• Preventive Nutrition and Food Science
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• v.5 no.3
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• pp.126-130
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• 2000

To enhance the antitumor activity of Chinese cabbage kimchi, four kinds of kimchi, which ere differently prepared in kinds and levels of sub-ingredients, were fermented at 15$^{\circ}C$ for 1 day and then at 5$^{\circ}C$ up to pH 4.3. The solid tumor formation, hepatic glutathione S-transferase activity and glutathione contents in the liver, and natural killer (NK) cell activity of spleen were determined from the sarcoma-180 cell injected Balb/c mice that were treated with methanol extracts of the kimchi samples. Kimchi IV, prepared with organically cultivated Chinese cabbage, red pepper powder, garlic, Chinese pepper powder mustard leaf and heat processed salt (Gueun salt), reduced the tumor formation by 39.3% compared to the sarcoma-180 cell treated group, resulting in the smallest tumor weight. Methanol extracts of the kimchi III and kimchi IV recovered the activities of hepatic glutathione S-transferase(GST) that was decreased by the transplantation of the sarcoma-180 cells to th mice. The injections of methanol extracts of kimchi II and kimchi IV increased glutathione contents in sarcoma-180 cells treated mice. The methanol extract of kimchi IV increased the natural killer (NK) cell activity of spleen lymphocytes a more effectively (p<0.05) than those the other kimchi samples. These results suggest that the anticancer activities of kimchi can be increased by changing the kinds and levels of sub-ingredients.

• Journal of Microbiology and Biotechnology
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• v.16 no.2
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• pp.318-324
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• 2006

In order to develop an oral vaccine to prevent H. pylori infection, we have expressed the hpaA gene of H. pylori K51 isolated from Korean patients, encoding 29-kDa HpaA that is known to be localized on the cell surface and flagella sheath, in a live delivery vector system, Lactococcus lactis. The hpaA gene, amplified by PCR using the genomic DNA of H. pylori K51, was cloned in the pGEX-2T vector, and the DNA sequence analysis revealed that the hpaA gene of H. pylori K51 had 99.7% and 94.8% identity with individual hpaA genes of the H. pylori 26695 strain (U.K) and the J99 strain (U.S.A). A polyclonal anti-HpaA antibody was raised in rats using GST-HpaA fusion protein as the antigen. The hpaA gene was inserted in an E. coli-L. lactis-shuttle vector (pMG36e) to express in L. lactis. Western blot analysis showed that the expression level of HpaA in the L. lactis transformant remained constant from the exponential phase to the stationary phase, without extracelluar secretion. These results indicate that the HpaA of H. pylori K51 was successfully expressed in L. lactis, and suggest that the recombinant L. lactis expressing HpaA may be applicable as an oral vaccine to induce a protective immune response against H. pylori.

• Journal of Microbiology and Biotechnology
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• v.18 no.7
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• pp.1235-1244
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• 2008

Indole-3-acetic acid (IAA) is produced commonly by plants and many bacteria, however, little is known about the genetic basis involving the key enzymes of IAA biosynthetic pathways from Bacillus spp. IAA intermediates from the Gram-positive spore-forming bacterium Paenibacillus polymyxa E681 were investigated, which showed the existence of only an indole-3-pyruvic acid (IPA) pathway for IAA biosynthesis from the bacterium. Four open reading frames (ORFs) encoding indole-3-pyruvate decarboxylase-like proteins and putative indole-3-pyruvate decarboxylase (IPDC), a key enzyme in the IPA synthetic pathway, were found on the genome sequence database of P. polymyxa and cloned in Escherichia coli DH5$\alpha$. One of the ORFs, PP2_01257, was assigned as probable indole-3-pyruvate decarboxylase. The ORF consisted of 1,743 nucleotides encoding 581 amino acids with a deduced molecular mass of 63,380 Da. Alignment studies of the deduced amino acid sequence of the ORF with known IPDC sequences revealed conservation of several amino acids in PP2_01257, essential for substrate and cofactor binding. Recombinant protein, gene product of the ORF PP2_01257 from P. polymyxa E681, was expressed in E. coli BL21 (DE3) as a glutathione S-transferase (GST)-fusion protein and purified to homogeneity using affinity chromatography. The molecular mass of the purified enzyme showed about 63 kDa, corresponding closely to the expected molecular mass of IPDC. The indole-3-pyruvate decarboxylase activity of the recombinant protein, detected by HPLC, using IPA substrate in the enzyme reaction confirmed the identity and functionality of the enzyme IPDC from the E681 strain.

• Environmental Analysis Health and Toxicology
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• v.23 no.4
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• pp.315-323
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• 2008

In this study, the backs with a hair cut of 6-week-old healthy ICR male mice were once exposed to a dose of $400\;mJ/cm^2$ UVB. An acute dermal inflammation was observed, and the certified 100% pure and natural lavender essential oil were applied to the UVB-exposed mice skin twice a day. It was observed that the mice exposed to UVB resulted in an acute inflammation, and when treated with lavender oil the degree of inflammation was much alleviated, and the inflamed skins of both the control and lavender oil-treatment groups were cured almost completely after 6 days of the UVB exposure. At 24 hours after UVB exposure, the epidermal keratinocytes in the control group showed a cell-membrane damage with the destruction of intercellular junctions, agglutination of tonofilaments within the cytoplasm and nucleus damage, while the lavender oil-treatment group had much less cell damage than the control group. While the control group showed a significant increase (p<0.05) in the activity of XO up to 144 hours, the lavender oil-treatment group did not show any significant increase except for 48 hours after the UVB exposure. Both the control and lavender essential oil-treatment groups had a significant decrease in the activities of CAT and SOD up to 96 hours. Particularly, the CAT activity was significantly lower(p<0.05) in the lavender oil-treatment group than the control group up to 48 hours, and higher than the control group at and after 96 hours. The GST activity was significantly decreased in both the control and lavender oil-treatment groups up to 96 hours after the UVB exposure except for the control group at 24 hours, and that of the lavender oil-treatment group was higher than the control group at and after 96 hours. Therefore, it is assumed that the application of the lavender oil to the ultraviolet-damaged mice skin can be effective in treatment for the damaged skin.

• Bulletin of the Korean Chemical Society
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• v.31 no.9
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• pp.2497-2502
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• 2010

Arg13 is a conserved active-site residue in all known Pi class glutathione S-transferases (GSTs) and in most Alpha class GSTs. To evaluate its contribution to substrate binding and catalysis of this residue, three mutants (R13A, R13K, and R13L) were expressed in Escherichia coli and purified by GSH affinity chromatography. The substitutions of Arg13 significantly affected GSH-conjugation activity, while scarcely affecting glutathione peroxidase or steroid isomerase activities. Mutation of Arg13 into Ala largely reduced the GSH-conjugation activity by approximately 85 - 95%, whereas substitutions by Lys and Leu barely affected activity. These results suggest that, in the GSH-conjugation activity of hGST P1-1, the contribution of Arg13 toward catalytic activity is highly dependent on substrate specificities and the size of the side chain at position 13. From the kinetic parameters, introduction of larger side chains at position 13 results in stronger affinity (Leu > Lys, Arg > Ala) towards GSH. The substitutions of Arg13 with alanine and leucine significantly affected $k_{cat}$, whereas substitution with Lys was similar to that of the wild type, indicating the significance of a positively charged residue at position 13. From the plots of log ($k_{cat}/{K_m}^{CDNB}$) against pH, the $pK_a$ values of the thiol group of GSH bound in R13A, R13K, and R13L were estimated to be 1.8, 1.4, and 1.8 pK units higher than the $pK_a$ value of the wild-type enzyme, demonstrating the contribution of the Arg13 guanidinium group to the electrostatic field in the active site. From these results, we suggest that contribution of Arg13 in substrate binding is highly dependent on the nature of the electrophilic substrates, while in the catalytic mechanism, it stabilizes the GSH thiolate through hydrogen bonding.

• Childhood Kidney Diseases
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• v.13 no.2
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• pp.170-175
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• 2009

Purpose : FSGS do not respond well to any kind of therapy and gradually progress to end-stage renal disease. This study was conducted to investigate the difference of protein expression between MCNS and FSGS as a preliminary study for understanding the pathophysiology of FSGS. Methods : Renal biopsy samples of MCNS and FSGS were obtained, which was diagnosed by one pathologist. They were solubilized with a conventional extraction buffer for protein extraction. The solution was applied on immobilized linear gradient strip gel (pH 4-7) using IPGphor system. Silver staining was carried out according to standard method. Protein identification was done by searching NCBI database using MASCOT Peptide Mass Fingerprint software. Results : The differences in protein expressions between MCNS and FSGS were shown by increased or decreased protein spots. Most prominently expressed spot among several spots in FSGS was isolated and analyzed, one of which was glutathione S-transferase (GST) P1-1, whereas it was not found in MCNS. So GSTP1-1 was considered as the one of the key biomarkers in pathogenesis of FSGS. Conclusion : This result would be helpful in diagnosing FSGS and researching FSGS. Further studies for glutathione S-transferase P1-1 might be necessary to elucidate the mechanisms regarding FSGS.

• The Korean Journal of Pesticide Science
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• v.14 no.3
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• pp.247-254
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• 2010

Two-spotted spider mite, Tetranychus urticae was collected from the rose greenhouse in Chilgok, Gyeongbuk Province in December 2000. This population has been selected for ten years with bifenazate (over 450 times), and increased 855.9 fold in resistance as compared with susceptible strain (S). Cross resistance of bifenazate resistant (BR) strain to eight miticides was investigated. The BR strain exhibited high and low cross resistance to acequinocyl (614.0 fold) and to chlorfenapyr (9.1 fold), respectively. Against fenazaquin (0.3 fold) and fenpyroximate (0.1 fold), however, showed the strain negatively correlated cross resistance. Each strain collected in Choeng-ju (CJ), Kang-jin (KJ), and Chung-ju (CUJ) showed 5.5-, 964.5-, and 21.8-fold resistance to bifenazate, respectively. The detoxifying enzymes of the BR strain showed 1.6-fold activity in cytochrome $P_{450}$-dependent monooxygenase ($P_{450}$) as compared with susceptible one. By comparing the mitochondrial cytochrome b (cytb) sequence, G126S point mutation was detected in the BR and KJ strains.

• Journal of the Korean Society of Food Science and Nutrition
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• v.34 no.9
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• pp.1330-1335
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• 2005

Red brownish p-pheylenediamine (PPD) has been widely used hair dye for women. The dye was known to cause systemic anaphylaxis, dermatitis and bladder cancer. But the effect of PPD toxicity with oxygen free radical has not been studied. This study investigated the degree of skin injury by PPD. PPD ($2.5\%$ PPD in $2\%\;NH_{4}OH$) was applied to the rat skin ($25 mg/16.5\;cm^2$) 3 or 5 times every other day. Histopathological findings demonstrated the proliferation of epithelial cells and the increased keratinization by PPD. The activities of glucose 6-phosphatase (G6Pase) was decreased and acid phosphatase (ACP) was increased in PPD-applied rat skin. Groups in which PPD was applied 5 times were more damaged than groups applied 3 times. To examine the relationship between tissue damage and oxygen free radicals, effect of PPD on xanthine oxidase (XO) activity was measured and XO activity was more significantly increased in the group treated with PPD 5 times than 3 times. However, reduced glutathione (GSH) content, and the activities of catalase (CAT), super-oxide dismutase (SOD) and glutathione S -transferase (GST) were more decreased in PPD-applied groups than in controls. Even though the activities of XOD was not changed in the group treated with PPD 3 times, the decreased activities of oxygen free radical system and the damaged skin tissue were observed. This result might be caused by the production of toxic PPD metabolites in rat skin. In conclusion, topical PPD application led to skin injury in a dose-dependent manner, probably due to the generation rate of oxygen free radical.

• Korean Journal of Food Science and Technology
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• v.37 no.4
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• pp.631-635
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• 2005

Effects of fermented milk, $Kupffer's^{\circledR}$, intake on hepatic antioxidative systems were investigated in rats fed ethanol (3 g/kg B.W.) for 2 weeks. Serum AST and ALT were $88.7{\pm}6.5\;and\;41.2{\pm}4.1IU/L$ in control group, $114.6{\pm}7.1\;and\;64.7{\pm}3.8IU/L$ in alcohol group, and $94.0{\pm}5.5\;and\;44.7{\pm}5.3IU/L$ in fermented milk (FM) group, respectively. Fermented milk intake decreased hepatic glutathione peroxidase and superoxide dismutase activities of FM group to level of control group (p<0.05). Glutathione S-transferase activity of fermented milk group increased by 122% compared to control group. These results suggest antioxidative activities of lactic acid bacteria and ingredients in $Kupffer's^{\circledR}$ improve antioxidative system in alcohol-treated rats.

• Proceedings of the Korean Society of Crop Science Conference
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• 2017.06a
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• pp.36-36
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• 2017

Out of twenty common protein amino acids, there are many kinds of non protein amino acids (NPAAs) that exist as secondary metabolites and exert ecological functions in plants. Mimosine (Mim), one of those NPAAs derived from L. leucocephala acts as an iron chelator and reversely block mammalian cell cycle at G1/S phases. Cysteine (Cys) is decisive for protein and glutathione that acts as an indispensable sulfur grantor for methionine and many other sulfur-containing secondary products. Cys biosynthesis includes consecutive two steps using two enzymes-serine acetyl transferase (SAT) and O-acetylserine (thiol)lyase (OASTL) and appeared in plant cytosol, chloroplast, and mitochondria. In the first step, the acetylation of the ${\beta}$-hydroxyl of L-serine by acetyl-CoA in the existence of SAT and finally, OASTL triggers ${\alpha}$, ${\beta}$-elimination of acetate from OAS and bind $H_2S$ to catalyze the synthesis of Cys. Mimosine synthase, one of the isozymes of the OASTLs, is able to synthesize Mim with 3-hydroxy-4-pyridone (3H4P) instead of $H_2S$ for Cys in the last step. Thus, the aim of this study was to clone and characterize the cytosolic (Cy) OASTL gene from L. leucocephala, express the recombinant OASTL in Escherichia coli, purify it, do enzyme kinetic analysis, perform docking dynamics simulation analysis between the receptor and the ligands and compare its performance between Cys and Mim synthesis. Cy-OASTL was obtained through both directional degenerate primers corresponding to conserved amino acid region among plant Cys synthase family and the purified protein was 34.3KDa. After cleaving the GST-tag, Cy-OASTL was observed to form mimosine with 3H4P and OAS. The optimum Cys and Mim reaction pH and temperature were 7.5 and $40^{\circ}C$, and 8.0 and $35^{\circ}C$ respectively. Michaelis constant (Km) values of OAS from Cys were higher than the OAS from Mim. Inter fragment interaction energy (IFIE) of substrate OAS-Cy-OASTL complex model showed that Lys, Thr81, Thr77 and Gln150 demonstrated higher attraction force for Cys but 3H4P-mimosine synthase-OAS intermediate complex showed that Gly230, Tyr227, Ala231, Gly228 and Gly232 might provide higher attraction energy for the Mim. It may be concluded that Cy-OASTL demonstrates a dual role in biosynthesis both Cys and Mim and extending the knowledge on the biochemical regulatory mechanism of mimosine and cysteine.