• Title/Summary/Keyword: Activity system

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High Yield Bacterial Expression and Purification of Active Cytochrome P450 p-coumarate-3-hydroxylase (C3H), the Arabidopsis Membrane Protein (대장균 시스템을 이용한 Arabidopsis 막 단백질 cytochrome P450 p-coumarate-3hydroxylase (C3H) 활성형의 과발현 및 분리정제)

  • Yang, Hee-Jung;Kim, Wan-Yeon;Yun, Young-Ju;Yoon, Ji-Won;Kwon, Tae-Woo;Youn, Hye-Sook;Youn, Bu-Hyun
    • Journal of Life Science
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    • v.19 no.8
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    • pp.1039-1046
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    • 2009
  • The cytochrome P450s (P450s) metabolizing natural products are among the most versatile biological catalysts known in plants, but knowledge of the structural basis for their broad substrate specificity has been limited. The activity of p-coumarate 3-hydroxylase (C3H) is thought to be essential for the biosynthesis of lignin and many other phenylpropanoid pathway products in plants however, all attempts to express and purify the protein corresponding C3H gene have failed. As a result, no conditions suitable for the unambiguous assay of the enzyme are known. The detailed understanding of the mechanism and substrate-specificity of C3Hdemands a method for the production of active protein on the milligram scale. We have developed a bacterial expression and purification system for the plant C3H, which allows for the quick expression and purification of active wild-type C3H via introduction of combinational mutagenesis. The modified cytochrome P450 C3H ($C3H_{mod}$) could be purified in the absence of detergent using immobilized metal affinity chromatography and size exclusion chromatography following extraction from isolated membranes in a high salt buffer and catalytically activated. This method makes the use of isotopic labeling of C3H for NMRstudies and X-ray crystallography practical, and is also applicable to other plant cytochrome P450 proteins.

Anti-neuroinflammatory Effect of Teleogryllus emma Derived Teleogryllusine in LPS-stimulated BV-2 Microglia (BV-2 미세아교세포에서 왕귀뚜라미 유래 Teleogryllusine의 신경염증 억제 효과)

  • Seo, Minchul;Shin, Yong Pyo;Lee, Hwa Jeong;Baek, Minhee;Lee, Joon Ha;Kim, In-Woo;Hwang, Jae-Sam;Kim, Mi-Ae
    • Journal of Life Science
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    • v.30 no.11
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    • pp.999-1006
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    • 2020
  • The suppression of neuroinflammatory responses in microglial cells, well known as the main immune cells in the central nervous system (CNS), are considered a key target for improving the progression of neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, and Huntington's disease. Teleogryllus emma is widely consumed around the world for its broad-spectrum therapeutic effect. In a previous work, we performed transcriptome analysis on T. emma in order to obtain the diversity and activity of its antimicrobial peptides (AMPs). AMPs are found in a variety of species, from microorganisms to mammals. They have received much attention as candidates oftherapeutic drugs for the treatment of inflammation-associated diseases. In this study, we investigated the anti-neuroinflammatory effect of Teleogryllusine (VKWKRLNNNKVLQKIYFVKI-NH2) derived from T. emma on lipopolysaccharide (LPS) induced BV-2 microglia cells. Teleogryllusine significantly inhibited nitric oxide (NO) production without cytotoxicity, and reducing pro-inflammatory enzymes expression such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). In addition, Telegryllusine also inhibited the expression of pro-inflammatory cytokines such as interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) through down-regulation of the mitogen-activated protein kinases (MAPKs) and nuclear factor kappa B (NF-κB) signaling pathway. These results suggest that T. emma-derived Teleogryllusine could be a good source of functional substances that prevent neuroinflammation and neurodegenerative diseases.

Potentiation of Antitumor Effect of Radiotherapy by Recombinant Tumor Necrosis Factor-$\alpha$ (방사선의 항암작용에 대한 재조합 TNF-$\alpha$의 효과)

  • Seong Jinsil;Shin Hang Chul;Kim Gwi Eon;Suh Chang Ok
    • Radiation Oncology Journal
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    • v.16 no.3
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    • pp.225-231
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    • 1998
  • Purpose : To determine whether TNF-$\alpha$ increases the antitumor effect of radiotherapy in murine syngeneic tumor system. Materials and Methods : Syngeneic murine tumors of MCa-K or MCa-4 (mammary carcinoma), OCa-I (ovarian carcinoma), or HCa-I(hepatocarcinoma were grown in hind legs of C3Hf/HeJ mice. When tumors were grown to 6 mm in mean diameter mice were treated with TNF-$\alpha$, radiation, or combination of the both. Gamma-radiation was given as a single dose of 30 Gy for HCa-I and 15 Gy for other tumors using Cobalt-60 teletherapy unit. A novel TNF-$\alpha$ mutein developed in Korea, was intraperitoneally administered daily at a dose of 10 ug per mouse for 7 days. In combination of radiation and TNF-$\alpha$, the drug was started 1 hour after radiation. Tumor growth delay assay was used to measure the tumor response to the treatment. Results : Among 4 tested tumors, TNF-$\alpha$ alone showed significant antitumor activity in MCa-K and OCa-I tumors, which showed absolute growth delay (AGD) of 5.0 days and 6.5 days, respectively. In combination with radiation, TNF-$\alpha$ showed significant delay of AGD (41.1 days) in OCA-I compared to AGDs of TNF-$\alpha$ alone and radiation, i.e., 6.5 days and 26.9 days, respectively(p<0.05). Enhancement factor was 1.29 in OCa-I, which showed supraadditive effect. TNF-$\alpha$ did not show significant delay of AGDs in the remaining 3 tumors compared to AGDs of TNF-$\alpha$ alone and radiation. Conclusions: TNF-$\alpha$ alone showed antitumor effects in MCa-K and OCa-I. In combination with radiation, TNF-$\alpha$ acted in supraadditive way in OCa-I only. The results of this study imply that the combination of TNF-$\alpha$ and radiation has different therapeutic potential depending on tumor model and further study is advocated.

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Effects of Cellulose and Pectin on Diet-induced Thermogenesis in Young Women (한국인 젊은 여성에서 셀룰로오스 및 펙틴이 식이성 발열효과에 미치는 영향)

  • Oh, Seung-Ho;Park, Jeong-Jin;Choi, In-Seon;Ro, Hee-Kyong
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.36 no.2
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    • pp.194-200
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    • 2007
  • The purpose of this study was to observe the effects of fiber free control diet (CD), cellulose diet (CED) and pectin diet (PTD) on diet induced thermogenesis (DIT) in healthy Korean woman for 3 hours. The three test diets were as follows: CD (carbohydrate intake: 63.4% of energy, protein intake: 14.0% of energy and fat intake: 25.8% of energy), CED (carbohydrate intake: 62.5% of energy, protein intake: 14.0% of energy and fat intake: 26.1% of energy) and PTD (carbohydrate intake: 62.7% of energy, protein intake: 14.0% of energy and fat intake: 26.2 of energy). Groups were served 10 g of cellulose for CED and 10 g of pectin for PTD, respectively. DIT was measured at fasting state and at 30, 60, 90, 120, 150 and 180 min after consuming each diet. The mean age of all subjects was $22.3{\pm}1.9$ years. Body weight was $52.5{\pm}8.6$ kg and body mass index was $20.6{\pm}2.7kg/m^2$. Preprandial resting energy expenditure was $0.79{\pm}0.02kcal/min$ and postprandial DIT were $14.05{\pm}0.62%$ for CD, $9.33{\pm}0.62%$ for CED, $11.07{\pm}1.35%$ for PTD as a percentage of the energy load. DIT of CD was significantly higher than those of CED and PTD. There was no significant difference in postprandial change in body temperature after consuming each test diets and the sympathetic nervous system activity measured by heart rate was significantly higher in CD than CED and PTD (p<0.05). With this study, it can be concluded that CED and PTD have significantly lowered in DIT (p<0.05). We didn't show the correlation of the factors that relate in DIT; thus, further experiments on that matter should be followed.

Hepatic Oxygen Free Radical and Alcolhol Metabolizing Enzyme Activities in Rats Fed Diets Supplemented with Lycium chinense Ethanol Extract (흰쥐에 있어서 구기자 추출물 첨가식이가 간조직의 유해산소 및 알콜대사 효소활성에 미치는 영향)

  • 윤종국;김현희;채순님;오만진;이규희
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.30 no.4
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    • pp.668-672
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    • 2001
  • To investigate the oxygen free radical and alcohol metabolizing system in liver of rats fed diets with 30% ethanol extract of Lycium chinense (LCEE), Sprague-Dawley male rats weighing 225~235g have been fed a diet supplemented with 2% or 4% LCEE for a month. The rats fed LCEE supplemented diets gained less body weight compared with the control, and had no remarkable changes of liver function. In rats fed 2% LCEE supplemented diet, hepatic cytochrome P450 contents appeared to be increased, but catalase (204.88$\pm$20.06 $H_2O$$_2$nmoles/mg protein/min), and superoxide dismutase (13.18$\pm$0.74 Unit/mg protein) activities were significantly increased compared with control 120.28$\pm$26.99 $H_2O$$_2$nmoles/mg protein/min and 10.49$\pm$0.80 Units/mg protein). There was no difference in hepatic glutathione content, glutathione peroxidase, and glutathione-S-transferase ctivities between the rats fed LCEE suplemented diets and the control diet. On the other hand, hepatic alcohol dehydrogenase activity were not changed by LCEE feeding, but hepatic aldehyde dehydrogenase activities were significantly increased in rats fed both 2 and 4% LCEE diets(5.01$\pm$0.21 and 4.47$\pm$0.06 $\mu$moles NADPH/mg protein/min) compared with control (3.28$\pm$0.21 $\mu$moles NADPH/mg protein/min) and its Vmax value was 1.9 fold increased in rats fed 2% LCEE and 1.5 fold in those fed 4% LCEE compared with control. In conclusion, it is likely that rats receiving a diet supplemented with LCEE may have the oxygen free radicals and alcohol detoxication potential.

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Effects of Fatty Acids and Vitamin E Supplementation on Antioxidant Systems in the Liver and Serum of the Second Generation Rat (지방산 및 비타민 E 보충 식이가 제2세대 흰쥐 간조직과 혈청의 항산화체계에 미치는 영향)

  • 황혜진;박정화;엄영숙;정은정;김수연;이양자
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.31 no.2
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    • pp.257-262
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    • 2002
  • Effects of dietary fatty acids and vitamin E on antioxidant system were studied in rat liver and serum. Sources of dietary fat (10 wt%) were safflower oil (SO) poor in $\omega$3 fatty acid and mixed oil (MO) with computer-adjusted fatty acid ratios (AA/DHA=1.4, $\omega$6/$\omega$3=6.3, P/M/S=1.0/l.5/1) with (ME) and without (MO) vitamin E (500 mg/kg diet). Rats were fed the three kinds of diet from 3~4 wks prior to the conception. At the age of 3 and 9 wks of the second generation rat, antioxidant vitamins and glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) activities were measured in the liver and serum. The concentrations of $\beta$-carotene were lower in ME than in MO and SO in the liver at the age of 3 wks. It seemed that vitamin E has an inhibitory action on the uptake of $\beta$-carotene or acts as a preferred antioxidant to $\beta$-carotene. The concentrations of lycopene were lower in SO than in MO in the liver at the age of 3 wks. The concentrations of cryptoxanthin showed no significant changes within groups. The activities of GSH-Px tended to increase in ME compared to MO and the ratios of SOD/GSH-Px tended to decrease in ME compared to MO in the liver at the age of 3 weeks. The activities of antioxidant enzyme at the age of 3 weeks and 9 weeks were similar. This suggested that the activity level of antioxidant enzymes reached to the adult level at the age of 3 weeks which is the end point of lactation period.

Effect of Hormones and Short Chain Fatty Acids on CYP7A1 Gene Expression in HepG2 Cell (호르몬과 단쇄지방산이 HepG2 Cell 내에서 CYP7A1 발현에 미치는 효과)

  • Yang, Jeong-Lye;Lee, Hyun-Jung;Kim, Yang-Ha
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.34 no.5
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    • pp.573-580
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    • 2005
  • Cholesterol $7\alpha-hydroxylase$ (CYP7A1) is the rate-limiting enzyme in the conversion of cholesterol to bile acids and plays a central role in regulating cholesterol homeostasis. We previously showed that a fermentable $\beta-glucan$ ingestion decreased plasma cholesterol levels due to fecal bile acid excretion elevation involved inincrease of cholesterol $7\alpha-hydroxylase$ mRNA expression and activity. It is proposed that short chain fatty acids (SCFA) produced by cecal and colonic fermentation of soluble fiber are associated with cholesterol-lowering effect of fiber. In the present study, we investigated whether CYP7A1 expression is up-regulated by short chain fatty acids or by hormones in cultured human hepatoma (HepG2) cells. Confluent HepG2 cell were incubated with acetate, propionate, or butyrate at 1 mM concentration for 24 hrs. Acetate as well as propionate increased to 1.8-fold expression of CYP7A1 mRNA than the control. Butyrate also increased 1.5-fold expression of CYP7A1 mRNA. Our data show for the first time that SCFA increase expression of CYP7A1 mRNA. Adding insulin, dexamethasone and triiodothyronine $(1\;{\mu}M)$ to HepG2 cell increased the expression of CYP7A1 mRNA to $150\%,\;173\%,\;141\%$, respectively. These results suggest that SCFA produced by cecal fermentation stimulate enteric nervous system, in which secreted some neuropeptides may be responsible for change in cholesterol and bile acid metabolism. These findings suggest that SCFA are involved in lowering plasma cholesterol levels due to the up-regulation of CYP7A1 and bile acid synthesis.

Effect of Raw Broun Rice and Job식s Tear Supplemented Diet on Serum and Hepatic Lipid Concentrations, Antioxidative System, and Immune Function of Rats (현미 및 율무 함유 생식이 영양불균형이 유도된 흰쥐의 체내 지질농도, 항산화체계 및 면역기능에 미치는 영향)

  • 박진영;양미자;전혜승;이진희;배희경;박태선
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.32 no.2
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    • pp.197-206
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    • 2003
  • Physiological functions of raw grain diet composed of brown rice and Job's Tear (1 : 1) were evaluated in rats raised with nutritionally unbalanced diet including 1% cholesterol, high proportion of animal lipids (lard: soybean oil : 8 : 2), sub-optimal levels of vitamin and mineral mixture along with 0.5% ethanol in drinking water for 4 weeks. Control rats were fed the AIN-93G diet for 9 weeks, and nutritionally unbalanced rats were divided into 3 groups, and fed one of the following diets with 0.5% ethanol in drinking water for another 5 weeks: unbalanced control diet (UCD), raw grain diet (RGD) (UCD +20% brown rice and Job's Tear mixture, and cooked grain diet (CGD)(autoclaved RGD at 121$^{\circ}C$ for 3 hours). Feeding UCD for 5 weeks significantly lowered the food efficiency ratio (FER) of rats than the value for control animals, and dietary supplementation of brown rice and Job's Tear mixture to UCD significantly restored the FER. Serum total cholesterol concentration was significantly lowered in rats fed RGD (24% decrease) or CGD (16% decrease) compared to the value for rats fed UCD. Feeding RGD for 5 weeks significaly lowered the serum LDL+VLDL-cholesterol concentration (26% decrease), as well as the hepatic cholesterol level (16% decrease) than the values for UCD rats. Animals fed CGD (38% decrease) or RGU (59% decrease) showed significantly lower level of hepatic thiobarbituric acid reactive substances (TBARS) compared to the value for rats fed UCD (p<0.05), although hepatic activities of antioxidative enzymes were not influenced by dietary supplementation. Feeding RGD for 5 weeks significantly increased CD4$^{+}$ T-cell population along with CD4$^{+}$/CD8$^{+}$ ratio of mesenteric lymph nodes compared to those for UCD rats (p<0.05). In conclusion, dietary supplementation of brown rice and Job's Tear mixture as raw grains exhibited superior activity lowering blood and hepatic levels of cholesterol, and improving mesenteric lymph nodes immune function of rats to the cooked grain mixture of identical ingredients.

Ethanol Induced Leucocytic and Hepatic DNA Strand Breaks Are Prevented by Styela clava and Styela plicata Supplementation in Male SD Rats (알코올로 인한 흰쥐의 백혈구 및 간 DNA 손상에 미치는 미더덕과 오만둥이 분말의 보충섭취 효과)

  • Kim, Jung-Mi;Park, Hae-Ryoung;Lee, Seung-Cheol;Park, Eun-Ju
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.36 no.10
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    • pp.1271-1278
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    • 2007
  • In this study, the ability of Styela clava or Styela plicata to reduce ethanol-induced hepatotoxicity and hepatic and leucocytic DNA damages was evaluated. Twenty four male SD rats were given 25% ethanol containing water (ad lib, p.o.) and divided into 3 groups; ethanol treated control group (EtOH), ethano1+3% S. clava (EtOH+SC), and ethano1+3% S. plicata (EtOH+SP). After 6 weeks, the supplementation of S. clava reduced the plasma ALT, ALP and LDH activities significantly (p<0.05), while S. plicata induced significant decrease in the plasma LDH activity only. The comet assay was employed to quantify the alcohol-induced DNA damage in rat hepatocytes and leucocytes. A significant protective effect on hepatic and leucocytic DNA damages was observed in S. clava or S. plicata supplemented groups compared to the EtOH control group. The hepatic DNA damage was correlated positively with plasma ALP and LDH activities. These results demonstrated that S. clava or S. plicata supplementation protected alcohol-induced hepatic and leucocytic DNA damage.

A Study of Technical Development of Mariculture in the Coastal Water (천해양식어업발달과정에 관한 연구 - 기술개발활동을 중심으로 -)

  • Choi, Jeang-Yoon
    • The Journal of Fisheries Business Administration
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    • v.16 no.1
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    • pp.91-124
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    • 1985
  • Mariculture is contrasted with inland aqua-culturing fisheries. It is defind as the Industry of rearing Aquaorganism in limited coastal area relatively shallow in depth. Then, It's coming into being realization of Mariculture in it is long in history that Mariculture was realized in Korea. But it is from the early part of 1960s, that this industry has normally developed. Owing to 200 miles economy-zone problems of coastal countries, the development of deep sea fishing was limited, so the Korean Government has now appreciated the importance of cultured industries in the field of coastal fisheries. And the Korean Mariculture the output of which was only 18, 000 M/T in '60s attained 540, 000M/T in 1980s, has now occupied its relative importance in Korean Fisheries Industry. So the purpose of this report is to suggest the prospect of technical development of mariculture in the future of Korea, through the analysis of the various problems that affect upon the individual management '||'&'||' fishing ground utilization, along with the appreciation of "how to extend of those technical innovation" and "how the fishermen's technique level is extended at this stage. According to this study, the result is summarized as follows. First, Maricultural technique is classified into 8 sub-techniques as follows, as shown in fig. 1.Fig. 1. The Formation structure of mariculture technique Second, the change of technical method of mariculture in coastal area of Korea has made as 5 stages; 1) Scattering of culturing organism 2) Culturing by putting stone and installing bamboo 3) Culturing by installing rope and seeding 4) Culturing of putting objectives in cages 5) Culturing fish by feed Third, the maricultural fisheries of Korea has about 70 years long in history. It began from 1910s. But at that time there was no special technique in aquaculture and its technique was confined in searching out the object of species. The species was laver, oyster ect.Forth, although realization of mariculture in Korea has been long time, it is of late from 1960s that this has been industrial with normal development, and its technique of mariculture has mainly has developed from 1970s. Its result not only contributed to the high growth in Korean ecconomy along with the well balanced development between industires, but also it played a great role for the resolution of nation's food problem. Especially maricultural production has shown its sustained annual increase of 13.8% during the last 20 years. So the portion of mariculture among total fisheries stucture was extended from 4.1% in the early 1960s to 22.4% in 1980s.Fifth, it could be safely said that such development in maricultural field is resulted from the activity of aquacultural institutes such as Fisheries Reseach '||'&'||' Development production of major kinds such as Oyster, Sea-mustard, and Laver etc. As well as in the innovation of aquaculturing method with synthetic fiber utilization. FRDA has played important role in the efficient propargation of new aquacultural technique.Sixth, as for the change in aquaculture structure and its during period between 1970s and 1980s, the private management participation shown 25% increase from household number of 45, 173 to 56, 268 in total number. And in the respect of the management scale, of their management decreased, while it showed an increase in relative large scale management, the increase over 3 employees compared with other fisheries field between '70s and 80s. This must be an major trait to be recorded, Now the data above mentioned are shown as in table 1 and 2.Table 1. The maricultural fishing ground development situation in 982.Table 2. The mariculture management as seen in the employmnet size in high seasion.Owing to the technical innovation, of the mariculture in coastal area new income of fishermen increased and it also is true that the number of fishermen participating in its industrialization increased. But the problem being from now on is the self-discharge of the destruction fishing ground considered resulted from rapid expansion in aquaculture industry and the preventive system of sentility of fishing ground. sentility of fishing ground.

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