• 통합자연과학논문집
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• 제8권4호
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• pp.267-272
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• 2015

Galactosemia is a potentially lethal disorder caused by the deficiency of the enzyme galactose-1-phosphate uridyltransferase (GALT) within the Leloir pathway. Galactokinase (GALK) is the enzyme in Leloir pathway which converts ${\alpha}$-D galactose to galactose 1-phosphate. The elevated level of galactose-1-phosphate, the product of GALK plays a major role in Galactosemia. Therefore the inhibition of GALK is a novel therapy for this disorder. Hence in the present study, we performed molecular docking of twenty inhibitors with different activity against galactokinase into the active site of galactokinase enzyme. The binding mode of these inhibitors was obtained using Surflex dock program interfaced in Sybyl-X2.0. The residues such as SER141, TYR109, ARG105, ARG228, TYR106, GLY346, GLY136, ASP86, ASP186 and SER142 found to interact with inhibitors.

• Journal of Microbiology and Biotechnology
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• 제4권2호
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• pp.119-125
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• 1994

Steroid $\Delta^1$-dehydrogenase purified from hydrocortisone-induced cells of Arthrobacter simplex converted various 3-ketosteroids into their corresponding $\Delta^1$-dehydrogenated products. The transformation efficiencies depend upon the chemical structure of the steroids, especially length of the side chain at 17 position and hydroxyl groups at 11 and 17 positions. The Km values for androstenedione, the most favorable substrate examined, and hydrocortisone were 74 ${\mu}M$ and 294 ${\mu}M$, respectively. The optimum temperature and pH of the enzyme reaction were 35$^{\circ}C$ and pH 9, respectively, and the enzyme was relatively stable at the range from 20 to 35$^{\circ}C$ and from pH 5 to 10 after one hour of incubation. The enzyme activity was markedly inhibited in the presence of $Cu^{2+},\;Fe^{3+},\;Hg^{2+},\;Mo^{6+}$ ions, and somewhat inhibited by $Zn^{2+}$ and $Fe^{2+}$. $\alpha,\alpha'$-Dipyridyl that inhibits 9$\alpha$-hydroxylase and accumulates 1,4-androstadiene-3,17-dione from sterols revealed no inhibitory effect on this enzyme. EGTA showed inhibitory effect. $\beta$-Estradiol competitively inhibited the enzyme activity. Chemical modifications of the enzyme were attempted with several reagents. p-Hydroxymer-curibenzoate showed inhibition of the enzyme activity and protection of the substrate. This suggests that cysteine residue may be involved in the active site of the enzyme.

• Fisheries and Aquatic Sciences
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• 제1권2호
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• pp.209-215
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• 1998

Trypsin-like enzyme was purified from shrimp hepatopancreas through Q-Sepharose ionic exchange, benzamidine Sepharose-6B affinity, and Superdex 75 gel chromatography. Purity of trypsin-like enzyme was increased 69-fold with $44\%$ yield. The enzyme consisted of a single polypeptide chain with a molecular weight (M.W.) of 32 kDa judged by sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE). The enzyme was completely inactivated by serine enzyme inhibitors such as soybean trypsin inhibitor (SBTI), tosyl-L­lysine chloromethyl ketone (TLCK), and leupeptin. However, the enzyme was not affected by tosyl-L-phenylalanine chloromethyl ketone (TPCK) which is a chymotrypsin specific inhibitor. The enzyme had no activity against benzoyl-tyrosine ethyl ester (BTEE) which is a chymotrypsin specific substrate. The enzyme showed high activity on the carboxyl terminal of Phe, Tyr. Glu, Arg, and Asp. However. no activity was detected against the carboxyl terminal of Pro, Trp, Cys, Gly, Val, and Ala.

• Food Science and Biotechnology
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• 제17권4호
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• pp.873-877
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• 2008

Native soy protein isolate (SPI) was hydrolyzed with 4 different proteolytic enzymes, including bromelain, papain, Neutrase, and Flavourzyme. SPI hydrolysates with the degree of hydrolysis (DH) in range of 6 to 15% were prepared by each enzyme. The angiotensin 1 converting enzyme (ACE) inhibitory and the antioxidant activities of the SPI hydrolysates, such as superoxide dismutase-like activity and inhibition of the linoleic acid autoxidation, were evaluated. Overall, as the DH increased, all evaluated bioactivities of the SPI hydrolysates significantly increased. The significantly highest ACE inhibitory and antioxidant activities were found in hydrolysates made with papain and bromelain, respectively. SPI hydrolysates by Flavourzyme showed the significantly lowest activity in all tested bioactivities. The results suggested that ACE inhibitory and antioxidant activities of SPI hydrolysates were determined by the DH and by the enzyme used.

• 한국미생물·생명공학회지
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• 제25권4호
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• pp.391-395
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• 1997

It was proved that both ethanol extracts from Mori Folium from Kangwon do and silk worm had higher inhibition acitivity on $\alpha$-glucohydrolase than the water extracts. In adding above 8.5 (mg/L) of silkworm extracts, the inhibition rate on $\alpha$-1,4 glucosidase was saturated while the inhibition rate was continuously increased in adding the extracts from Moli Folium. It was also found that the diethyl ether fraction showed much better inhibition activity than water fraction from ethanol extracts, yielding ca. 85% of inhibition rate for the extract of Moli Folium, compared to 91% for a commercially available hypoglycemic drug, Chloropropamide. In separating the diethyl ether fractions by Consecutive Sephadex gel filtration and Thin layer chromatography, three and four active spots were obtained from Moli Folium and silkworm, respectively. It is interesting that the similar Rf spots from both species among several spots in TLC have the highest inhibition acitivity on a target enzyme, which can imply that the active substances from both species are same or similar molecular weight and structure. Glucose-lowering activities of both speciese were also examined in vivo, showing that the fraction from Moli Folium had better activity than that from silkworm, and its activity was similar to that of a commercial drug.

• Journal of Microbiology and Biotechnology
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• 제2권4호
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• pp.260-267
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• 1992

Pectate lyase from alkalitolerant Bacillus sp. YA-14 is an endo-type pectate lyase which acts randomly at the $\alpha$-1, 4-galacturonan linkage, and requires calcium or strontium ions for its activity. The enzyme is active on low methyl esterified pectin, but the activity toward a high methyl esterified substrate is reduced. The apparent Km's of the enzyme toward sodium polygalacturonic acid, polygalacturonic acid, and various pectins such as apple pectin, citrus pectin, and genu pectin are 0.826 mg/ml, 0.685 mg/ml, and 1.14 mg/ml, respectively. The enzyme activity is inhibited by SDS, urea, and sodium azide, but not by various reducing reagents, such as $\beta$-mercaptoethanol, Na-thiosulfate, Na-sulfate, cystein, and L-ascorbic acid. The enzyme is inactivated by N-bromosuccinimide, $I_2, H_2O_2$. PMSF, and iodoacetate. Judging from the results of their inhibition types, we speculate that tryptophan and serine residues are directly involved in enzyme activity, while tyrosine and methionine residues are indirectly involved in its activity.

• 약학회지
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• 제47권6호
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• pp.432-437
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• 2003

It has been known that quercetin, a bioflavonoid widely distributed in fruits and vegetables as dietary-derived flavonoid and exert significant multiple biological effects such as antioxidant and anti-inflammatory, anti-tumor effects. In addition, it has been shown to have a chemoprotective role in cancer, though complex effects on signal transduction involved in cell proliferation and angiogenesis. The present study investigated whether quercetin can enhance antioxidant enzyme activity (glutathione peroxidase: GPx, superoxide dismutase: SOD, catalase: CAT) and regulate the reactive oxygen species (ROS) generation in the presence of vitamin E, L-ascorbic acid, reduced glutathione (GSH) on B16F10 murine melanoma cells. After 48h treatment of cells with quercetin in the presence of vitamin E, L-ascorbic acid, GSH, we measured the cytotoxicities by MTT assay. The cells exhibited a dose-dependent inhibition in their proliferation in the presence of vitamin E, L-ascorbic acid, GSH respectively. We also investigated the effects of antioxidant enzyme activity and ROS generation. The antioxidant enzyme activity of quercetin in the presence of vitamin E was stronger than GSH, L-ascorbic acid, the same treatments decreased ROS generation in B16F10 murine melanoma cells. Taken together, these result demonstrate that the antioxidant effect of quercetin can enhanced in the presence of vitamin E and it might plays an important role in anti-oxidative effects.

• 미생물학회지
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• 제24권3호
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• pp.251-262
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• 1986

High-molecular-weight ${\beta}-glucosidase$ (EC 3.2.1.21) was purified from the culture filtrate of Trichoderma koningii through a four-step procedure including chromatography on Bio-Gel P-150, DEAE-Sephadex A-50 and SP-Sephadex C-50; and chromatofocusing on Polybuffer exchanger PBE 94. The molecular weight of the enzyme was determined to be about 101,000 by SDS-polyacrylamide gel electrophoreses, and the isoelectric point was estimated to be 4.96 by analytical isoelectric focusing. The temperature optimum for activity was about $55^{\circ}C$, and the pH optimumwas 3.5. The enzyme was considerably thermostable, for no loss of activity was observed when the enzyme was preincubated at $60^{\circ}C$ for 5h. Km values for cellobiose, gentiobiose, sophorose, salicin and $p-nitrophenyl-{\betha}-D-glucoside$ were 99.2, 14.7, 7.09, 3.15 and 0.70 mM, respectively, which indicates that the enzyme has much higher affinity towards $p-nitrophenyl-{\betha}-D-glucoside$ than towards the other substrates, especially cellobiose. Substrate inhibition by $p-nitrophenyl-{\betha}-D-glucoside$ and salicin was observed at the conecntrations exceeding 5mM. Gluconolactone was a powerful inhibitor against the action of the enzyme on $p-nitrophenyl-{\betha}-D-glucoside\;(K_i\;37.9\;{\mu}M)$, wherease glucose was much less effective ($K_i$ 1.95 mM). Inhibition was of the competitive type in each case. Transglucosylation activity was detected shen the readtion products formed from $p-nitrophenyl-{\betha}-D-glucoside$ by the enzyme were analysed using high-performance liquid chromatography.

• Archives of Pharmacal Research
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• 제27권2호
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• pp.199-205
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• 2004

Cytochrome P450 (P450) 1 enzymes such as P450 1A1, 1A2, and 181 are known to be involved in the oxidative metabolism of various procarcinogens and are regarded as important target enzymes for cancer chemoprevention. Previously, several hydroxystilbene compounds were reported to inhibit P450 1 enzymes and were rated as candidate chemopreventive agents. In this study, we investigated the inhibitory effect of 2-[2-(3,5-dimethoxyphenyl)vinyl]-thiophene (DMPVT), produced from the chemical modification of oxyresveratrol, on the activities of P450 1 enzymes. The inhibitory potential by DMPVT on the P450 1 enzyme activity was evaluated with the Escherichia coli membranes of the recombinant human cytochrome P450 1A1, 1A2, or 1B1 coexpressed with human NADPH-P450 reductase. DMPVT significantly inhibited ethoxyresorufin O-deethylation (EROD) activities with $IC_{50}$ values of 61, 11, and 2 nM for 1A1, 1A2, and 1B1, respectively. The EROO activity in OMBA-treated rat lung microsomes was also significantly inhibited by OMPVT in a dose-dependent manner. The modes of inhibition by DMPVT were non-competitive for all three P450 enzymes. The inhibition of P450 1B1-mediated EROD activity by OMPVT did not show the irreversible mechanism-based effect. The loss of EROD activity in P450 1B1 with OMPVT incubation was not blocked by treatment with the trapping agents such as glutathione, N-acetylcysteine, or dithiothreitol. Taken together, the results suggested DMPVT to be a strong noncompetitive inhibitor of human P450 1 enzymes that should be considered as a good candidate for a cancer chemopreventive agent in humans.

• 한국식품과학회지
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• 제29권6호
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• pp.1316-1318
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• 1997

산 casein으로부터 FPLC를 이용하여 gel permeation column으로 ${\kappa}-Casein$을 분획한 다음, 이를 chymosin, pepsin, trypsin 으로 각각 처리하여 3% TCA에서 soluble한 부분을 증류수로 투석(MW cut-off 1kDa)시킨 후 ACE저해 효과를 측정한 결과, trypsin으로 분해 시킨 경우 ACE 저해율이 94.7%로 가장 높게 나타났으며, chymosin 가수분해물은 가장 낮았다. GMP를 투석막의 종류에 따라 투석 시킨 후 $IC_{50}$을 측정한 결과, MW cut-off 의 크기가 증가할수록 ACE저해효과는 감소하는 것으로 나타났으며, MW cut-off 2 kDa의 경우가 가장 높은 저해율을 보였고 MW cut-off 5kDa에서는 저해율이 가장 낮았다.