• Microbiology and Biotechnology Letters
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• v.41 no.2
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• pp.153-159
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• 2013

A mannitol dehydrogenase (MDH; EC 1.1.1.67) gene was cloned from the Sinorhizobium meliloti 1021 (KCTC 2353) genome and expressed in Escherichia coli. It was seen to have an open reading frame consisting of 1,485 bp encoding 494 amino acids (about 54 kDa), which shares approximately 35-55% of amino acid sequence identity with some known long-chain dehydrogenase/ reductase family enzymes. The recombinant S. meliloti MDH (SmMDH) showed the highest activity at $40^{\circ}C$, and pH 7.0 (D-fructose reduction) and pH 9.0 (D-mannitol oxidation), respectively. SmMDH could catalyze the oxidative/reductive reactions between D-mannitol and D-fructose in the presence of $NAD^+/NADH$ as a coenzyme, but not with NADP+/NADPH. These results indicate that SmMDH is a typical $NAD^+/NADH$-dependent mannitol dehydrogenase.

• Journal of the Korean Society of Food Science and Nutrition
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• v.26 no.4
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• pp.682-688
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• 1997

This study was performed to investigate whether ascorbic acid can modulate the induction of CYP2E1 and prevent the lipid peroxidation which may cause diabetic chronic complication. Diabetes was induced by intraperitoneal injection of streptozotocin to 5-week-old male Sprague-Dawley rats(150~170g). Normal and diabetic group was randomly divided into three groups each; Control(CON, no supplementation), SUP1 (50mg/d ascorbate supplementation) and SUP2(250mg/d ascorbate supplementation). Ascobic acid was prepared daily by dissolving in drinking water and supplied for 4 weeks. There was no difference in hepatic microsomal and mitochondrial P450 contents between normal and diabetes. Hepatic microsomal N-nitrosodimethylamine(NDMA) demethylase activity, which repre-sents contents of CYP2E1, was elevated in diabetes, but not significantly. The NDMA demethylase activity of diabetic SUP2 group was significantly lower activity than that of the diabetic CON group. However, no difference in hepatic mitochondrial NDMA demethylase activity was observed between the diabetes and the normal group. The result suggests that the induction of CYP2E1 in diabetes can be alleviated by ascorbic acid supplementation at the dose of 50mg1d. In addition, ascorbic acid supplementation showed dose-dependent reduction of hepatic microsomal TBARS contents in diabetic rats.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.40 no.4
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• pp.504-511
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• 1995

This study was conducted to investigate the superoxide dismutase (SOD) characteristics and antioxidant activity by nonenzymatic(Fe$^{2+}$/Ascorbate) and Fe$^{3+}$-ADP/NADPH method in perilla(Perilla frutescens var. japonica Hara.) and jaso(Perilla frutescens Briton var. acuta Kudo.) leaves. The characteristics were evaluated by the nitro blue tetrazolium reduction method. Perilla leaves contained three or four major SODs depending on the varieties. The inhibitor test indicated that the Perilla leaves contained two Cu /ZnSODs and one or two FeSODs, but Jaso leaves have only Cu/ZnSOD. However, no varietal differences were detected in the Cu /ZnSOD isozyme patterns. FeSODs, however, showed different varietal isozyme patterns through the different combinations of the two FeSOD isozymes. Among MeOH extractes, "mil yang 2" showed very strong antioxidant activity. Relatively large differences in the levels of SOD and antioxidant activity detected in the Perilla varietites. There was significantly different in the comparison between perilla leaves and red jaso leaves.s.etween perilla leaves and red jaso leaves.

• Journal of Pharmacopuncture
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• v.4 no.1
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• pp.123-124
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• 2001

Purpose. Free radicals are implicated in the pathophysiology of aging, ischemic injury and neurodegenerative disorders. To deform]no whether Hominis Placenta extract prevents $H_2O_2$-induced apoptosis, we have performed morphological and biochemical analyses for the detection of apoptotic phenomena in the pineal tumor cell line $PGT-{\beta}$ We have also peformed cytochemical and immunocytochemical analyses for the detection of changes in nitric oxide synthase (NOS) activity and estimated the expression . of apoptotic genes using reverse transcription-polymerase chain reaction (RT-PCR) Methods. $PGT-{\beta}\;cells$ were pretreated with Hominis Placenta extracts $(0,\;10^{-2}\;{\mu}g/ml)$ for 2 hours and then exposed to $H_2O_2\;(0,\;50\;{\mu}M)$ for 3 hours. Appearance of apoptotic characteristics were monitored using 4, 6-diamidino-2-phenylindole dihydrochloride (DAPI) staining assay, terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick end labeling (TUNEL) assay and flow cytometric analysis. NOS activity was measured by NADPH-diaphorase cytochemistry. Expression of inducible NOS (iNOS) and nuclear factor kappa B (NF k B) was assessed via immunocytochemistry. The expression of apoptotic genes was examined by RT-PCR. Results. After 3 flours of exposure to $H_2O_2$, it was shown that $PGT-{\beta}\;cells$ treated with $H_2O_2(50\;{\mu}M)$ exhibit classical apoptotic features and increases in NOS activity and caspase-3 expression. Treatment with Hominis Placenta extract resulted in a reduced occurrence of apoptotic features. DAPI staining, TUNEL and flow cytometric assays revealed decreases in the occurrence of nuclear fragmentation and in the sub-Gl fraction in the $PGT-{\beta}\;cells$ treated with Hominis Placenta extract. Cells treated with Hominis Placenta extract also showed lower activity of NADPH-diaphorase and immunoreactivities of both iNOS and NF k B than those of $H_2O_2$-treated cells which were not treated with Hominis Placenta extract. By RT-PCR, it was shown that the level of caspase-3 mRNA was derreased In the cells treated with Hominis Placenta . extract. Conclusions. This study shows that Hominis Placenta extract prevents $H_2O_2$-induced apoptosis in $PGT-{\beta}\;cells$; inhibitions of iNOS and caspnse-3 are possible mechanisms of the protection against apoptosis.

• Journal of Pharmacopuncture
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• v.18 no.2
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• pp.7-18
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• 2015

Objectives: Lawsone (1,4 naphthoquinone) is a non redox cycling compound that can be catalyzed by DT diaphorase (DTD) into 1,2,4-trihydroxynaphthalene (THN), which can generate reactive oxygen species by auto oxidation. The purpose of this study was to evaluate the toxicity of the phytomarker 1,4 naphthoquinone and its metabolite THN by using the molecular docking program AutoDock 4. Methods: The 3D structure of ligands such as hydrogen peroxide ($H_2O_2$), nitric oxide synthase (NOS), catalase (CAT), glutathione (GSH), glutathione reductase (GR), glucose 6-phosphate dehydrogenase (G6PDH) and nicotinamide adenine dinucleotide phosphate hydrogen (NADPH) were drawn using hyperchem drawing tools and minimizing the energy of all pdb files with the help of hyperchem by $MM^+$ followed by a semi-empirical (PM3) method. The docking process was studied with ligand molecules to identify suitable dockings at protein binding sites through annealing and genetic simulation algorithms. The program auto dock tools (ADT) was released as an extension suite to the python molecular viewer used to prepare proteins and ligands. Grids centered on active sites were obtained with spacings of $54{\times}55{\times}56$, and a grid spacing of 0.503 was calculated. Comparisons of Global and Local Search Methods in Drug Docking were adopted to determine parameters; a maximum number of 250,000 energy evaluations, a maximum number of generations of 27,000, and mutation and crossover rates of 0.02 and 0.8 were used. The number of docking runs was set to 10. Results: Lawsone and THN can be considered to efficiently bind with NOS, CAT, GSH, GR, G6PDH and NADPH, which has been confirmed through hydrogen bond affinity with the respective amino acids. Conclusion: Naphthoquinone derivatives of lawsone, which can be metabolized into THN by a catalyst DTD, were examined. Lawsone and THN were found to be identically potent molecules for their affinities for selected proteins.

• Molecules and Cells
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• v.28 no.4
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• pp.321-329
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• 2009

Rapid production of nitric oxide (NO) and reactive oxygen species (ROS) has been implicated in the regulation of innate immunity in plants. A potato calcium-dependent protein kinase (StCDPK5) activates an NADPH oxidase StRBOHA to D by direct phosphorylation of N-terminal regions, and heterologous expression of StCDPK5 and StRBOHs in Nicotiana benthamiana results in oxidative burst. The transgenic potato plants that carry a constitutively active StCDPK5 driven by a pathogen-inducible promoter of the potato showed high resistance to late blight pathogen Phytophthora infestans accompanied by HR-like cell death and $H_2O_2$ accumulation in the attacked cells. In contrast, these plants showed high susceptibility to early blight necrotrophic pathogen Alternaria solani, suggesting that oxidative burst confers high resistance to biotrophic pathogen, but high susceptibility to necrotrophic pathogen. NO and ROS synergistically function in defense responses. Two MAPK cascades, MEK2-SIPK and cytokinesis-related MEK1-NTF6, are involved in the induction of NbRBOHB gene in N. benthamiana. On the other hand, NO burst is regulated by the MEK2-SIPK cascade. Conditional activation of SIPK in potato plants induces oxidative and NO bursts, and confers resistance to both biotrophic and necrotrophic pathogens, indicating the plants may have obtained during evolution the signaling pathway which regulates both NO and ROS production to adapt to wide-spectrum pathogens.

• Archives of Pharmacal Research
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• v.27 no.1
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• pp.48-52
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• 2004

DA-11004 is a synthetic, potent NADP-dependent isocitrate dehydrogenase (IDPc) inhibitor where $IC_{50}$ for IDPc is 1.49 $\mu$M. The purpose of this study was to evaluate the effects of DA-11004 on the high fat high sucrose (HF)-induced obesity in male C57BL/6J mice. After completing a 8-week period of experimentation, the mice were sacrificed 1hr after the last DA-11004 treatment and their blood, liver, and adipose tissues (epididymal and retroperitoneal fat)were collected. There was a significant difference in the pattern of increasing body weight between the HF control and the DA-11004 group. In the DA-11004 (100 mg/kg) treated group the increase in body weight significantly declined and a content of epididymal fat and retroperitoneal fat was also significantly decreased as opposed to the HF control. DA-11004 (100 mg/kg) inhibited the IDPc activity, and thus, NADPH levels in plasma and the levels of free fatty acid (FFA) or glucose in plasma were less than the levels of the HF control group. In conclusion, DA-11004 inhibited the fatty acid synthesis in adipose tissues via IDPc inhibition, and it decreased the plasma glucose levels and FFA in HF diet-induced obesity of C57BL/6J mice.

• Journal of Microbiology and Biotechnology
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• v.27 no.5
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• pp.956-964
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• 2017

Compactin and pravastatin are competitive cholesterol biosynthesis inhibitors of 3-hydroxy-3-methylglutaryl-CoA reductase and belong to the statin drugs; however, the latter shows superior pharmacokinetic characteristics. Previously, we reported that the bacterial P450, CYP105D7, from Streptomyces avermitilis can catalyze the hydroxylation of 1-deoxypentalenic acid, diclofenac, and naringenin. Here, we demonstrate that CYP105D7 could also catalyze compactin hydroxylation in vitro. In the presence of both bacterial and cyanobacterial redox partner systems with an NADPH regeneration system, the reaction produced two hydroxylated products, including pravastatin (hydroxylated at the C6 position). The steady-state kinetic parameters were measured using the redox partners of putidaredoxin and its reductase. The $k_m$ and $k_{cat}$ values for compactin were $39.1{\pm}8.8{\mu}M$ and $1.12{\pm}0.09min^{-1}$, respectively. The $k_{cat}/K_m$ value for compactin ($0.029min^{-1}{\cdot}{\mu}M^{-1}$) was lower than that for diclofenac ($0.114min^{-1}{\cdot}{\mu}M^{-1}$). Spectroscopic analysis showed that CYP105D7 binds to compactin with a $K_d$ value of $17.5{\pm}3.6{\mu}M$. Molecular docking analysis was performed to build a possible binding model of compactin. Comparisons of different substrates with CYP105D7 were conclusively illustrated for the first time.

• Proceedings of the Botanical Society of Korea Conference
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• 1999.08a
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• pp.83-90
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• 1999

Electron crystallography of two-dimensional crystalsand electron cryo-microscopy is becoming an established method for determining the structure and function of a variety of membrane proteins that are providing difficult to crystallize in three dimension. In this study this technique has been used to investigate the structure of a ~160 kDa reaction centre sub-core complex of photosystem II. Photosystem II is a photosynthetic membrane protein consisting of more than 25 subunits. It uses solar energy to split water releasing molecular oxygen into the atmosphere and creates electrochemical potential across the thylakoid membrane, which is eventually utilized to generate ATP and NADPH. Images were taken using Philips CM200 field emission gun electron microscope with an acceleration voltage of 200kW at liquid nitrogen temperature. In total, 79 images recorded dat tilt angles ranging from 0 to 67 degree yielded amplitudes and phases for a three-dimensional map with an in-plant resolution of 6$\AA$ and 11.4$\AA$ in the third dimension shows at least 23 transmembrane helices resolved in a monomeric complex, of which 18 were able to be assigned to the D1, D2, CP47 , and cytochrome b559 alfa beta-subunits with their associated pigments that ae active in electron transport (Rhee, 1998, Ph.D.thesis). The D1/D2 heterodimer is located in the central position within the complex and its helical scalffold is remarkably similar to that of the reaction centres not only in purple bacteria but also in plant photosystem I (PSI) , indicating a common evoluationary origin of all types of reaction centre in photosynthetic organism known today 9RHee et al. 1998). The structural homology is now extended to the inner antenna subunit, ascribed to CP47 in our map, where the 6 transmembrane helices show a striking structural similarity to the corresponding helices of the PSI reaction centre proteins. The overall arrangement of the chlorophylls in the D1 /D2 heterodimer, and in particular the distance between the central pair, is ocnsistent with the weak exciton coupling of P680 that distinguishes this reaction centre from bacterial counterpart. The map in most progress towards high resolution structure will be presented and discussed.

• Journal of Microbiology and Biotechnology
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• v.28 no.8
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• pp.1346-1351
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• 2018

Oxidoreductases are effective biocatalysts, but their practical use is limited by the need for large quantities of NAD(P)H. In this study, a whole-cell biocatalyst for NAD(P)H cofactor regeneration was developed using the economical substrate glycerol. This cofactor regeneration system employs permeabilized Escherichia coli cells in which the glpD and gldA genes were deleted and the gpsA gene, which encodes $NAD(P)^+-dependent$ glycerol-3-phosphate dehydrogenase, was overexpressed. These manipulations were applied to block a side reaction (i.e., the conversion of glycerol to dihydroxyacetone) and to switch the glpD-encoding enzyme reaction to a gpsA-encoding enzyme reaction that generates both NADH and NADPH. We demonstrated the performance of the cofactor regeneration system using a lactate dehydrogenase reaction as a coupling reaction model. The developed biocatalyst involves an economical substrate, bifunctional regeneration of NAD(P)H, and simple reaction conditions as well as a stable environment for enzymes, and is thus applicable to a variety of oxidoreductase reactions requiring NAD(P)H regeneration.