• Preventive Nutrition and Food Science
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• v.8 no.1
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• pp.36-39
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• 2003

Many defined cell culture media were formulated over 3() years ago and may be deficient in certain micronutrients whose essentiality has only subsequently been recognised. The objective of this study was to evaluate whether alpha-minimal essential medium (MEM) supplemented with 10% foetal bovine serum contained sufficient selenium for optimal activity of the selenium containing enzymes cytosolic glutathione peroxidase (cGPx) and phospholipid hydroperoxide glutathione peroxidase (PHGPx) in cultured Chinese hamster ovary (CHO) cells. Additionally, the effect of zinc deficiency and metallothionein (MT) overexpression on cGPx and PHGPx activity was studied. The addition of 100 nM of selenous acid to the culture medium increased cGPx expression by 10-fold and PHGPx by about 2-fold in both wild-type CHO-K1 cells and CHO-K1 cells overexpressing mouse MT-1. Zinc deficiency had no significant effect on enzyme activity, but cells overexpressing mouse MT-1 had higher levels of cGPx activity. Zinc deficiency decreased cell survival but overexpression of MT-1 was partially protective, probably because its presence in quantity favoured the uptake, sequestration and cellular retention of any remaining zinc. This study demonstrates that selenium in complete alpha-MEM is insufficient for optimal cGPx and PHGPx activity and may compromise the cellular response to oxidative stress.

• Journal of Nutrition and Health
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• v.25 no.2
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• pp.116-122
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• 1992

This study determined hepatic microsomal lipid peroxide values glucose 6-phosphatase NA-DPH-cytochrome P450 reductase and cytosolic glutathione S-transferase activites to examine the effects of methyl group deficiency on hepatic lipid peroxidation in rats treated with diethylni-trosamine(DEN) and 2-acetylamionfluorene(AAF) Weanling sprague Dawley male rats were fed the diet with methyl group supplemented or deficient. Two weeks after feeding rate were injected with a single of 200mg/kg body weight DEN intraperitoneally and after four weeks 0.02% AAF containing diets were fed for two weeks. Animals were sacrificed at 6th week. Microsomal lipid peroxide values were tended to increase in methyl group deficiency(MD). Especially in case of carcinogen tratments lipid peroxide values were increased significantly in MD. Microsomal glucose 6-phophatase activities were decreased by MD and carcinogens and in MD with carcinogen group (MD+C) the enzyme activites were the lowest Glucose 6-phosphatase activities were negatively correlated with lipid peroxidation. Microsomal NADPH-cytochrome P450 reductase activities were the highest in MD+C and correlated positively with lipid peroxidation. Cytosolic glutathione S-transferase activities were the highest in MD+C Methyl group deficiency induces lipid peroxidation especially in case of being exposed to carcinogens. Therefore the results suggest that lipid peroxidation may be one of the meachanisms of carcinogensis by methyl group deficiency.

• Journal of the Korean Society of Food Science and Nutrition
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• v.22 no.2
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• pp.116-126
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• 1993

The purposes of this study were to investigate the effect of seleniumc (Se) and vitamin E on activity of enzyme relevant to lipid peroxidation in alcohol administrated rats. Seventy two male rats of Sprague-Dawley strain weighing about 58~62g were divided into 12groups. The dietary Se levels were 0, 0.4 and 10mg and the dietary vitamin E levels were 0 and 150mg per kg diet, respectively. Alcohol-administrated groups received drinking water solution containing 10% of ethanol from the 3-weeks of experimental periods. The obtained experimental results are summarized as follow: The ${\gamma}$-GTP activity in plasma was higher in alcohol administrated groups and high selenium group (HSe) and low selenium group (LSe) than in control groups (CSe). The ${\gamma}$-GOT and GPT activities were higher in alcohol groups. The ${\gamma}$-GTP activity was significantly influenced by alcohol in LSe groups than in other groups. The glutathione peroxidase (GSH-Px) activity of plasma was significantly lower in LSe groups than HSe and CSe groups. The GSH-Px activity of microsomal and cytosolic fraction was slightly lower in alcohol groups and was about a half value lower in HSe and LSe groups than CSe groups. There was negative correlation between plasma Se level and GSH-Px activity of cytosolic fraction in HSe groups (r=- 0.662, p<0.001) and positive correlation in LSe groups (r=0.640, p<0.001). The GSH S-transferase activity in microsomal and cytosolic fraction was slightly higher in alcohol administrated but vitamin E nonadministrated groups, and significantly higher in LSe groups than in other groups. The catalase activity in mitochondria was lower in HSe than CSe groups, but rather higher in LSe groups. The superoxide dismutase (SOD) activity in cytosolic fraction of liver was not found any effect in all groups. The cytochrome P-450 was higher in alcohol groups, but significantly lower in HSe groups. In conclusion, the deficiency of Se and vitamin E develops the hyperoxidation of liver lipid through the increase of activity of enzyme related to the lipid peroxidation and alcohol administration appears to further increase of hyperoxidation of liver lipid.

• The Korean Journal of Pharmacology
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• v.21 no.1
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• pp.1-11
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• 1985

The effects of feeding indole, indole-3-carbinol and benzofuran (all at 5 mmole/kg body wt./day) on various hepatic microsomal and cytosolic enzyme activities involved in xenobiotic metabolism have been compared. Benzofuran was found to elevate the activities of many enzymes both in microsomes (e.g., aniline hydroxylase, 7-ethoxycoumarin O-deethylase, p-nitrophenol UDPGA-transferase and epoxide hydrolase) and in cytosol (e.g., glutathione reductase, glutathione S-transferase, NADH:quinone reductase and UDP-glucose dehydrogenase). The structures of indole and indole-3-carbinol are similar to benzofuran except for the substitution of nitrogen with oxygen atom within the furan ring. Results showed that the activities of UDPGA-transferase and NADH:quinone reductase were not elevated by these indole compounds. While the chemical structure of these two indole compounds are identical except for the presence of the carbinol (methanol) group in indole-3-carbinol, there were marked differences in the types and activities of microsomal enzymes that were enhanced. Among the microsomal enzyme activities determined, indole elevated only the NADPH:cytochrome c reductase, while indole-3-carbinol increased several mixed function oxidase and particularly the epoxide hydrolase activities. Based on the chemical structures of tested compounds and the observed results, possible explanations for the mechanisms involved in elevating epoxide hydrolase activity by benzofuran and indole-3-carbinol are discussed.

• Parasites, Hosts and Diseases
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• v.29 no.3
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• pp.259-266
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• 1991

In cytosolic (raction of adult Paragonimus westermani, superoxide dismutase activity was identified (4.3 units/mg of specific activity) using a xanthine-xanthine oxidase system. The enzyme was purified 150 fold in its activity using the ammonium sulfate precipitation, DEAE-Trisacryl M anion-exchange chromatography and Sephadex G-100 molecular sieve chromatography. The enzyme exhibited the enhanced activity at pH 10.0. The enzyme activity totally disappeared in 1.0mM cyanide while it remained 77.8% even in 10 mM azide. These findings indicated that the ensyme was Cu, Zn-SOD type. Molecular mass of the enzyme was estimated to be 34 kDa by gel filtration and 17 kDa on reducing SDS-polyacrylamide gel electrophoresis which indicated a dimer protein.

• BMB Reports
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• v.31 no.3
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• pp.227-232
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• 1998

The leukocyte NADPH oxidase of neutrophils is a membrane-bound enzyme that catalyzes the production of $O_2^-$ from oxygen using NADPH as an electron donor. Dormant in resting neutrophils, the enzyme acquires catalytic activity when the cells are exposed to appropriate stimuli. During activation, the cytosolic oxidase components $p47^{phox}$ and $p67^{phox}$ migrate to the plasma membrane, where they associate with cytochrome $b_{558}$, a membrane-bound flavohemoprotein, to assemble the active oxidase. The oxidase can be activated in a cell-free system; the activating agent usually employed is an anionic amphiphile such as sodium dodecyl sulfate (SDS). Because $p47^{phox}$ can translocate by itself during activation, the conformational change in $p47^{phox}$ may be responsible for the activation of NADPH oxidase. We show here that the treatment of $p47^{phox}$ with SDS leads to an increase in the reactivity of the sutbydryl group of cysteines toward N-ethylmaleimide, indicating that the conformational change occurs when $p47^{phox}$ is exposed to SDS. We propose that this change in conformation results in the appearance of a binding site through which $p47^{phox}$ interacts with cytochrome $b_{558}$during the activation process.

• Journal of Microbiology and Biotechnology
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• v.18 no.2
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• pp.295-298
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• 2008

Creatine kinase (CK; E.C. 2.7.3.2) is an important enzyme that catalyzes the reversible transfer of a phosphoryl group from ATP to creatine in energy homeostasis. The brain-type cytosolic isoform of creatine kinase (BB-CK), which is found mainly in the brain and retina, is a key enzyme in brain energy metabolism, because high-energy phosphates are transfered through the creatine kinase/phosphocreatine shuttle system. The recombinant human BB-CK protein was overexpressed as a soluble form in Escherichia coli and crystallized at $22^{\circ}C$ using PEG 4000 as a precipitant. Native X-ray diffraction data were collected to $2.2{\AA}$ resolution using synchrotron radiation. The crystals belonged to the tetragonal space group $P4_32_12$, with cell parameters of a=b=97.963, $c=164.312{\AA},\;and\;{\alpha}={\beta}={\gamma}=90^{\circ}$. The asymmetric unit contained two molecules of CK, giving a crystal volume per protein mass $(V_m)$ of $1.80{\AA}^3\;Da^{-1}$ and a solvent content of 31.6%.

• BMB Reports
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• v.31 no.1
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• pp.31-36
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• 1998

Phenylalanine ammonia-lyase (PAL; EC 4.3.1.5), the first enzyme in the phenylpropanoid biosynthesis, catalyzes the elimination reaction of ammonium ion from L-phenylalanine. PAL was purified from the cytosolic fraction of Chinese cabbage (Brassica campestris ssp. napus var. pekinensis) through ammonium sulfate fractionation, DEAE-cellulose chromatography, Sephadex G-200 chromatography, and Q-Sepharose chromatography. It consists of four identical subunits, the molecular mass of which was estimated to be about 38,000 daltons on SDS-PAGE. The optimal pH and temperature of the purified enzyme are 8~9 and $45^{\circ}C$, respectively. Its activity is greatly inhibited by $Zn^{2+}$ ion, and strongly activated by caffeic acid. The purified PAL has some different characteristics compared to those obtained with other PALs.

• Journal of Microbiology and Biotechnology
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• v.28 no.6
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• pp.909-916
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• 2018

Previously, a cytosolic trehalase (TreH) from the hyperthermophilic archaeon Sulfolobus acidocaldarius was reported; however, the gene responsible for the trehalase activity was not identified. Two genes, saci_1816 and saci_1250, that encode the glycoside hydrolase family 15 type glucoamylase-like proteins in S. acidocaldarius were targeted and expressed in Escherichia coli, and their abilities to hydrolyze trehalose were examined. Recombinant Saci_1816 hydrolyzed trehalose exclusively without any help from a cofactor. The mass spectrometric analysis of partially purified native TreH also confirmed that Saci_1816 was involved in proteins exhibiting trehalase activity. Optimal trehalose hydrolysis activity of the recombinant Saci_1816 was observed at pH 4.0 and $60^{\circ}C$. The pH dependence of the recombinant enzyme was similar to that of the native enzyme, but its optimal temperature was $20-25^{\circ}C$ lower, and its thermostability was also slightly reduced. From the biochemical and structural results, Saci_1816 was identified as a trehalase responsible for trehalose degradation in S. acidocaldarius. Identification of the treH gene confirms that the degradation of trehalose in Sulfolobus species occurs via the TreH pathway.

• Journal of the East Asian Society of Dietary Life
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• v.1 no.2
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• pp.185-190
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• 1991

Effects of water extracts of endocarps and seeds of omija(Schizandra chinensis Baillon)on alloxan and CCl4 induced liver injuries were determined by measuring the contents of metabolites and enzyme activities both in liver and serum in male Sprague-Dawley rats. The treatment with water extracts in parts of omija showed low levels of glucose, GOT, LDH and FFA of serum but high level of hepatic G-6-P DH in alloxan treated rats. CCl4 treatment with water extracts of parts of omija resulted in low levels of GOT, LDH, UN of serum and hepatic pyruvate but hepatic cytosolic protein has been increased.