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Cho S.S.,Sungkyunkwan University | Sun Y.,Sungkyunkwan University | Yu M.,Sungkyunkwan University | Kwon S.H.,Rexgene Biotech Co. | Kwon S.-T.,Sungkyunkwan University
Enzyme and Microbial Technology | Year: 2012

We cloned and sequenced the gene encoding Thermococcus pacificus dUTPase (Tpa dUTPase). The Tpa dUTPase gene consists of 471. bp and encodes a 156-amino acid protein. The deduced amino acid sequence of Tpa dUTPase has high sequence similarity with other archaeal dUTPases. The Tpa dUTPase had an 18-kDa major protein band consistent with the 17,801. Da molecular mass calculated based on the amino acid sequence. The specific activity of Tpa dUTPase on dUTP at 85. °C was 90,909. U/mg. For Tpa dUTPase activity, we determined an optimum pH of 8.5 and temperature of 85. °C. Magnesium ions strongly induced activity, with an optimum concentration of 0.75. mM. The half-life of the enzyme at 94. °C was about 7. h. The specific activity of the Tpa dUTPase on dUTP was about 10-20-fold higher than that of Tpa dUTPase on dCTP. Tpa dUTPase enhanced the PCR amplification efficiency of long targets when Pfu and Vent DNA polymerases were used. © 2012 Elsevier Inc. Source


Jung I.-S.,Konkuk University | Lee S.-H.,Konkuk University | Yang M.-K.,Konkuk University | Park J.-W.,Konkuk University | And 6 more authors.
Archives of Pharmacal Research | Year: 2010

To investigate the cardioprotective effects and mechanism of action of KR-32560 {[5-(2-methoxy-5-fluorophenyl)furan-2-ylcarbonyl]guanidine}, a newly synthesized NHE-1 inhibitor, we evaluated the effects of KR-32560 on cardiac function in a rat model of ischemia/reperfusion (I/R)-induced heart injury as well as the role antioxidant enzymes and pro-survival proteins play these observed effects. In isolated rat hearts subjected to 25 min of global ischemia followed by 30 min of reperfusion, KR-32560 (3 and 10 μM) significantly reversed the I/Rinduced decrease in left ventricular developed pressure and increase in left ventricular enddiastolic pressure. In rat hearts reperfused for 30 min, KR-32560 (10 μM) significantly decreased the malondialdehyde content while increasing the activities of both glutathione peroxidase and catalase, two important antioxidant enzymes. Western blotting analysis of left ventricles subjected to I/R showed that KR-32560 significantly increased phosphorylation of both Akt and GSK-3β in a dose-dependent manner, with no effect on the phosphorylation of eNOS. These results suggest that KR-32560 exerts potent cardioprotective effects against I/Rinduced rat heart injury and that its mechanism involves antioxidant enzymes and the Akt-GSK-3β cell survival pathway. © 2010 The Pharmaceutical Society of Korea and Springer Netherlands. Source


Jung J.-C.,Rexgene Biotech Co. | Moon H.-I.,Dongguk University | Moon H.-I.,Wonkwang University
Pharmaceutical Biology | Year: 2011

Context: Coumarins are natural substances found in a variety of plants. It is well known that plant-derived natural products are extensively used as biologically active compounds. Among them, coumarins were the first preservatives used by man, originally in their natural state within plant tissues and then as natural materials obtained by water distillation. During our search for new types of coumarin derivatives possessing a larvicidal activity, we investigated the synthesis of 4-hydroxycoumarin derivatives. Objective: The coumarin derivatives were synthesized and the structure determination and larvicidal effects were studied. Materials and methods: The structure analyses were conducted by nuclear magnetic resonance (NMR), and mass (MS) spectroscopy revealed that the coumarin derivatives were obtained in good yields, and the eight coumarin derivatives were 3-{1,2,3,4-tetrahydro-3-[4-(4- trifluoromethylbenzyloxy)phenyl}-1-naphthalen-1-on (1), 3-{1,2,3,4-tetrahydro-3- [4-(4-trifluoro methylbenzyloxy)phenyl}-1-naphthalen-1-ol (2), brodifacoum (3), difethialone (4), bromadiolone (5), 4-hydroxy-3-(1,2,3,4-tetrahydronaphthalen-1- yl)-2H-chromen-2-one (coumatetralyl) (6), cis-flocoumafen (7) and trans-flocoumafen (8). Results: The compounds were tested against the F 21 laboratory strain of Aedes aegypti L. Brodifacoum and cis-flocoumafen mediated strong activity with an LC50 values of 8.23 and 9.34 ppm, respectively. Discussion and conclusion: The above indicates that brodifacoum may play a more important role in the toxicity of 4-hydroxycoumarin derivatives. © 2011 Informa Healthcare USA, Inc. Source


Son D.J.,Chungbuk National University | Jung J.C.,Rexgene Biotech Co. | Hong J.T.,Chungbuk National University
PLoS ONE | Year: 2016

Microtubule stabilizing agents (MTSA) are known to inhibit vascular smooth muscle cell (VSMC) proliferation and migration, and effectively reduce neointimal hyperplasia and restenosis. Epothilones (EPOs), non-taxane MTSA, have been found to be effective in the inhibition of VSMC proliferation and neointimal formation by cell cycle arrest. However, effect of EPOs on apoptosis in hyper-proliferated VSMCs as a possible way to reduce neointimal formation and its action mechanism related to VSMC viability has not been suited yet. Thus, the purposes of the present study was to investigate whether EPOs are able to inhibit neointimal formation by inducing apoptosis within the region of neointimal hyperplasia in balloon-injured rat carotid artery, as well as underlying action mechanism. Treatment of EPO-B and EPO-D significantly induced apoptotic cell death and mitotic catastrophe in hyper-proliferated VSMCs, resulting in cell growth inhibition. Further, EPOs significantly suppressed VSMC proliferation and induced apoptosis by activation of p53-dependent apoptotic signaling pathway, Bax/cytochrome c/caspase-3. We further demonstrated that the local treatment of carotid arteries with EPOs potently inhibited neointimal lesion formation by induction of apoptosis in rat carotid injury model. Our findings demonstrate a potent antineointimal hyperplasia property of EPOs by inducing p53-depedent apoptosis in hyper-proliferated VSMCs. © 2016 Son et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Source


Kim J.-H.,Sungkyunkwan University | Lee K.-K.,Sungkyunkwan University | Sun Y.,Sungkyunkwan University | Seo G.-J.,Sungkyunkwan University | And 3 more authors.
Extremophiles | Year: 2013

The nucleotide cofactor specificity of the DNA ligase from the hyperthermophilic crenarchaeon Hyperthermus butylicus (Hbu) was studied to investigate the evolutionary relationship of DNA ligases. The Hbu DNA ligase gene was expressed under control of the T7lac promoter of pTARG in Escherichia coli BL21-CodonPlus(DE3)-RIL. The expressed enzyme was purified using the IMPACT™-CN system (intein-mediated purification with an affinity chitin-binding tag) and cation-ion (Arg-tag) chromatography. The optimal temperature for Hbu DNA ligase activity was 75 °C, and the optimal pH was 8. 0 in Tris-HCl. The activity was highly dependent on MgCl2 or MnCl2 with maximal activity above 5 mM MgCl2 and 2 mM MnCl2. Notably, Hbu DNA ligase can use ADP and GTP in addition to ATP. The broad nucleotide cofactor specificity of Hbu DNA ligase might exemplify an undifferentiated ancestral stage in the evolution of DNA ligases. This study provides new evidence for possible evolutionary relationships among DNA ligases. © 2013 Springer Japan. Source

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