• Title/Summary/Keyword: Mg - dependent enzyme

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Observation of Asymmetry amongst Nucleotide Binding Sites of F1-ATPase of Escherichia coli by 31P NMR Spectroscopy

  • Jun, Nam-Kung;Sohn, Joon-Hyung;Yeh, Byung-Il;Choi, Jong-Whan;Kim, Hyun-Won
    • Bulletin of the Korean Chemical Society
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    • v.32 no.2
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    • pp.531-535
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    • 2011
  • It was regarded that the $^{31}P$ resonances of inherent nucleotides in $F_1$-ATPase (EF1), as large as 380KDa, could not be observed by $^{31}P$ NMR spectroscopy. However, our $^{31}P$ NMR spectroscopy could differentiate between different nucleotide binding sites on EF1 from Escherichia coli. When EF1 was prepared in the absence of $Mg^{2+}$, EF1 contained only ADP. Multiple $^{31}P$ resonances from $\beta$-phosphates of ADP bound to the EF1 were observed from the enzyme prepared without $Mg^{2+}$, suggesting asymmetry or flexibility amongst nucleotide binding sites. $^{31}P$ resonances from enzyme bound ATP could be observed only from EF1, when the enzyme was prepared in the presence of $Mg^{2+}$. This $Mg^{2+}$ dependent ATP binding was very tight that, once bound, nucleotide could not be removed even after removal of $Mg^{2+}$. $^{31}P$ NMR proved to be a valuable tool for investigating phosphorous related enzymes.

Antioxidant, Antibacterial and Anti-inflammatory Effects of Stachys sieboldii Extract (초석잠 추출물의 항산화, 항균 및 항염 활성)

  • Lee, Jeong Ho
    • Korean Journal of Plant Resources
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    • v.34 no.5
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    • pp.420-432
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    • 2021
  • This study was conducted to evaluate antioxidant, antibacterial, anti-inflammatory, and digestive enzyme activity in water extract (SAW) and 60% ethanol extract (SAE) from Stachys sieboldii. As the treatment concentration of each extract S. sieboldii extract increased, antibacterial and antioxidant activity increased. The total polyphenol and total flavonoid contents of SAW were 106.25 ± 0.94 mgGAE/g, 24.4 ± 0.24 mgQE/g and SAE were 124.61 ± 1.11 mgGAE/g, 45.2 ± 3.52 mgQE/g, respectively. The 400 ㎍/mL of SAW and SAE performed more than 53% protective effects against oxidative stress in HepG2 cell lines. All extracts were not showed cytotoxicity to RAW 264.7 cell line at 100 ㎍/mL. NO production was reduced to 44.3 ± 1.4% for SAW and 45.1 ± 1.0% for SAE at a concentration of 100 ㎍/mL. The production of inflammatory cytokines each TNF-α, IL-1β, and IL-6 was inhibited in a concentration-dependent. S. sieboldii extract did not showed Caco-2 cells cytotoxicity and inhibited NO production in concentration-dependent. As the concentration of the S. sieboldii extract increased of α-amylase and protease enzymes activity, which are digestive enzyme. As a result of the experiment, it is judged that it can be used as basic data for the development of health food using S. sieboldii.

Purification and Characterization of Protein Phosphatase 2C from Rat Liver

  • Oh, Joung-Sook;Hwang, In-Seong;Choi, Myung-Un
    • BMB Reports
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    • v.30 no.3
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    • pp.222-228
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    • 1997
  • Protein phosphatase 2C (PP2C) is one of the four major serine/threonine phosphatases which is dependent on $Mg^{2+}$ for its activity. PP2C was purified from rat liver cytosol and its characteristics were investigated. The substrate employed for routine assay was $[^{32}P]casein$ phosphorylated by PKA. The purification process involved DEAE chromatography, ammonium sulfate fractionation, phenyl sepharose chromatography, sephacryl 5-200 gel filtration, and histone agarose chromatography. The SDS-PAGE of PP2C showed one major single protein band at a position corresponding to a molecular mass of 43 kd and the purification fold was 637. The enzyme showed a pH optimum of 8 and $K_M$ value was $1.9\;{\mu}M$. However, when the substrate was changed to $[^{32}P]histone$, the pH optimum was shifted to 7 and $K_M$ value was $2.3\;{\mu}M.\;Mg^{2+}$ was essential to the enzyme activity and okadaic acid did not exert any inhibitory effect on the enzyme. To examine residue in the active site of PP2C effects of some protein-modifying reagents were tested.

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Alterations of Glutathione and Glutathione-Dependent Enzyme Activities by Monosodium-L-Glutamate in Rats with Carbon Tetrachloride-Induced Liver Damage (사염화탄소와 Monosodium-L-Glutamate 병용투여에 의한 간조직의 환원형글루타치온 함량 및 그의 관련효소활성의 변화)

  • 김형춘;이왕섭;전완주;김수희;주왕기
    • YAKHAK HOEJI
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    • v.35 no.5
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    • pp.384-388
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    • 1991
  • To explore the effect of monosodium-L-glutamate(MSG) on CCI$_{4}$-damaged liver in Wister male rat, 5% MSG solution as drink water were administered after S.C. injection of 0.1 mg/kg CC1$_{4}$ twice a week for 4 weeks. After last administration of MSG, heptic glutathione(GSH) dependent system was assayed. It showed that MSG increased significanly hepatic glutathione(GSH) and glutathione peroxidase(GSH$_{px}$), but decreased glutathione-S-transferase(GST) acivity in normal rats. MSG increased significantly the GSH$_{px}$ and GST activities in rats with CCI$_{4}$-induced liver damage. These results indicate that decrease of GSH dependent systems in CC1$_{4}$ liver injury might be partially elevated by coadministration of MSG.

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The Effect of Repeated Betaine Treatment on Hepatotoxicity and Cytochrome P-450 Dependent Drug Metabolizing Enzyme System (반복적인 Betaine 투여가 간독성 및 Cytochrome P-450 의존성 약물대사효소계 활성에 주는 영향)

  • Kim, Sang-Gyeom;Kim, Yeong-Cheol
    • YAKHAK HOEJI
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    • v.40 no.4
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    • pp.449-455
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    • 1996
  • Betaine is one of the major water-soluble components in Lycii Fructus. In the present study the effect of repeated betaine treatment on the hepatotoxicity and the cytochrome P-4 50-dependent enzyme system was examined in adult female rats. Administrations of betaine (100 or 1,000mg/kg/day, ip) to rats repeatedly for 4 or 9 days did not evoke hepatotoxic response as determined by increases in glutamic pyruvic transaminase(GPT) and glutamic oxaloacetic transaminase(GOT) activities measured 24 hours following the final dose of betaine. The activities of aminopyrine N-demethylase, p-nitroanisole O-demethylase and p-nitrophenol hydroxylase as well as the contents of cytochrome P-450 were determined in hepatic microsomes of rats treated with betaine(1,000mg/kg/day, ip) for 4 or 9 days. Repeated treatment of rats with betaine for a period of 4 days induced a marginal decrease in the contents of cytochrome P-450, but did not influence the activities of p-nitrophenol hydroxylase, p-nitroanisole O-demethylase, or aminopyrine N-demethylase. Extension of the betaine treatment to 9 consecutive days failed to alter the parameters for hepatic drug metabolizing activity determined in the present study. Since repeated large doses of betaine were demonstrated to be tolerated by rats without showing any toxicity or changes in drug metabolizing enzyme activities in the liver, this compound appears to be relatively safe to animals upon long-term ingestion.

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Scavenging Effects of Hydroxycinnamic acids on Paraquat Induced Hepatotoxicity (II) (Paraquat 유도 간독성에 대한 Hydroxycinnamic acid계 화합물의 독성 경감 효과 (II))

  • 최병기;오은정
    • Environmental Analysis Health and Toxicology
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    • v.14 no.3
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    • pp.87-93
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    • 1999
  • Antioxidative and scavenging effects were investigated by using two hyaroxycinnamic acids (caffeetannins). such as caffeic acid and chlorogenic acid, on oxidative stress and hepatotoxicity that induced by paraquat. The results are summerized as follows: 1. To assess radical scavenging ability, reduction concentration (IC$\sub$50/) of 1.1 diphenyl-2-dipicrylhydrazine (DPPH) were measured. IC$\sub$50/ values of caffeic acid and chlorogenic acid were 29.7 ${\pm}$0.6 ${\mu}$M and 26.0${\pm}$0.5 ${\mu}$M respectively. Their radical scavenging activities showed concentration-dependent manner. 2. In H$_2$O$_2$-induced hemolysis assay to rat blood, caffeic acid and chlorogenic acid led to different effects, whose hemolysis inhibition ratios at 100 ${\mu}$M were 45.2${\pm}$7.1% and 11.6${\pm}$3.1% respectively 3. In hypoxanthine-xanthine oxidase system producing superoxide anion, caffeic acid and chlorogenic acid showed different inhibitory activities of xanthine oxidase showing 36.8${\pm}$4.3% and 5.4${\pm}$2.3% respectively. 4. To microsomal NADPH dependent cytochrome p-450 reductase in rat liver, paraquat consumed NADPH at a dose-dependent manner from 0 to 1 ${\mu}$M paraquat concentration. Caffeic acid and chlorogenic acid blocked NADPH consumption rates at concentration-dependent manner and inhibition ratios at 100 ${\mu}$M were 67.6% and 59.2% respectively. 5. Administration (30mg/kg, iv) of paraquat to rats caused the marked elevation of glutamic oxaloacetic transaminase (GOT), glutamic pyruvic transaminase (GPT), lactate dehydrogenase (LDH), alkaline phosphatase (ALP) and lipid peroxides (LPO) in the serum and lipid peroxides in the microsome as compared to the control group. Serum GOT, GPT, LDH, ALP and LPO and liver microsomal LPO were reduced significantly by caffeic acid (50mg/kg), chlorogenic acid (25mg/kg) and silymarin (150 mg/kg) as compared to the paraquat group. From these results, caffeic acid and chlorogenic acid exerted their antioxidative agents by removing reactive oxygen substance (ROS) and scavenging effects by inhibiting ROS generating enzyme. As a general, two hydroxyeinnamic acids showed the useful compounds for scavenger and reducer on the paraquat induced hepatotoxicity.

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Increase in Hepatic DT-Diaphorase Activity by Chronic Administration of Panax ginseng Extract to Mice (생쥐에서의 인삼추출액의 장기간 투여에 의한 간장 DT-Diaphorase 활성의 증가)

  • Lee, Kang-Mee;Wie, Myung-Bok;Song, Dong-Keun;Kim, Yong-Sik;Kim, Yung-Hi
    • Journal of Ginseng Research
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    • v.17 no.2
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    • pp.123-126
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    • 1993
  • Effects of chronic administration of ginseng extracts (30 or 150 mg/kg/day for 52 days, p.o.) to mice on the activities of DT-diaphorase and glutathione S-transferase (GST) in the liver and the brain were studied. The DT-diaphorase activity in the liver was increased over 2-fold at the dose of both 30 and 150 mg/kg/day, while there was no change in the activity of the enzyme in the brain. The GST activity in the liver was increased in a dose-dependent fashion up to 142% of the control value at the dose of 150 mg/kg/day. while there was no change in the activity of the enzyme in the brain. The ginseng-induced increase in the activities of these hepatic phase II drug-metabolizing enzymes which are involved in the detoxification of carcinogens, is suggested to underlie, at least in part, the anticarcinogenic activity of Panax ginseng.

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Antioxidant and digestive enzyme inhibitory effects of Eisenia bicyclis extracted by different methods (추출방법에 따른 대황 추출물의 항산화 및 소화효소 저해 효과)

  • Kim, Jin-Hak;Park, La Young;Lee, Shin-Ho
    • Food Science and Preservation
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    • v.23 no.1
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    • pp.97-103
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    • 2016
  • The antioxidant and digestive enzyme inhibitory effects of Eisenia bicyclis extracted by various extraction methods (RE, reflux extraction; SE, ultrasonification extraction; AE, autoclave extraction; LE, low-temperature high-pressure extraction) were investigated. The extraction yield (55.21%) and the laminarin (39.03%), fucoidan (24.75%), total polyphenol (115.68 mg GAE/g) and flavonoid (36.67 mg RHE/g) contents of AE were higher than those in other methods. The DPPH radical (86.60%, 500 mg%), ABTS radical (58.56%, 25 mg%), nitrite (86.38%, 100 mg%) scavenging activities of the Eisenia bicyclis extracted by AE were higher than those of Eisenia bicyclis extracted by other methods. The ABTS radical and nitrite scavenging activities were above 98% in all tested Eisenia bicyclis extracts and these activities were dependent on its concentration. The inhibitory effects of AE against amylase (50 mg%) and ${\alpha}$-glucosidase (5 mg%) were 64.76% and 86.71%, respectively. The AE showed the best inhibitory effect of Eisenia bicyclis extracts (50 mg%) against trypsin (24.37%) and ${\alpha}$-chymotrypsin (49.05%), respectively. These results suggest that Eisenia bicyclis extracted by AE can be used as a bioactive and functional material in the food industry.

Effects of Tributyltin Chloride (TBTCI) on Reproductive Organs and Steroidogenic Enzymes

  • Ki, Ho-Youn;Lee, Su-Jung;Shin, Jae-Ho;Kang, Il-Hyun;Moon, Hyun-Ju;Kim, Tae-Sung;Hoon Bae;Dong, Mi-Sook;Yoon, Yong-Dal
    • Proceedings of the Korean Society of Developmental Biology Conference
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    • 2003.10a
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    • pp.67-67
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    • 2003
  • Tributyltin chloride (TBTCI) is an organotin compounds that have been widely used as antifouling agents and bioaccumulated in the food chain. TBTCI has been known to induce imposex in female gastropods. There are several reports that TBTCI increased testosterone level and inhibited the conversion of testosterone to estradiol by the aromatase cytochrome P450 enzyme. In this studies, we investigated the effects of TBTCI on steroidogenesis in testes, We dosed to 4-week-old Spragus-Dawleys (SD) male rats with TSTCI (0, 1, 5, 10, and 20mg/kg/day) daily by gavage for 14 days. TBTCI significantly decreased the weights of seminal vesicle, prostate, cowper's gland and LABC at 10 and 20mg/kg/day but significantly Increased the weights of liver at 10 and 20mg/kg/day and adrenals at 20mg/kg/day. mRNA levels of steroidogenic acute regulatory (StAR) and P450 aromatase were decreased and mRNA levels of cytochrome P450 17$\alpha$-hydroxylase/$C_{17-20}$ lyase (P450c17) were increased by TBTCI. TBTCI significantly increased serum testosterone level in dose-dependent manner. From above results, we found that TBTCI altered mRNA levels of enzymes related steroidogenesis, weights of organs and serum testosterone levels. This suggests that change of hormone levels may be due to alteration of mRNA levels of steroidogenic enzyme in testes, but further studies are necessary to investigate hormone levels in testis organ in order to find a relation of enzyme related to steroidogenesis with hormone levels. This work was supported by the Korea FDA Grant KFDA-03131-EDS-010.

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Scavenging Effects of Lonicera Japonica Extracts on Paraquat Induced Toxicity(IV) (Paraquat 유도독성에 대한 금은화 엑스의 효과(IV))

  • 최병기
    • Environmental Analysis Health and Toxicology
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    • v.15 no.1_2
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    • pp.7-12
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    • 2000
  • Scavenging effects on paraquat induced toxicity were investigated by using methanol (MeOH) and ethylacetate (EtoAC) extracts of Lonicera japonica. The results are summerized as follows: 1. To Fe(III)-ADP-NADPH induced microsomal lipid peroixdation, MeOH and EtoAC extracts showed antioxidative activiies and inhibition ratio at 100 $\mu\textrm{g}$/$m\ell$ 44.4% and 73.8% respectively 2. To microsomal NADPH dependent cytochrome p -450 reductase in rat liver, MeOH and EtoAC extracts inhibited the enzyme activiies and inhibition ratio were 26.3% and 44.8% respectively. 3. Administration (30 mg/kg, iv) of paraquat to rats caused the marked elevation of GOT, GPT, LDH, ALP in the serum and lipid peroxides in the microsome as compared to the control group. Serum GTP, LDH, ALP and liver microsomal LPO were reduced significantaly by administration of MeOH extract. (1,000 mg/kg), EtoAC extract (40 mg/kg) and Silymarin (150 mg/kg) as compared to the paraquat group. From the results, MeOH and EtoAc exuacts. of Lonicera japonica showed the useful scavenger and reducer on the paraquat induced hepatotoxicty.

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