• Korean Journal of Food Science and Technology
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• v.30 no.3
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• pp.709-716
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• 1998

Three kinds of anti-complementary system and macrophage activating polysaccharides, AB-20-Ia, AB-20-IIa-2a and AB-20-IVa-2 were isolated from the fruit body of Agaricus bisporus and their structures were characterized. The proteoglycan, AB-20-IVa-2 showing the most potent anti-complementary and macrophage activity was composed of glucose, galactose, mannose, xylose, fucose and arabinose in a molar ratio of 3.48:1.83:1.00:0.79:0.74:0.11 and its main component amino acids were phenylalanine (34.72%) and valine (27.84%). The neutral polysaccharides, AB-20-Ia and AB-20-IIa-2a showing lower activity than AB-20-IVa-2, consisted of xylose, glucose, mannose, fucose and arabinose in molar ratios of <0.05:<0.05:2.07:1.00:2.72 and 2.16:1.58:1.00:0.20:0.14, respectively. The molecular weights of AB-20-Ia, AB-20-IIa-2a and AB-20-IVa-2 were 840,000, 750,000 and 650,000 respectively. In the $^1H-\;and\;^{13}C-NMR$ spectra of AB-20-Ia and AB-20-IIa-2a, AB-20-Ia showed only ${\beta}-configuration\;(^1H:\;4.8\;ppm,\;^{13}C:\;107.0\;ppm)$ in the anomerization of the glycosidic linkages, while AB-20-IIa-2a had both ${\alpha}-anomer\;(^1H:\;5.4\;ppm,\;^{13}C:\;102.0\;ppm)\;and\;{\beta}-anomer$. Especially, AB-20-Ia and AB-20-IIa-2a showed acetyl signals $(^1H:\;2.5\;ppm,\;^{13}C:\;21.0\;ppm)$. In the methylation analysis of the three polysaccharides, high proportion of 1,6-linked glucofuranosyl residues were detected in AB-20-Ia, whereas 1,6-linked glucopyranosyl residues and branches linked at position 4 of those mainly contained in AB-20-IIa-2a. AB-20-IVa-2 consisted mainly of 1,2-linked xylofuranosyl residues and 1,6-linked glucopyranosyl residues and branches linked at position 3 of those.

• Journal of the Korean Society of Food Science and Nutrition
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• v.44 no.5
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• pp.664-672
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• 2015

In order to develop new immuno-stimulating ingredients from mature leaves of green tea, crude polysaccharides were isolated from pectinase digests of tea leaves (green tea enzyme digestion, GTE-0), after which their immuno-stimulating activities and chemical properties were examined. GTE-0 mainly contained neutral sugars (54.9%) such as glucose (14.2%), arabinose (12.2%), rhamnose (11.1%), and galacturonic acid (45.1%), which are characteristic of pectic polysaccharides. The anti-complementary activity of GTE-0 was similar to that of polysaccharide K (used as positive control). Number of morphologically activated macrophages was significantly increased in the GTE-0-treated group. GTE-0 significantly augmented $H_2O_2$ and reactive oxygen species production by murine peritoneal macrophage cells in a dose-dependent manner, whereas production of nitric oxide showed the highest activity at a dose of $100{\mu}g/mL$ among all tested concentrations. Murine peritoneal macrophages stimulated with GTE-0 showed enhanced production of various cytokines such as interleukin (IL)-6, IL-12, and tumor necrosis factors-${\alpha}$ in a dose-dependent manner. Further, GTE-0 induced higher phagocytic activity in a dose-dependent manner. In ex vivo assay for cytolytic activity of murine peritoneal macrophages, GTE-0-treated group showed significantly higher activity compared to the untreated group at an effector-to-target cell ratio of 20. The above results lead us to conclude that polysaccharides from leaves of green tea have a potent immuno-stimulating effect on murine peritoneal macrophage cells.

• Journal of Life Science
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• v.15 no.3 s.70
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• pp.415-422
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• 2005

library of the mutants was prepared by transposon mutagenesis of the Mycobacterium bovis BCG. We screened this library for the resistance to an anti-tuberculosis antibiotic, PA-824. Most of the mutants resistant to the PA-824 were not able to synthesize the coenzyme $F_{420}$ which is normally produced by the wild type M. bovis BCG strains. HPLC analysis of the cellular extract showed that one of those mutants which lost the ability to synthesize $F_{420}$ still produced F0. The insertion site of the transposon in this mutant was determined by an inverse PCR and the transposon was found to be inserted in the Rv2435c open reading frame (ORF). Rv2435c ORF is predicted to encode an 80.3 kDa protein. Rv2435c protein appears to be bound to the cytoplasmic membrane, its N-terminal present in the periplasm and C-terminal in the cytoplasm. The C-terminal portion of this protein is highly homologous with the adenylyl cyclases of both prokaryotes and eukaryotes. There are 15 ORFs which have homology with the class III AC proteins in the genome of the M. tuberculosis and M. bovis. Two of those, Rv1625c and Rv2435c, are highly homologous with the mammalian ACs. We cloned the cytoplasmic domain of the Rv2435c ORF and expressed it with six histidine residues attached on its C-terminal in Escherichia coli to find out if this protein is a genuine AC. Production of that protein in E. coli was proved by purifying the histidine-tagged protein by using the Ni-NTA resin. This protein, however, failed to complement the cya mutation in E. coli, indicating that this protein lacks the AC activity. All of the further attempts to convert this protein to a functional AC by a mutagenesis with UV or hydroxylamine, or construction of several different fusion proteins with Rv1625c failed. It is, therefore, possible that Rv2435c protein might affect the conversion of F0 to $F_{420}$ not by synthesizing cAMP but by some other way.