- Purification and Characterization of Manganese Peroxidase of the White-Rot Fungus Irpex lacteus
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Kwang-Soo Shin , Young Hwan Kim , Jong-Soon Lim
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J. Microbiol. 2005;43(6):503-509.
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DOI: https://doi.org/2298 [pii]
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The production of manganese peroxidase (MnP) by Irpex lacteus, purified to electrophoretic homogeneity by acetone precipitation, HiPrep Q and HiPrep Sephacryl S-200 chromatography, was shown to correlate with the decolorization of textile industry wastewater. The MnP was purified 11.0-fold, with an overall yield of 24.3%. The molecular mass of the native enzyme, as determined by gel filtration chromatography, was about 53 kDa. The enzyme was shown to have a molecular mass of 53.2 and 38.3 kDa on SDS-PAGE and MALDI-TOF mass spectrometry, respectively, and an isoelectric point of about 3.7. The enzyme was optimally active at pH 6.0 and between 30 and 40oC. The enzyme efficiently catalyzed the decolorization of various artificial dyes and oxidized Mn (II) to Mn (III) in the presence of H2O2. The absorption spectrum of the enzyme exhibited maxima at 407, 500, and 640 nm. The amino acid sequence of the three tryptic peptides was analyzed by ESI Q-TOF MS/MS spectrometry, and showed low similarity to those of the extracellular peroxidases of other white-rot basidiomycetes.
- Purification and Characterization of Extracellular Temperature-Stable Serine Protease from Aeromonas hydrophila
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Soo-Jin Cho , Jong-Ho Park , Seong Joo Park , Jong-Soon Lim , Eung Ho Kim , Yeon-Jae Cho , Kwang-Soo Shin
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J. Microbiol. 2003;41(3):207-211.
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Extracellular protease, from Aeromonas hydrophila Ni 39, was purified 16.7-fold to electrophoretic homogeneity with an overall yield of 19.9%, through a purification procedure of acetone precipitation, and Q Sepharose and Sephacryl S-200 chromatographies. The isoelectric point of the enzyme was 6.0 and the molecular mass, as determined by Sephacryl S-200 HR chromatography, was found to be about 102 kDa. SDS/PAGE revealed that the enzyme consisted of two subunits, with molecular masses of 65.9 kDa. Under standard assay conditions, the apparent K_m value of the enzyme toward casein was 0.32 mg/ml. About 90% of the proteolytic activity remained after heating at 60 ℃ for 30 min. The highest rate of azocasein hydrolysis for the enzyme was reached at 60℃, and the optimum pH of the enzyme was 9.0. The enzyme was inhibited by the serine protease inhibitor, phenylmethylsulfonyl fluoride (PMSF), by about 87.9%, but not by E64, EDTA, pepstatin or 1,10-phenanthroline. The enzyme activity was inhibited slightly by Ca_2^+, Mg_2^+ and Zn_2^+ ions.
- Cytokine-Inducing and T Cell Mitogenic Effects of Cordyceps hepialidicola
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Jong-Soon Lim , Seung-Hyung Kim , Jeong-Youl Choi , Jin-Seo Park , Seong Joo Park , Kwang-Soo Shin
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J. Microbiol. 2001;39(3):181-185.
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The morphological characteristics of newly isolated Cordyceps hepialidicola were characterized, and the phylogenetic relationships with other Cordyceps species were investigated using a sequence analysis of the internal transcribed spacer (ITS). The PCR product of 592 bp showed a homology of 92 and 91% with C. militaris and C. nutans, respectively. In an in vitro model using mouse peripheral blood mononuclear cells (PBMC), a methanol extract of C. hepialidicola induced multiple cytokines, including IFN-[gamma], IL-4, and IL-18. The extract also enhanced the percentages of the CD4^+ and CD8^+ T cells in the healthy murine PBMCs to 56.1% and 13.0%, respectively. The percentages of CD4^+ and CD8^+ in the untreated controls were 28.4 and 7.3%, and concanavalin A-treated positive controls were 62.4 and 18.3%, respectively.
- Anti-tumor Activity of the Fruitbody Extract of Basidiomycete, Phellinus linteus
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Jong-Soon Lim , Seung-Hyung Kim , Jin-Seo Park , Jeong-Youl Choi , Seong Joo Park , Kwang-Soo Shin
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J. Microbiol. 2001;39(2):121-125.
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Methanol extract prepared from the fruitbody of Phellinus linteus (EPL) showed anti-tumor and immuno-stimulating activities. The invasion activity of B16-F10 melanoma cells through a reconstituted basement membrane to the collagen-coated lower surface of the filters was inhibited about 67% by EPL (100 ug/ml). Also, EPL inhibited the expression of the mRNA for MMP-2 and MMP-9. In vivo treatment of C57BL/6 mice (150 mg/kg) with EPL for 14 days, the pulmonary colonization was found to be inhibited about 75%. Using reverse transcriptionpolymerase chain reaction (RT-PCR) analysis, we found that cytokine IL-12 and INF-[gamma] genes were induced by EPL. Furthermore, EPL stimulated the proliferation of CD4^+ (33.5%) and CD8^+ (17.7%) in mouse splenocytes.
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