• Journal of Microbiology and Biotechnology
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• v.24 no.2
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• pp.245-253
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• 2014

A fibrinolytic enzyme was produced by an edible mushroom of Pleurotus ostreatus using submerged culture fermentation. The enzyme was purified from the culture supernatant by applying a combination of freeze-thaw treatment, ammonium sulfate precipitation, hydrophobic interaction, and gel filtration chromatographies. The enzyme was purified by a 147-fold, with a yield of 7.54%. The molecular masses of the enzyme an determined by gel filtration and SDS-PAGE were 13.6 and 18.2 kDa, respectively. The isoelectric point of the enzyme was 8.52. It hydrolyzed fibrinogen by cleaving the ${\alpha}$ and ${\beta}$ chains of fibrinogen followed by the ${\gamma}$ chains, and also activated plasminogen into plasmin. The enzyme was optimally active at $45^{\circ}C$ and pH 7.4. The enzyme activity was completely inhibited by EDTA, whereas protease inhibitors of TPCK, SBTI, PMSF, aprotinin and pepstatin showed no inhibition on its activity. The partial amino acid sequences of the enzyme as determined by Q-TOF2 were ATFVGCSATR, GGTLIHESSHFTR, and YTTWFGTFVTSR. These sequences showed a high degree of homology with those of metallo-endopeptidases from P. ostreatus and Armillaria mellea. The purified enzyme can also be applied as a natural agent for oral fibrinolytic therapy or prevention of thrombosis.

• Journal of Life Science
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• v.14 no.5
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• pp.815-820
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• 2004

A bacterum containing antitumor activity was isolated from traditional korean food, Jeotgal. Through the 16s rRNA sequence analysis, the bacterium was identitied as a strain of Bacillus subtilis SW-l. The best culture condition for antitumor activity of the bacterium is 3% of soluble starch and 1 % of yeast extract as corbon and nitrogen sources, respectively. Cytotoxicitic concentrations of the culture supernatant of B. subtilis SW-1 against cancer cell lines, A549 and SK-OV3 were 30 ul/ml and 40 ul/ml, respectively, as $IC_{50}$/ values. In DNA fragmentation assay, the culture supernatant showed the programmed cell death (apoptosis) to cause degrading the chromosomal DNA like ladder. Taken together, the culture supernatant of the B. subtilis SW-1 has some possibility to be used as an antitumor agent.

• Mycobiology
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• v.36 no.1
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• pp.13-18
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• 2008

Eight distinct bacteria were isolated form diseased mycelia of the edible mushroom, Pleurotus eryngii. 16S rDNA sequence analysis showed that the isolates belonged to a variety of bacterial genera including Bacillus (LBS5), Enterobacter (LBS1), Sphingomonas (LBS8 and LBS10), Staphylococcus (LBS3, LBS4 and LBS9) and Moraxella (LBS6). Among them, 4 bacterial isolates including LBS1, LBS4, LBS5, and LBS9 evidenced growth inhibitory activity on the mushroom mycelia. The inhibitory activity on the growth of the mushroom fruiting bodies was evaluated by the treatment of the bacterial culture broth or the heat-treated cell-free supernatant of the broth. The treatment of the culture broths or the cell-free supernatants of LBS4 or LBS9 completely inhibited the formation of the fruiting body, thereby suggesting that the inhibitory agent is a heat-stable compound. In the case of LBS5, only the bacterial cell-containing culture broth was capable of inhibiting the formation of the fruiting body, whereas the cell-free supernatant did not, which suggests that an inhibitory agent generated by LBS5 is a protein or a heat-labile chemical compound, potentially a fungal cell wall-degrading enzyme. The culture broth of LBS1 was not inhibitory. However, its cell-free supernatant was capable of inhibiting the formation of fruiting bodies. This indicates that LBS1 may produce an inhibitory heat-stable chemical compound which is readily degraded by its own secreted enzyme.

• Journal of Veterinary Clinics
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• v.40 no.3
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• pp.175-181
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• 2023

Fucoidan extracted from brown seaweed has a variety of biological activities. Neutrophil extracellular traps (NETs) formation is an immune response for the invasion of pathogens. Neutrophils release granule protein and chromatin that form extracellular fibers that bind microbes. These NETs degrade virulence factors and kill bacteria. The aim of this study was to investigate the effect of fucoidan on NET formation of porcine peripheral blood polymorphonuclear cells (PMNs). The NET formation was determined by fluorescence emission of propidium iodide (PI) in PMNs by a fluorescence microplate reader. The production of tumor necrosis factor (TNF)-α from peripheral blood mononuclear cells (PBMCs) was measured by ELISA method. Fucoidan itself did not show any direct effect on NET formation. However, NET formation of PMNs was increased by the culture supernatant from PBMCs treated with fucoidan. The NET formation of PMNs were also enhanced by treatment with recombinant porcine (rp) TNF-α. The ability of culture supernatant from PBMCs treated with fucoidan to increase the NET formation of PMNs was inhibited by addition of goat anti-rp TNF-α polyclonal antibody (pAb) (IgG) prior to the culture. The increase of NET formation by rp TNF-α was also inhibited by goat anti-rp TNF-α pAb (IgG). The level of TNF-α in culture supernatant from PBMCs was increased by treatment with fucoidan. These results suggest that fucoidan increases porcine NET formation, which is mediated by TNF-α produced from PBMCs.

• Journal of Microbiology and Biotechnology
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• v.8 no.1
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• pp.42-48
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• 1998

In order to maximize the secretory expression of human serum albumin (HSA) in the yeast Saccharomyces cerevisiae, a series of HSA expression vectors were constructed with a combination of different promoters, 5' untranslated regions (5'UTR), and secretion signal sequences. The expression vector composed of the galactose-inducible promoter GALl0, the natural 5'UTR, and the natural signal sequence of HSA directed the most efficient expression and secretion of HSA among the constructed vectors when introduced into several S. cerevisiae strains. Although the major form of HSA expressed and secreted in the yeast transformants was the mature form of 66 kDa, the truncated form of 45 kDa was also detected both in the cell extract and in the culture supernatant. The level of the intact HSA protein in the culture supernatant reached up to 30 mg/l at 24 h of cultivation in a shake-flask culture but began to decrease afterwards, indicating that the secreted HSA protein was unstable in a prolonged culture of yeast.

• Journal of Veterinary Clinics
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• v.16 no.1
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• pp.37-41
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• 1999

The immunostimulating effects of egg white derivatives (EWD) on the phagocytic response of feline peripheral blood polymorphonuclear cells (PMN) as well as mono- nuclear cells (MNC) were examined. The phagocytic activity was analyzed by a flow cytometry system. The EWD showed directly an enhanced effect on the phagocytic response of MNC but not PMN. The phagocytic activity of MNC was enhanced by culture supernatant from MNC and PMN treated with EWD, respectively. Similarly, the phagocytic activity of PMN was enhanced by culture supernatant from MNC but not PMN treated with EWD. It was, therefore, indicated that the enhanced phagocytic activity of feline PMN could be mainly mediated by humoral factor(s) released from MNC treated with EWD. These results suggested that EWD could enhance the phagocytosis of feline peripheral blood phagocytes.

• Journal of Life Science
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• v.25 no.7
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• pp.810-818
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• 2015

We analyzed whether lactic acid bacteria could control the expression of IL-4 and IL-13 in activated mast cells and whether these bacteria could inhibit the activity of transcription factors such as GATA-1, GATA-2, NF-AT1, NF-AT2, and NF-κB p65. We previously described a technique for identification of lactic acid bacteria with anti-atopy functionality by confirming increased expression of CD4+/CD25+/foxp3+ in T cells. We also confirmed that a double-culture method increased the antibacterial activity of these lactic acid bacteria against Staphylococcus aureus (S. aureus). In the present study, we characterized the effect of lactic acid bacteria cultured by this double-culture method on inhibition of allergic inflammatory reactions of RBL-2H3 mast cells, a cellular model of atopic dermatitis. The strongest anti-allergic effects of the lactic acid bacteria were seen in the following order: Lactococcus lactis broth cultured with medium containing Lactobacillus plantarum culture supernatant > Lc. lactis > Lc. lactis broth cultured with medium containing Lb. plantarum culture supernatant > Lb. plantarum. Thus, Lc. lactis cultured in medium containing Lb. plantarum culture supernatant had the strongest inhibitory effect on the differentiation of mast cells during allergic reactions, which may be mediated through the selective regulation of expression of relevant genes.

• Korean Journal of Microbiology
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• v.41 no.1
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• pp.74-80
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• 2005

For the reseach of the natural marine antioxidant, an antioxidant-producing bacterium was isolated from seawater in Jeju costal area. The isolated strain SC2-1 was Gram-positive, catalase positive, oxidase negative, motile and small rods. The strain utilized sucrose, dextrose, fructose, mannitol and maltose as a sole carbon and energy source and NaCl required for growth. The radical scavenging activity of the culture supernatant was detemined by DPPH method. This bacterium was identified based on morphological, biochemical characteristics and 16S rDNA sequence analysis, and then was named Exiguobacterium sp. SC2-1. The optimum conditions of culture for production antioxidnat were $25^{\circ}C$, pH 6-8 and $4\%$ NaCl. The stain showed the highest activity and growth cultured in medium which added $1\%$ maltose. Hydroxyl radical activity of the supernatant of Exiguobacterium sp. SC2-1 was $73\%$. The SOD activity of the culture supernatant was estimated about $35\%$.

• Journal of Dairy Science and Biotechnology
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• v.26 no.1
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• pp.5-10
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• 2008

This study was conducted to isolate protein components from culture supernatant of Lactobacillus casei. and measure anti-cancer activity. The protein components were isolated A and B on Ultrafiltration membrane(3, 10, 30, 100 KDa). And the protein components A and B were isolated fractions(number $3{\sim}9$) on FPLC. Experimental studies were progressed through the cell cytotoxicity and anti-cancer activities. Cell cytotoxicity test using human kidney normal cell(293) showed cytotoxicity of below 20% by the protein components A and B($100{\mu}g/mL$). The anti-cancer activity was increased up to 70% by the protein components A and B($100{\mu}g/mL$) in AGS(stomach cancer), A549(lung cancer), MCF-7 (breast cancer), SK-OV-3(ovary cancer) and LoVo(colon cancer). Cell cytotoxicity test was showed cytotoxicity of about 50% by the fractions(number 3, 8, 9) isolated FPLC. The others have not the cytotoxicity about the human normal cell. The anti-cancer activity was increased up to 70% by the fraction number 7 in cancer cell line. Therefore the components isolated from culture supernatant of Lactobacillus casei were showed anti-cancer activity.

• Journal of Life Science
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• v.19 no.5
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• pp.676-679
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• 2009

In order to produce a new antibiotic material against Jurkat T cells using horizontal gene transfer among microbes, co-cultures between soil bacteria AY2000 and the multiple antibiotic producer S. griseus was carried out. It showed that the highest active substance against Jurkat T cells was produced at 48 hr of co-culture time with MTT assay. Moreover, a morphological change of nuclear of Jurkat T cells treated with co-cultured substance was observed in DAPI staining. This result suggests that a new material was produced with co-culture supernatant, and that co-culture between microboes can develop new antibiotic materials.