−oxygen complexes are possible and alternate chemistries can be considered. Another issue regarding P450-catalyzed reactions is the delineation of rate-limiting steps in the catalytic cycle and the contribution to reaction selectivity. In addition to the rather classical oxidations, P450s also catalyze less generally discussed reactions including reduction, desaturation, ester cleavage, ring expansion, ring formation, aldehyde scission, dehydration, ipso attack, one-electron oxidation, coupling reactions, rearrangement of fatty acid and prostaglandin hydroperoxides, and phospholipase activity. Most of these reactions are rationalized in the context of high-valent...

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

Abstract: 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.

The paleozoic origin of enzymatic lignin decomposition reconstructed from 31 fungal genomes

Abstract: Wood is a major pool of organic carbon that is highly resistant to decay, owing largely to the presence of lignin. The only organisms capable of substantial lignin decay are white rot fungi in the Agaricomycetes, which also contains non-lignin-degrading brown rot and ectomycorrhizal species. Comparative analyses of 31 fungal genomes (12 generated for this study) suggest that lignin-degrading peroxidases expanded in the lineage leading to the ancestor of the Agaricomycetes, which is reconstructed as a white rot species, and then contracted in parallel lineages leading to brown rot and mycorrhizal species. Molecular clock analyses suggest that the origin of lignin degradation might have coincided with the sharp decrease in the rate of organic carbon burial around the end of the Carboniferous period.

Lignin peroxidase of Phanerochaete chrysosporium

Abstract: Publisher Summary Several procedures have been described for growing Phanerochaete chrysosporiurn for ligninase production. These procedures differ somewhat in the medium formulation and types of growth vessels: (1) shallow stationary cultures, (2) agitated liquid cultures, and (3) rotating biological contactors (RBCs; disk fermenters). The recently developed use of agitated culture for production of ligninase permits easier “scale up.” Although ligninase can be produced in agitated flask cultures, the reliable use of stirred tank fermenters awaits further development, which is ongoing in several laboratories. This chapter describes the production of ligninase in shallow stationary cultures and in agitated cultures. Shallow stationary cultures (10 ml) are grown in rubber-stoppered, 125-ml Edenmyer flasks at 39 ° under 100% oxygen. The stationary cultures give somewhat more reliable and reproducible results than the agitated cultures.