• Title/Summary/Keyword: glutathione S

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An Efficient System for the Expression and Purification of Yeast Geranylgeranyl Protein Transferase Type I

  • Kim, Hyun-Kyung;Kim, Young-Ah;Yang, Chul-Hak
    • BMB Reports
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    • v.31 no.1
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    • pp.77-82
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    • 1998
  • To purify the geranylgeranyl protein transferase type I (GGPT-I) efficiently, a gene expression system using the pGEX-4T-1 vector was constructed. The cal1 gene, encoding the ${\beta}$ subunit of GGPT-I, was subcloned into the pGEX-4T-1 vector and co-transformed into E. coli cells harboring the ram2 gene, the ${\alpha}$ subunit gene of GGPT-I. GGPT-I was highly expressed as a fusion protein with glutathione S-transferase (GST) in E. coli, purified to homogeneity by glutathione-agarose affinity chromatography, and the GST moiety was excised by thrombin treatment. The purified yeast GGPT-I showed a dose-dependent increase in the transferase activity, and its apparent $K_m$ value for an undecapeptide fused with GST (GST-PEP) was $0.66\;{\mu}M$ and the apparent value for geranylgeranyl pyrophosphate (GGPP) was $0.071\;{\mu}M$.

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Human Glutathione S-Transferase P1 Suppresses MEKK1-mediated Apoptosis by Regulating MEKK1 Kinase Activity in HEK293 Cells

  • Zhao, Xin;Fan, Yumei;Shen, Jiayin;Wu, Yifan;Yin, Zhimin
    • Molecules and Cells
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    • v.21 no.3
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    • pp.395-400
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    • 2006
  • Glutathione S-transferase P1 (GSTP1) plays an important role in detoxification and the metabolism of xenobiotics. Here we show that GSTP1 also regulates the MEKK1-MKK7 signaling pathway. Over-expression of GSTP1 in HEK293 cells inhibited both ${\Delta}MEKK1$- and etoposide-induced apoptosis, and inhibited procaspase-3 activation and PARP cleavage. MEKK1- induced apoptosis requires both its kinase activity and proteolytic cleavage. ${\Delta}MEKK1$ activity was inhibited by over-expression of GSTP1 in vivo and MEKK1 kinase activity was also inhibited by GSTP1 in vitro when assayed with bacterially-expressed MKK7(KM) protein as substrate. GSTP1 inhibition of etoposide-induced cell apoptosis was mainly due to its ability to suppress MEKK1 kinase activity. The glutathione-conjugating activity of GSTP1 was essential for the above effects. These findings provide insight into the mechanism by which GSTP1 protects cells from genotoxin-induced apoptosis.

Effects of Acori Graminei Rhizoma Aqua-acupunture Solution(AGRAS) on Induction of Cancer Chemopreventive Enzymes (석창포(石菖蒲) 약침액(藥鍼液)의 암(癌) 예방(豫防) 관련 효소 유도 효과)

  • Roh Dong-Il;Lim Jong-Kook
    • Korean Journal of Acupuncture
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    • v.19 no.2
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    • pp.51-56
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    • 2002
  • Induction of phase II enzymes such as quinone reductase (QR) and glutathione S-transferase (GST) is considered a major mechanism of protection against initiation of carcinogenesis. The present study was performed to evaluate the chemopreventive activity of Acori Graminei Rhizoma aqua-acupuncture solution (AGRAS) and Acori Graminei Rhizoma water-extracted solution (AGRWS) by measuring the induction of phase II enzymes. AGRAS and AGRWS are potent inducers of quinone reductase activity in murine hepatoma Hepa1c1c7 cells. The levels of GSH and GST was increased sightly with AGRAS and AGRWS. These results suggest that AGRAS and AGRWS may act as blocking agents against carcinogenesis by induction of phase II enzymes.

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Effects of γ-Irradiated Pork Diet on Cytochrome P-450 System, Microsome Glucose 6-Phosphatase Activity and Antioxidative Defense Systems in Rat Hepatocarcinogenesis

  • Kang, Il-Jun;Kim, Jung-Hee;Chung, Cha-Kwon
    • Preventive Nutrition and Food Science
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    • v.8 no.2
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    • pp.158-161
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    • 2003
  • This study investigated the effects of a ${\gamma}$-irradiated pork (0-30 kGy) diet on lipid peroxidation, cytochrome P-450 content, microsomal glucose 6-phosphatase (G-6-Pase) activity and antioxidative defense systems in diethylnitrosamine (DEN)-induced rat hepatocarcinogenesis. The body weight of rats fed irradiated diets did not change significantly. Liver weight was significantly increased by the administration of DEN, but not by irradiated diets at any dose level. There were no significant effects of gamma irradiation on the content of microsomal malondialdehyde (MDA), cytochrome P-450, or on the activity of G-6-Pase. However, with DEN treatment, cytochrome P-450 content was significantly increased while microsomal G-6-Pase activity was significantly decreased. The ${\gamma}$-irradiated diet supplement did not affect serum retinol or $\alpha$-tocopherol concentrations. However, it did cause a significant decrease in hepatic retinol at 30 kGy. With DEN treatment, hepatic retinol content was even more significantly (p<0.05) decreased compared to the non-irradiated control. The enzyme activities related to antioxidative defense systems, including glutathione peroxidase (GSH-Px), glutathione reductase (GSH-Rx) and glutathione S-transferase (GST) were not affected by gamma irradiation. Those results suggest that an irradiated pork diet up to 30 kGy may not cause a health hazard in experimental animals.

The Effect of Lycii fructus beer intake on serum lipid profiles and antioxidant activity in rats (구기자 맥주의 섭취가 흰쥐의 혈청 지질패턴 및 항산화효소 활성에 미치는 영향)

  • Chung, Hae-Kyung;Choi, Chang-Suk;Yang, Eun-Ju;Kang, Myung-Hwa
    • Journal of the Korean Society of Food Culture
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    • v.19 no.1
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    • pp.52-60
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    • 2004
  • This study was performed to investigate the effect of Lycii fructus beer on serum lipid profiles and antioxidant activity in rat Sprague-Dawley (SD) rats weighting about 190g were divided into the following 5 groups ; distillate water (Control), 5% ethanol in distillate water (Ethanol), commercial beer (CB), Lycii fructus beer (LFB) and 5% alcohol red wine diluted with distillate water (RW). Body weight, total food intake, FER and percent organ (liver, kidney) weight per body weight were not significantly changed by Lycii fructus beer drinking. After 6 weeks, serum total cholesterol, triglyceride and HDL cholesterol level were not significantly different. But, Lycii fructus beer intake tended to decrease serum triglyceride level and atherogenic index. Also, GOT and GPT levels were expressed lower than Ethanol group. There was not significantly different in hepatic glutatiione (GSH) content and glutathione-S-transferase (GST) activities among 5 groups. Lipid peroxidation in the hepatic was decreased by Lycii fructus beer intake. The results demonstrated that Lycii fructus beer was potential and effective antioxidant that can protect the decrease associated with alcohol.

Effect of Cnidii Rhizoma on Phase II Enzyme and Ornithine Decarboxylase Activities (천궁이 Phase II 효소 유도와 Ornithine Decarboxylase 활성에 미치는 영향)

  • Shon, Yun-Hee;Kim, Mee-Kyung;Cho, Hyun-Jung;Nam, Kyung-Soo
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.20 no.6
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    • pp.1572-1575
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    • 2006
  • Water extract from Cnidii Rhizoma (CRW) was tested for colon cancer chemopreventive activity by measuring the induction of phase II detoxification enzyme activity [quinone reductase (QR) and glutathione S-transferase (GST)] and glutathion (GSH) levels and ornithine decarboxylase (ODC) activity in cultured human colorectal adenocarcinoma HT-29 cells. CRW inhibited cell proliferation in cultured HT-29 cells. CRW induced QR activity in a dose-dependent manner in a concentration range of 0.1${\sim}$5.0 $mg/m{\ell}$. GST activity was also induced with the treatment of CRW in HT-29 cells. In addition GSH levels was increased with CRW. CRW inhibited ODC activity, a key enzyme of polyamine biosynthesis, which is enhanced in tumor promotion. These results suggest that CRW has colon cancer chemopreventive activity by increasing phase II enzyme activity and GSH levels and inhibiting ODC activity in vitro.

Neuroprotective Activity of Boesenbergia rotunda Against Glutamate Induced Oxidative Stress in HT22 Cells (글루타메이트에 의해 산화적 스트레스를 받은 HT22 세포에서 핑거루트의 신경세포 보호활성)

  • Kim, Eun Seo;Ma, Choong Je
    • Korean Journal of Pharmacognosy
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    • v.53 no.2
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    • pp.79-86
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    • 2022
  • Excessive glutamate causes oxidative stress in neuronal cells, which can cause degenerative neurological disorders. We tried to find medicinal plant showed neuroprotective activity by using glutamate-injured HT22 cell as a model system. In this study, we found that Boesenbergia rotunda methanol extract showed neuroprotective activity against glutamate induced neurotoxicity in mouse hippocampal HT22 cells. B. rotunda methanol extract suppressed the formation of reactive oxygen species and decreased intracellular Ca2+concentration. Also, B. rotunda made mitochondrial membrane potential maintain to normal levels. In addition, B. rotunda increased total glutathione amount and activated antioxidative enzyme such as glutathione reductase and glutathione peroxidase compared to glutamate-treated groups. These results suggested that B. rotunda decreased neuronal cell death damaged by high concentrations of glutamate treatment, via antioxidative mechanism and might be one of candidate of development of new drug to treat neurodegenerative disease such as Alzheimer's disease.

Molecular Characterization and Expression Analysis of a Glutathione S-Transferase cDNA from Abalone (Haliotis discus hannai) (북방전복 (Haliotis discus hannai)에서 분리한 Glutathione S-transferase 유전자의 분자생물학적 고찰 및 발현분석)

  • Moon, Ji Young;Park, Eun Hee;Kong, Hee Jeong;Kim, Dong-Gyun;Kim, Young-Ok;Kim, Woo-Jin;An, Cheul Min;Nam, Bo-Hye
    • The Korean Journal of Malacology
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    • v.30 no.4
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    • pp.399-408
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    • 2014
  • Glutathione S-transferases (GSTs) are a superfamily of detoxification enzymes that primarily catalyze the nucleophilic addition of reduced glutathione to both endogenous and exogenous electrophiles. In this study, we isolated and characterized a full-length of alpha class GST cDNA from the abalone (Haliotis discus hannai). The abalone GST cDNA encodes a 223-amino acid polypeptide with a calculated molecular mass of 25.8 kDa and isoelectric point of 5.69. Multiple alignments and phylogenetic analysis with the deduced abalone GST protein revealed that it belongs to the alpha class GSTs and showed strong homology with disk abalone (Haliotis discus discus) putative alpha class GST. Abalone GST mRNA was ubiquitously detected in all tested tissues. GST mRNA expression was comparatively high in the mantle, gill, liver, and digestive duct, however, lowest in the hemocytes. Expression level of abalone GST mRNA in the mantle, gill, liver, and digestive duct was 182.7-fold, 114.8-fold, 4675.8-fold, 406.1-fold higher than in the hemocytes, respectively. Expression level of abalone GST mRNA in the liver was peaked at 6 h post-infection with Vibrio parahemolyticus and decreased at 12 h post-infection. While the expression level of abalone GST mRNA in the hemocytes was drastically increased at 3 h post-infection with Vibrio parahemolyticus. These results suggest that abalone GST is conserved through evolution and may play roles similar to its mammalian counterparts.

Increase of Salt and Low Temperature Tolerance by Overexpressing Glutathione S-Transferase (GST) Gene (염분과 저온에 대한 내성증진을 위한 GST 유전자의 과발현)

  • Jun Chol Kim;Il Seop Kim;Won Hee Kang
    • Journal of Bio-Environment Control
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    • v.11 no.3
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    • pp.139-143
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    • 2002
  • Cotton Glutathione S-Transferase (GST: EC 2.5.1.18) was cloned and overexpressed in tobacco (Nicotiana tabacum) plants. Northern blot analysis confirmed the successful transformation of cotton gst gene in tobacco plant. Type I and Type ll transcript patterns were identified in transgenic tobacco plants and only Type I transcripts were discussed in this paper, The activity of GST in the type II transgenic plants was about 1.5-fold higher than those of the wild type and non-expresser by using 1-chloro-2,4-dinitrobenzene (CDNB) and reduced glutathione as the substrate. The expression of cotton GST in tobacco plants proved that Gh-5 could be translated into functional protein. Type II transgenic plants produced functional GST in the cells. The effects of cotton GST in the seedlings was evaluated by growing the control and transgenic seedlings at $15^{\circ}C$ in the growth chamber in the light. Overexpressors were grown well compared to the control plants (non-expressors). lo test far tolerance to salinity, seeds of Gh-5 overexpressors and the wild type Xanthi seedlings were grown at 0, 50, 100, 150, and 200 mM NaCl solution. Gh-5 transgenic seedlings showed higher growth rate over control seedlings on 50 and 100 mM NaCl solution. There was no difference in growth rate at 150 and 200mM NaCl concentration.