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Michael H. Gold - One of the best experts on this subject based on the ideXlab platform.

  • Homologous expression of Phanerochaete chrysosporium manganese peroxidase, using bialaphos resistance as a dominant selectable marker
    Current Genetics, 2003
    Co-Authors: Biao Ma, Mary B. Mayfield, Michael H. Gold
    Abstract:

    Manganese peroxidase (MnP) is a major extracellular component of the lignin-degrading system of the white-rot fungus, Phanerochaete chrysosporium . Homologous expression of recombinant MnP isozyme 1 (rMnP1) in P. chrysosporium was achieved using a novel transformation system for this fungus, which utilizes the Streptomyces hygroscopicus bialaphos-resistant gene, bar , as the selectable marker. The transformation frequency for this system is approximately 100 bialaphos-resistant transformants per microgram of plasmid DNA. Transformed strains all contain plasmid DNA, ectopically integrated into the fungal genome. Using this transformation system, the promoter region of the P. chrysosporium translation elongation factor gene was used to drive expression of mnp1 , encoding MnP1, in primary metabolic cultures of P. chrysosporium , where endogenous MnP was not expressed. Approximately 2–3 mg of active recombinant MnP1 per liter of extracellular medium was produced in agitated cultures of transformants.

  • degradation of pentachlorophenol by Phanerochaete chrysosporium intermediates and reactions involved
    Microbiology, 2000
    Co-Authors: Vijay Bhasker G Reddy, Michael H. Gold
    Abstract:

    Under nitrogen-limiting, secondary metabolic conditions, the lignin-degrading basidiomycete Phanerochaete chrysosporium rapidly degrades pentachlorophenol. The pathway for the degradation of pentachlorophenol has been elucidated by the characterization of fungal metabolites and oxidation products generated by purified lignin peroxidase (LiP) and manganese peroxidase (MnP). The multi-step pathway is initiated by a LiP- or MnP-catalysed oxidative dechlorination reaction to produce tetrachloro-1,4-benzoquinone. Under primary or secondary metabolic conditions, the quinone is further degraded by two parallel pathways with cross-links. The quinone is reduced to tetrachlorodihydroxybenzene, which can undergo four successive reductive dechlorinations to produce 1,4-hydroquinone, and the latter is o-hydroxylated to form the final aromatic metabolite, 1,2,4-trihydroxybenzene. Alternatively, the tetrachloro-1,4-benzoquinone is converted, either enzymically or nonenzymically, to 2,3,5-trichlorotrihydroxybenzene, which undergoes successive reductive dechlorinations to produce 1,2,4-trihydroxybenzene. Finally, at several points, hydroxylation reactions convert chlorinated dihydroxybenzenes to chlorinated trihydroxybenzenes, linking the two pathways at each of these steps. Presumably, the 1,2,4-trihydroxybenzene produced in each pathway is ring-cleaved with subsequent degradation to CO2. In contrast to the oxidative dechlorination step, the reductive dechlorinations and hydroxylations occur during both primary and secondary metabolic growth. Apparently, all five chlorine atoms are removed from the substrate prior to ring cleavage.

  • homologous expression of recombinant lignin peroxidase in Phanerochaete chrysosporium
    Applied and Environmental Microbiology, 1994
    Co-Authors: Mary Mayfield, Margaret Alic, K Kishi, Michael H. Gold
    Abstract:

    The white rot basidiomycete Phanerochaete chrysosporium has been the focus of numerous studies on the degradation of lignin (6, 15, 22) and aromatic pollutants (5, 17). Two peroxidases, manganese peroxidase (MnP) and lignin peroxidase (LiP), along with an extracellular H2O2-generating system, are thought to be the major extracellular components of the lignin-degrading system (14, 18, 22) of this organism. Both MnP and LiP occur as a series of isozymes encoded by a family of genes which are expressed under secondary metabolic growth conditions (9, 12, 14). The major isozymes, MnP1 (H3) and LiPH8, have been characterized in detail (14), and the X-ray structures of MnP1 (38) and LiPH8 (30, 31) have been reported. In addition, a homologous expression system (28) and several heterologous expression systems for MnP have been established (37, 41), allowing structure-function studies of mutant MnPs (24, 25, 42). In contrast, the efficient expression of active recombinant LiPH8 (rLiPH8) has not been achieved. The use of Escherichia coli as a LiP expression host has resulted in expression; however, refolding of denatured LiP from E. coli inclusion bodies resulted in the isolation of active rLiPH8 (10) and rLiPH2 (29) in relatively low yield. In addition, neither isozyme was glycosylated and, in one case, the recombinant protein contained seven extra N-terminal amino acids (10). In this paper, we report the first successful homologous expression of rLiPH8 in P. chrysosporium and the characterization of the recombinant enzyme.

  • isolation and transformation of uracil auxotrophs of the lignin degrading basidiomycete Phanerochaete chrysosporium
    Current Genetics, 1993
    Co-Authors: Lakshmi Akileswaran, Margaret Alic, Edith K Clark, Jason L Hornick, Michael H. Gold
    Abstract:

    Uracil auxotrophs of Phanerochaete chrysosporium were isolated using 5-fluoroorotate resistance as a selection scheme. The ura3 auxotrophs deficient in orotidylate decarboxylase and ura5 auxotrophs deficient in orotate phosphoribosyl transferase were characterized by enzyme assays and complementation tests. The ura5 auxotrophs were transformed to prototrophy with the ura5 gene from the ascomycete Podospora anserina. The ura3 auxotrophs were transformed to prototrophy with the ura3 gene from the basidiomycete Schizophyllum commune. The P. chrysosporium ura3 gene was isolated from a γEMBL3 genomic library using the S. commune ura3 gene as a probe. A 6.6-kb fragment incorporating the ura3 gene was subcloned into Bluescript SK+(pURA3.1) and used to transform P. chrysosporium ura3 auxotrophic strains. The pURA3.1 insert was mapped for restriction sites and the approximate location of the ura3 gene within the insert was determined. Double auxotrophic strains were transformed with either of two marker genes and the resulting single auxotrophic strains were crossed to demonstrate genetic recombination between two nuclei of identical genetic background.

Laila, Maulida Husni - One of the best experts on this subject based on the ideXlab platform.

  • PENGARUH PENGGUNAAN LIMBAH SAWIT DAN DEDAK PADI YANG DIFERMENTASI DENGAN Phanerochaete chrysosporium dan Neurospora crassa DALAM RANSUM TERHADAP KUALITAS TELUR PUYUH
    2021
    Co-Authors: Laila, Maulida Husni
    Abstract:

    Telur puyuh merupakan produk hasil peternakan yang memiliki gizi yang cukup seimbang, namun dewasa ini telur puyuh dihindari karena mengandung kolesterol yang cukup tinggi. Penelitian ini bertujuan untuk mengetahui berapa batasan dan bagaimana pengaruh penggunaan campuran limbah sawit dan dedak padi yang difermentasi dengan Phanerochaete chrysosporium dan Neurospora crassa (LSDF) dalam ransum terhadap kualitas telur puyuh. Penelitian ini menggunakan 200 ekor puyuh (Coturnix-coturnix japonica) umur 20 minggu dengan produksi telur 70%. Penelitian ini menggunakan metode eksperimen dengan Rancangan Acak Lengkap (RAL) dengan 4 perlakuan dan 5 ulangan. Perlakuan adalah penggunaan 0%, 8%, 16%, dan 24% LSDF dengan Phanerochaete chrysosporium dan Neurospora crassa. Peubah yang diamati yaitu kandungan kolesterol kuning telur (mg/100 g), kandungan lemak kasar (%) dan warna kuning telur. Hasil analisis keragaman menunjukan bahwa penggunaan campuran limbah sawit dan dedak padi yang difermentasi dengan Phanerochaete chrysosporium dan Neurospora crassa dalam ransum puyuh memberikan pengaruh berbeda sangat nyata (P

Masahiro Samejima - One of the best experts on this subject based on the ideXlab platform.

  • Crystal structure of a family 6 cellobiohydrolase from the basidiomycete Phanerochaete chrysosporium
    Acta Crystallographica Section F Structural Biology Communications, 2017
    Co-Authors: Mikako Tachioka, Kiyohiko Igarashi, Akihiko Nakamura, Takuya Ishida, Masahiro Samejima
    Abstract:

    Cellobiohydrolases belonging to glycoside hydrolase family 6 (CBH II, Cel6A) play key roles in the hydrolysis of crystalline cellulose. CBH II from the white-rot fungus Phanerochaete chrysosporium (PcCel6A) consists of a catalytic domain (CD) and a carbohydrate-binding module connected by a linker peptide, like other known fungal cellobiohydrolases. In the present study, the CD of PcCel6A was crystallized without ligands, and p-nitrophenyl β-d-cellotrioside (pNPG3) was soaked into the crystals. The determined structures of the ligand-free and pNPG3-soaked crystals revealed that binding of cellobiose at substrate subsites +1 and +2 induces a conformational change of the N-terminal and C-terminal loops, switching the tunnel-shaped active site from the open to the closed form.

  • transcriptional response of the cellobiose dehydrogenase gene to cello and xylooligosaccharides in the basidiomycete Phanerochaete chrysosporium
    Applied and Environmental Microbiology, 2012
    Co-Authors: Chiaki Hori, Kiyohiko Igarashi, Hitoshi Suzuki, Masahiro Samejima
    Abstract:

    Cellobiose dehydrogenase (CDH) gene transcripts were quantified by reverse transcription-PCR (RT-PCR) in cultures of Phanerochaete chrysosporium supplemented with various cello- and xylooligosaccharides in order to elucidate the mechanism of enhanced CDH production in xylan/cellulose culture. Cellotriose and cellotetraose induced cdh expression, while xylobiose and xylotriose induced expression of cellobiohydrolase genes, especially cel7C.

  • differential transcription of β glucosidase and cellobiose dehydrogenase genes in cellulose degradation by the basidiomycete Phanerochaete chrysosporium
    Fems Microbiology Letters, 2004
    Co-Authors: Makoto Yoshida, Kiyohiko Igarashi, Katsumi Aida, Rie Kawai, Masahiro Samejima
    Abstract:

    Transcriptional analysis of β-glucosidase gene (bgl) and cellobiose dehydrogenase gene (cdh) in relation to cellobiose metabolism in the basidiomycete Phanerochaete chrysosporium was performed using real-time quantitative RT-PCR. Addition of glucose to cellulose-degrading culture significantly decreased the number of both transcripts. In contrast, addition of cellobiose repressed only transcription of bgl but no effect for that of cdh. Moreover, to investigate induction of the two genes, the mycelia grown on glucose medium were transferred to medium containing glucose, cellobiose or no carbon source. In cellobiose medium, the number of bgl transcripts was slightly lower, whereas that of cdh transcripts was 2.3-fold higher than those in glucose medium. Consequently, cellobiose represses transcription of bgl, whereas it induces that of cdh.

  • production and characterization of recombinant Phanerochaete chrysosporium β glucosidase in the methylotrophic yeast pichia pastoris
    Bioscience Biotechnology and Biochemistry, 2003
    Co-Authors: Rie Kawai, Kiyohiko Igarashi, Makoto Yoshida, Tomomi Tani, Tsuyoshi Ohira, Hiromichi Nagasawa, Masahiro Samejima
    Abstract:

    The extracellular beta-glucosidase from the white-rot fungus Phanerochaete chrysosporium was expressed heterologously in the methylotrophic yeast Pichia pastoris. After 7 days' cultivation in an induction medium containing 1% (v/v) methanol, the expression level of the recombinant enzyme was 28,500 U/l, 38 times that of the wild-type enzyme. The specific activity of the crude recombinant enzyme for p-nitrophenyl-beta-D-glucoside was 52 U/mg, 37 times that of the wild-type enzyme; this difference made the purification of the enzyme simple. On a SDS-PAGE, the molecular mass of the recombinant enzyme was 133 kDa, and that of the wild-type enzyme was 116 kDa, but the difference had no effect on the hydrolysis of cellobiose or p-nitrophenyl-beta-D-glucoside. We concluded that the recombinant enzyme produced by Pichia pastoris retains the catalytic properties of the wild-type enzyme from Phanerochaete chrysosporium.

Sri, Des Utari - One of the best experts on this subject based on the ideXlab platform.

  • PENGARUH PENGGUNAAN CAMPURAN LIMBAH SAWIT DAN DEDAK FERMENTASI DENGAN Phanerochaete chrysosporium DAN Neurospora crassa DALAM RANSUM TERHADAP PERFORMA PUYUH PETELUR DAN INCOME OVER FEED COST
    2020
    Co-Authors: Sri, Des Utari
    Abstract:

    Penelitian ini bertujuan untuk mengetahui berapa batasan dan bagaimana pengaruh penggunaan campuran limbah sawit dan dedak yang difermentasi (LSF) dengan Phanerochaete chrysosporium dan Neurospora crassa dalam ransum terhadap performa puyuh petelur. Penelitian ini menggunakan 200 ekor puyuh (Coturnix-coturnix japonica) umur 20 minggu dengan produksi telur 70%. Penelitian ini menggunakan metode eksperimen dengan Rancangan Acak Lengkap (RAL) dengan 4 perlakuan (0%, 8%, 16%, dan 24% LSDF dengan Phanerochaete chrysosporium dan Neurospora crassa) dan 5 kali ulangan. Peubah yang diamati yaitu konsumsi ransum (gr/ekor/hari), produksi telur harian (%), berat telur (g/butir), massa telur (gr/ekor/hari), konversi ransum dan income over feed cost (Rp/kg). Hasil analisis keragaman menunjukkan bahwa penggunaan campuran limbah sawit dan dedak yang difermentasi dengan Phanerochaete chrysosporium dan Neurospora crassa dalam ransum memberikan pengaruh berbeda tidak nyata (P>0.05) terhadap konsumsi ransum, produksi telur harian, berat telur, massa telur, dan konversi ransum. Kesimpulan dari penelitian ini adalah campuran limbah sawit dan dedak yang difermentasi dengan Phanerochaete chrysosporium dan Neurospora crassa dapat digunakan sampai 24% dalam ransum puyuh petelur. Pada kondisi ini diperoleh konsumsi ransum 24,22 gr/ekor/hari, produksi telur 79,93%, berat telur 9,96 gr/butir, massa telur 7,97 gr/ekor/hari, konversi ransum 3,04 dan income over feed cost Rp. 8.510/kg Kata Kunci : Limbah sawit, Neurospora crassa Performa produksi puyuh, Phanerochaete chrysosporium

Dan Cullen - One of the best experts on this subject based on the ideXlab platform.

  • extracellular oxidative systems of the lignin degrading basidiomycete Phanerochaete chrysosporium
    Fungal Genetics and Biology, 2007
    Co-Authors: Phil Kersten, Dan Cullen
    Abstract:

    The US Department of Energy has assembled a high quality draft genome of Phanerochaete chrysosporium, a white rot Basidiomycete capable of completely degrading all major components of plant cell walls including cellulose, hemicellulose and lignin. Hundreds of sequences are predicted to encode extracellular enzymes including an impressive number of oxidative enzymes potentially involved in lignocellulose degradation. Herein, we summarize the number, organization, and expression of genes encoding peroxidases, copper radical oxidases, FAD-dependent oxidases, and multicopper oxidases. Possibly relevant to extracellular oxidative systems are genes involved in posttranslational processes and a large number of hypothetical proteins.