• Journal of Microbiology and Biotechnology
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• v.14 no.4
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• pp.789-795
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• 2004

A promoter-probe shuttle vector pSK1Cat was constructed for the isolation of transcriptional signal sequences from Corynebacterium glutamicum. Besides conferring resistance to kanamycin in Escherichia coli and C. glutamicum, the vector carried a promoterless cat gene to confer resistance to chloramphenicol upon insertion of the appropriate transcriptional signals in the multiple cloning site. By utilizing the vector, a series of transcriptionally active fragments were isolated from the genome of C. glutamicum. The clones, ranging from 200 bp to 1 kb in size, were grouped into 3 classes of strong, medium, and weak, based on the chloramphenicol acetyltransferase (CAT) activity and sensitivity to the chloramphenicol of the clone-carrying C. glutamicum cells. C. glutamicum cells carrying the $P_{19}$ clone, a representative in the strong class, were able to grow on minimal agar plates containing over $40 mg/mell$ chloramphenicol, and showed CAT activity of 10 m㏖/mgㆍmin, performing slightly better than the cells carrying $P_{tac}$ , a strong E. coli promoter. Subcloning analysis of the $P_{19}$ clone identified a 180 bp intergenic fragment ($P_{180}$), which was located upstream of a gene encoding a hypothetical membrane protein. The expression conferred by $P_{180}$ was not affected by either the kinds of carbon sources or changes in temperature. These properties make the $P_{180}$ clone useful for the deregulated expression of biosynthetic genes in C. glutamicum during amino acid fermentation.

• Korean Journal of Plant Resources
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• v.25 no.3
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• pp.349-356
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• 2012

The antioxidant potential and enzyme activities in Salicornia herbacea, Corylopsis coreana, Erythronium japonicum, Phragmites communis, Momordica Charantia, Nelumbo nucifera, Salvia plebeia, Portulaca oleracea, Ficus carica, Citrus junos and Cornus officinalis were determined. Their antioxidant activities were measured using DPPH radical scavenging and nitrite scavenging activity. Enzyme activities in investigated plants were evaluated as superoxide dismutase (SOD), catalase (CAT), peroxidase (POD), and ascorbate peroxidase (APX). The DPPH scavenging rate from 100 to 2500 $mgL^{-1}$ was the highest in the flower of Corylopsis coreana. However, it was not detected in most of the samples at concentration below 100 $mgL^{-1}$. The nitrite scavenging activity according to each kind of resource plants was significantly higher in the stem of Corylopsis coreana and leaf of Nelumbo nucifera. The root extract of Erythronium japonicum had the highest SOD enzyme activity of 94.0% while leaf of Salvia plebeia showed the lowest SOD enzyme activity of 30.4%. The activity of CAT and APX showed higher values in the stem of Corylopsis coreana, root of Erythronium japonicum and root of Phragmites communis in comparison with other plants. The activity of POD showed significantly high values in stem of Corylopsis coreana, Momordica Charantia and pericarp of Citrus junos extracts. The antioxidant enzyme activities differ significantly in different plants. In conclusion, we showed that Corylopsis coreana, Erythronium japonicum Cornus officinalis, and Momordica Charantia had the potent biological activities. Therefore, these plant resources showing antioxidant activity could be good materials for development of source of functional healthy food.

• Imaging Science in Dentistry
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• v.53 no.2
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• pp.127-135
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• 2023

Purpose: This study assessed the intensity of artifacts produced by 2 metal posts, 2 cements, and different exposure parameters using 2 cone-beam computed tomography (CBCT) units. Materials and Methods: The sample was composed of 20 single-rooted premolars, divided into 4 groups: Ni-Cr/zinc phosphate, Ni-Cr/resin cement, Ag-Pd/zinc phosphate, and Ag-Pd/resin cement. Samples were scanned before and after post insertion and cementation using a CS9000 3D scanner with 4 exposure parameters (85/90 kV and 6.3/10 mA) and an i-CAT scanner with 120 kV and 5 mA. The presence of artifacts was assessed subjectively by 2 observers and objectively by a trained observer using ImageJ software. The Mann-Whitney, Wilcoxon, weighted kappa, and chi-square tests were used to assess data at a 95% confidence level(α<0.05). Results: In the subjective analyses, AgPd presented more hypodense and hyperdense lines than NiCr (P<0.05), and more hypodense halos were found using i-CAT (P<0.05) than using CS9000 3D. More hypodense halos, hypodense lines, and hyperdense lines were observed at 10 mA than at 6.3 mA (P<0.05). More hypodense halos were observed at 85 kV than at 90 kV (P<0.05). CS9000 3D presented more hypodense and hyperdense lines than i-CAT (P<0.05). In the objective analyses, AgPd presented higher percentages of hyperdense and hypodense artifacts than NiCr (P<0.05). Zinc phosphate cement presented higher hyperdense artifact percentages on CS9000 3D scans(P<0.05). CS9000 3D presented higher artifact percentages than i-CAT(P<0.05). Conclusion: High-atomic-number alloys, higher tube current, and lower tube voltage may increase the artifacts present in CBCT images.

• Journal of Life Science
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• v.28 no.6
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• pp.656-662
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• 2018

Flavin-containing monooxygenases from Corynebacterium (cFMOs) were mutagenized based on homology modeling to develop variants with an enhanced indigoid production capability. The four mutants, F170Y, A210G, A210S, and T326S, which fused to a maltose-binding protein (MBP), were constructed, and their biochemical properties were characterized. Of these, purified MBP-T326S required a higher concentration of exogenous FAD (100 mM) than the wild-type MBP-cFMO for optimal activity and showed a 3.8-fold increase in the $k_{cat}/K_m$ value at $100{\mu}M$ FAD compared to that of MBP-cFMO at $2{\mu}M$ FAD. The indole oxygenase activities of MBP-T326S decreased to 63-77% compared to that of the MBP-cFMO In addition, MBP-T326S displayed a very low level of futile NADPH oxidase activities (21-24%) in the absence of a substrate. Mutant proteins except for T326S displayed similar $K_m$ and increased $k_{cat}/K_m$ values compared to the wild-type. MBP-F170Y and -A210S mutants showed elevated indole oxygenase activity higher than 3.1- and 2.9-fold, respectively, in comparison with MBP-cFMO. When indigoid production was carried out in LB broth with 2.5 g/l of tryptophan, Escherichia coli expressing cFMO produced 684 mg/l of indigo and 104 mg/l of indirubin, while cells harboring T326S produced 1,040 mg/l of indigo and 112 mg/l of indirubin. The results indicate that the production of indigo was 13% higher when compared to a previous report in which an E. coli expressing FMO from Methylophaga produced 920 mg/l of indigo. The protein engineering of cFMO based on homology modeling provided a more rational strategy for developing indigoid-producing strains.

• Korean Journal of Poultry Science
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• v.41 no.3
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• pp.205-215
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• 2014

The aim of this study was to determine the effects of stocking density and strain on the performance and physiological adaptive responses including the plasma corticosterone content and the level of mRNA expression of pro-inflammatory cytokines and antioxidant enzymes in broiler chicks. A total of 300 birds of two strains (150 Ross strain vs. 150 Cobb strain) aged 3-d old were allotted into two stocking densities (standard stocking density,$0.046m^2/bird$ vs. high stocking density, $0.023m^2/bird$) in battery cages by $2{\times}2$ factorial designs with ten replicates until 35 d of age. There was no significant strain effect on body weight, feed intakes and feed to gain ratio and the relative organ weights. However body weight, feed intakes and relative organ weight were found to be significantly (P<0.05) affected by the effect of stocking density. Plasma corticosterone level was not affected by both stocking density and strain effects. Hepatic mRNA expression of pro-inflammatory cytokines including interleukin-$1{\beta}$ (IL-$1{\beta}$), IL-6, IL-18 and interferon-gamma (IFN-${\gamma}$) was not significantly changed by the effects of strain and stocking density. However, the mRNA expression of glutathione peroxidase (GPX) was affected by strain, showing that Ross strain decreased (P<0.05) the GPX expression. With respect to the effect of stocking density, there was a significant (P<0.05) increase in superoxide dismutase (SOD) and catalase (CAT) and GPX mRNA expression in the liver from high stocking density group. Splenic pro-inflammatory cytokine expression was not also affected by stocking density and strain, except that IL-18 mRNA significantly (P<0.05) decreased in Cobb strain under high stocking density. The mRNA expression of SOD and CAT was significantly (P<0.05) affected by the effects of stocking density and strain. In conclusion, growth performance was not affected by strain but stocking density. Although mRNA expression of major pro-inflammatory cytokines was not changed by stocking density and strain, antioxidant enzyme was significantly affected by stocking density, strain or even organ in birds under summer conditions. More detailed studies still needed to be explored to elucidate the effects of environmental conditions and genetic background on physiological responses in birds.

• Journal of the Korean Wood Science and Technology
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• v.35 no.6
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• pp.159-165
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• 2007

An extracellular xylanase from the brown-rot fungus Fomitopsis palustris grown on rice straw culture was purified to a single protein band. On SDS-PAGE, the molecular mass of purified enzyme was estimated to be about 43 kDa. The amino acid sequence of the proteolytic fragments showed high homology with fungal glycoside hydrolase family 10 xylanases. The $K_m$, $K_{cat}$ and $V_{max}$ for birch xylan were $31mg/m{\ell}$, $2.3{\times}10^4/min$ and 252.3 U/mg, respectively. The optimal activity of the purified xylanase from F palustris was observed at pH 4.0~5.0 and $70^{\circ}C$.

• BMB Reports
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• v.50 no.1
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• pp.31-36
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• 2017

High-dose caffeine uptake is a developmental stressor and causes food-avoidance behavior (aversion phenotype) in C. elegans, but its mode of action is largely unknown. In this study, we investigated the molecular basis of the caffeine-induced aversion behavior in C. elegans. We found that aversion phenotype induced by 30 mM caffeine was mediated by JNK/MAPK pathway, serotonergic and dopaminergic neuroendocrine signals. In this process, the dopaminergic signaling appears to be the major pathway because the reduced aversion behavior in cat-2 mutants and mutants of JNK/MAPK pathway genes was significantly recovered by pretreatment with dopamine. RNAi depletion of hsp-16.2, a cytosolic chaperone, and cyp-35A family reduced the aversion phenotype, which was further reduced in cat-2 mutants, suggesting that dopaminergic signal is indeed dominantly required for the caffeine-induced food aversion. Our findings suggest that aversion behavior is a defense mechanism for worms to survive under the high-dose caffeine conditions.

• Proceedings of the Korean Society of Crop Science Conference
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• 2017.06a
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• pp.203-203
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• 2017

The Earth's climate is rapidly changing because of increasing carbon dioxide content in atmosphere so, climate prediction models anticipate that earth surface temperature will rise by 3 to $5^{\circ}C$ in next 50 to 100 years. Therefore, frequency of un-expected weather events such as drought, salinity, low or high temperature and flooding etc. will be increasing worldwide. Furthermore, increased atmosphere temperature can influence pests and pathogens spread as well. Therefore, to protect enormous grain loss from unexpected weather conditions, studies related with combine stress conditions like abiotic plus biotic stress condition are really required. Thus, our research focused on physiological responses under combined abiotic and biotic stress condition in rice plant. To induce uniform stress condition, we used NaCl (100 mM) and salicylic acid (0.5 and 1.0 mM SA) as each stress a stimulator. Each artificial abiotic and biotic stress inducer was applied to hydroponically grown rice seedlings alone or together for four day. The data were collected in a time-dependent manner [1, 2, 3 and 4 day(s) after treatment (DAT)] and were matched with our anticipation that shoot length and shoot fresh weight was decreased in solo and combined abiotic and biotic stress condition. The lipid peroxidation content was significantly increased ($1.5{\pm}0.2$ to $2.7{\pm}0.1mg$ mg of $MDA\;g^{-1}FW$) in the first two days in both stress exposed plants, and showed the opposite trend ($0.5{\pm}0.01$ to $0.1{\pm}0.001mg$ of $MDA\;g^{-1}FW$) in last two days under multi stress condition. Superoxide dismutase (SOD) activity did not showed difference in only biotic stress condition (alone 0.5 and 1.0 mM SA) as compared to control however, it was significantly increased in multi stress condition or solo abiotic stress condition whereas, catalase (CAT), and ascorbate peroxidase (APX) activities were significantly decreased in solo biotic and combined abiotic and biotic condition. In particular, both enzymes activities were more decreased in multi stress condition as compared to solo biotic stress condition. The results for relative mRNA expression level of CAT and APX enzymes were in agreement with results of spectrophotometric values. Correlation value between each stress condition and phenotypic data showed that biotic stress condition showed high correlation with activity of CAT and APX whilst, abiotic stress condition revealed significant correlation with SOD activity.

• Proceedings of the KAIS Fall Conference
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• 2010.11a
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• pp.495-499
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• 2010

The biocatalytic capture of $CO_2$, and its precipitationas $CaCO_3$, over bovine carbonic anhydrase (BCA) immobilized on a pore-expanded SBA-15 support was investigated. SBA-15 was synthesized using TMB as a pore expander, and the resulting porous silica was characterized by XRD, BET, IR, and FE-SEM analysis. BCA was immobilized on SBA-15 through various approaches, including covalent attachment (BCA-CA), adsorption (BCA-ADS), and cross-linked enzyme aggregation (BCA-CLEA). The immobilization of BCA on SBA-15 was confirmed by the presence of zinc metal in the EDXS analysis. The effects of pH, temperature, storage stability, and reusability on the biocatalytic performance of BCA were characterized by examining para-nitrophenyl acetate (p-NPA) hydrolysis. The $K_{cat}/K_m$ values for p-NPA hydrolysis were 740.05, 660.62, and $680.11M^{-1}s^{-1}$, respectively, where as $K_{cat}/K_m$ for free BCA was $873.76M^{-1}s^{-1}$. The amount of $CaCO_3$ precipitate was measured quantitatively using anion-selective electrode and was found to be 12.41, 11.82, or 11.28 mg $CaCO_3$/mg for BCA-CLEA, BCA-ADS, or BCA-CA, respectively. The present results indicate that the immobilized BCA-CLEA, BCA-ADS, and BCA-CA are green materials, and are tunable, reusable, and promising biocatalysts for $CO_2$ sequestration.

• Biomolecules & Therapeutics
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• v.20 no.6
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• pp.562-568
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• 2012

Cytochrome P450 BM3 (CYP102A1) from Bacillus megaterium is a self-sufficient monooxygenase that consists of a heme domain and FAD/FMN-containing reductase domain (BMR). In this report, the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and 5-cyano-2,3-ditolyl tetrazolium chloride (CTC) by BMR was evaluated as a method for monitoring BMR activity. The electron transfer proceeds from NADPH to BMR and then to BMR substrates, MTT and CTC. MTT and CTC are monotetrazolium salts that form formazans upon reduction. The reduction of MTT and CTC followed classical Michaelis-Menten kinetics ($k_{cat}=4120\;min^{-1}$, $K_m=77{\mu}M$ for MTT and $k_{cat}=6580\;min^{-1}$, $K_m=51{\mu}M$ for CTC). Our continuous assay using MTT and CTC allows the simple, rapid measurement of BMR activity. The BMR was able to metabolize mitomycin C and doxorubicin, which are anticancer drug substrates for CPR, producing the same metabolites as those produced by CPR. Moreover, the BMR was able to interact with CYP1A2 and transfer electrons to promote the oxidation reactions of substrates by CYP1A2 and CYP2E1 in humans. The results of this study suggest the possibility of the utilization of BMR as a surrogate for mammalian CPR.