Interactions of heavy metals with white-rot fungi

Purine nucleoside phosphorylases: properties, functions, and clinical aspects.

Publisher Summary Carbohydrate oxidases are found in all kingdoms of life but are mostly found in fungi. Their natural role is not always clear. Usage of molecular oxygen as electron acceptor is not a logical choice when the enzyme is part of a catabolic pathway. This chapter provides an overview of the occurrence and properties of carbohydrate oxidases. The physiological role of the different enzymes is discussed in relation to their origin, and the catalytic and structural properties are discussed in relation to their family background. It also provides a summary of the biocatalytic applications of carbohydrate oxidases. Carbohydrate oxidases are valuable enzymes for several applications. They are relatively stable and do not need expensive coenzymes. Carbohydrate oxidases are widely used in diagnostic applications, in the food and drinks industry, and for carbohydrate synthesis. They are also used for bleaching (production of H2O2) and as oxygen scavenger.

Occurrence and biocatalytic potential of carbohydrate oxidases.

The production of the H(2)O(2)-generating enzyme pyranose oxidase (POD) (EC 1.1.3.10) (synonym, glucose 2-oxidase), two ligninolytic peroxidases, and laccase in wood decayed by three white rot fungi was investigated by correlated biochemical, immunological, and transmission electron microscopic techniques. Enzyme activities were assayed in extracts from decayed birch wood blocks obtained by a novel extraction procedure. With the coupled peroxidase-chromogen (3-dimethylaminobenzoic acid plus 3-methyl-2-benzothiazolinone hydrazone hydrochloride) spectrophotometric assay, the highest POD activities were detected in wood blocks degraded for 4 months and were for Phanerochaete chrysosporium (149 mU g [dry weight] of decayed wood), Trametes versicolor (45 mU g), and Oudemansiella mucida (1.2 mU g), corresponding to wood dry weight losses of 74, 58, and 13%, respectively. Mn-dependent peroxidase activities in the same extracts were comparable to those of POD, while lignin peroxidase activity was below the detection limit for all fungi with the veratryl alcohol assay. Laccase activity was high with T. versicolor (422 mU g after 4 months), in trace levels with O. mucida, and undetectable in P. chrysosporium extracts. Evidence for C-2 specificity of POD was shown by thin-layer chromatography detection of 2-keto-d-glucose as the reaction product. By transmission electron microscopy-immunocytochemistry, POD was found to be preferentially localized in the hyphal periplasmic space of P. chrysosporium and O. mucida and associated with membranous materials in hyphae growing within the cell lumina or cell walls of partially and highly degraded birch fibers. An extracellular distribution of POD associated with slime coating wood cell walls was also noted. The periplasmic distribution in hyphae and extracellular location of POD are consistent with the reported ultrastructural distribution of H(2)O(2)-dependent Mn-dependent peroxidases. This fact and the dominant presence of POD and Mn-dependent peroxidase in extracts from degraded wood suggest a cooperative role of the two enzymes during white rot decay by the test fungi.

Pyranose Oxidase, a Major Source of H(2)O(2) during Wood Degradation by Phanerochaete chrysosporium, Trametes versicolor, and Oudemansiella mucida.

Lignin is the most abundant aromatic polymer in nature. It is synthesized by higher plants, reaching levels of 20–30% of the dry weight of woody tissue. Although white-rot fungi were long recognized as efficient lignin-degrading microbes, research on their enzymology and genetics was somewhat neglected until the past decade. The impetus for increased research interest can be traced to the discovery of “ligninases” and potential commerical applications in the pulp and paper industry and in the degradation of xenobiotics. In this chapter, we briefly summarize several decades of research on white-rot fungi, and readers are referred to several comprehensive reviews for additional background information. Recent developments in molecular biology and enzymology are emphasized. Areas where knowledge is incomplete are highlighted.

https://www.fpl.fs.fed.us/documnts/pdf2004/fpl_2004_cullen002.pdf

Enzymology and Molecular Biology of Lignin Degradation

Coriolus versicolor KY2912 grown on a medium containing glucose, sucrose or glycerol produced pyranose oxidase. Pyranose oxidase (glucose-2-oxidase) was purified by HPA-75 chromatography, Sepharose 4B and Sephadex G-100 gel filtration, and hydroxyapatite chromatography. The purified enzyme preparation showed a single protein band on acrylamide gel electrophoresis. The highest activity was obtained when D-glucose was employed as substrate and molecular oxygen as electron acceptor. The enzyme was most active at pH 6.2 and 50°C, stable in the pH region between 5.0 and 7.4, and the activity was completely lost above 70°C. The activity was inhibited by Ag+ , Cu2+ and PCMB. The enzyme contained FAD covalently bound to the polypeptide chain. The enzyme consisted of identical subunits with a molecular weight of 68,000, and showed a total molecular weight of 220,000.

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Purification and properties of pyranose oxidase from Coriolus versicolor.

Uricase activity was found in Enterobacter cloacae KY3074 grown on guanine, hypoxanthine, uric acid, and xanthine media. The enzyme was purified from cells grown on uric acid as a source of nitrogen. The purification procedure included ammonium sulfate fractionation, gel filtration on Sephadex G-150, and column chromatography on DEAE-cellulose and DEAE-Sephadex. The enzyme had a molecular weight of about 105,000 and was specific for uric acid. The optimum pH was around 9.5, and the activity was inhibited by the presence of potassium cyanide, Ag+ or Cu2+. This uricase can be used for estimation of uric acid.

Purification and properties of uricase from Enterobacter cloacae.

Utilization of bacterial xanthine oxidase for inorganic phosphorus determination.

Enterobacter cloacae KY 3074 grown in a medium containing xanthine, hypoxanthine, guanine, or their nucleosides and nucleotides produced xanthine oxidase. The purified enzyme preparation showed a major protein band and a few minor bands in acrylamide gel electrophoresis. Molecular oxygen was the most effective electron acceptor. Ferricyanide and 2,6-dichlorophenolindophenol also served as electron acceptors, but NAD and NADP did not. Xanthine and hypoxanthine were good substrates, and guanine was also an effective substrate. The activity was inhibited by Ag2+, Cu2+, PCMB, and ascorbate. The spectrum of the Enterobacter enzyme resembled that of some known xanthine oxidizing enzymes, and this suggests a similarity in the prosthetic groups of these enzymes. The molecular weight of the native enzyme and subunit was 128,000 and 69,000, respectively.

Purification and properties of xanthine oxidase from Enterobacter cloacae.

The substrate for xanthine oxidase such as xanthine, hypoxanthine, guanine, 2-hydroxypurine, etc. are determined by oxidizing the substrate by the action of xanthine oxidase derived from a microorganism and determining the formed hydrogen peroxide by the system for the determination of hydrogen peroxide comprising peroxidase and coloring reagent. The determination is carried out in one step by using the test composition for the determination of the substrate for xanthine oxidase which comprises xanthine oxidase and a system for the determination of hydrogen peroxide. The determination method can be applied to the determination of the compound which participates in the system wherein the substrate is stoichiometrically formed by the reaction or decomposition. When the system is an enzymatic reaction, the compound is determined in one step. Xanthine oxidase used in the present invention is different from known xanthine oxidase in molecular weight and is prepared by culturing a microorganism belonging to the genus Enterobacter.

Yozo Machida

Papers

Purification and properties of pyranose oxidase from Coriolus versicolor.

Purification and properties of uricase from Enterobacter cloacae.

Utilization of bacterial xanthine oxidase for inorganic phosphorus determination.

Purification and properties of xanthine oxidase from Enterobacter cloacae.

Method and test composition for the determination of the substrate for xanthine oxidase