• BMB Reports
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• v.34 no.1
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• pp.15-20
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• 2001

One of the innate immune reactions in invertebrates is the pro-phenoloxidase (pro-PO) activation system that is involved in the generation of superoxide, melanin synthesis, and the subsequent sequestration of foreign matter entering the hemocoel of the invertebrates. However, the molecular mechanism of this biological reaction is still obscure. To expand our understanding of the biological roles of the pro-PO activation system in invertebrates, we performed a yeast two-hybrid screening by using three regions of pro-PO as bait and a yeast two-hybrid cDNA library from Tenebrio molitor larvae as prey We isolated a novel partial cDNA clone that encodes a glycine-rich protein that interacted with the active phenoloxidase (termed phenoloxidase interacting protein, POIP). POIP consists of two domains: One is an N-terminal unique domain and the other is a C-terminal glycine-rich domain. The C-terminal glycine-rich domain showed sequential homology with those of insect antifungal proteins. Also, the yeast two-hybrid screen in a reverse orientation (using POIP as bait) yielded PO, suggesting that the PO-POIP interaction is specific. By using a 315 bP PCR fragment of the N-terminal unique region of POIP, we cloned the full-length cDNA of POIP from the Tenebruo cDNA library constructed by using E. coli injected larvae. The interaction analysis between PO, and a truncated fragment lacking the N-terminal unique region of POIP, indicated that the N-terminal unique region is necessary for interaction between PO and POIP. The expression level of the POIP mRNA is increased by bacterial injection into T. molitor larvae. This suggests that POIP might be engaged in the humoral defense reaction.

• Proceedings of the Korean Vacuum Society Conference
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• 2000.02a
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• pp.60-60
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• 2000

The adhesion interface formation between copper and poly(ethylene terephthalate)(PET), poly(methyl methacrylate)(PMMA) and Polyimide films was treated using Ion assisted reaction system to sequential sputter deposition by High-Frequency ion source. The ion beam modification system used a new type of low power HF ion thruster for space application as new low thruster electric propulsion system. Low power HF ion thruster with diameter 100mm gives the opportunity to obtain beams of Ar+ with currents 20~150 mA (current density 0.5~3.5 mA/cm2) and energy 200~2500eV at HF power level 10~150 W. Using Ar as a working gas it is possible to obtain thrust within 3~8 mN. Contact angles for untreated films were over 95$^{\circ}$ and 80 for Pet, 10o for PMMA and 12o for PI samples as a condition of ion assisted reaction at the ion dose of 10$\times$1016 ions/cm2, the ion beam potential of 1.2 keV and 4 ml/min for environmental gas flow rate. 900o peel tests yielded values of 15 to 35 for PET, 18 to 40 and 12 to 36 g/min. respectively. High resolution X-ray photoelectron spectrocopy is the Cls region for Cu metal on these polymer substrates showed increases in C=O-O groups for polymide, whereas PET and PMMA treated samples showed only C=O groups with increase the ion dose. Finally, unstable polymer surface can be changed from hydrophobic to hydrophilic formation such as C-O and C=O that were confirmed by the XPS analysis, conclusionally, the ion assisted reaction is very effective tools to attach reactive ion species to form functional groups on C-C bond chains of PET, PMMA and PI.

• Bulletin of the Korean Chemical Society
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• v.15 no.5
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• pp.394-399
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• 1994

The catalytic mechanism of malonyl-CoA synthetase from Rhizobium trifolii was investigated by the steady state kinetics and intermediate identification. Initial velocity studies and the product inhibition studies with AMP and PPi strongly suggested ordered Bi Uni Uni Bi Ping-Pong Ter Ter system as the most probable steady state kinetic mechanism of malonyl-CoA synthetase. Michaelis constants were $0.17{\pm}0.04 {\mu}M,\;0.24{\pm}0.18 {\mu}M\;and\;0.045{\pm}0.26 {\mu}$M for ATP, malonate and CoA, respectively. The TLC analysis of the $^{32}P-labelled$ products in reaction mixture containing $[{\gamma}-^{32}P]$ ATP in the absence of CoA showed that PPi was produced after the sequential addition of ATP and malonate. Formation of malonyl-AMP, suggested as an intermediate in the kinetically deduced mechanism, was confirmed by the analysis of $^{31}P-NMR$ spectra of AMP product isolated from the $^{18}O$ transfer experiment using $[^{18}O]$malonate. Two resonances were observed, corresponding to AMP labelled with zero and one atom of $^{18}O$, indicating that one atom of $^{18}O$ transferred from $[^{18}O]$malonate to AMP through the formation of malonyl-AMP. Formation of malonyl-AMP was also confirmed through the TLC analysis of reaction mixture containing $[{\alpha}-^{32}P]$ATP. These results strongly support the ordered Bi Uni Uni Bi Ping-Pong Ter Ter mechanism deduced from the initial velocity and product inhibition studies.

• Journal of Microbiology and Biotechnology
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• v.28 no.11
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• pp.1859-1864
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• 2018

Synthesis of flavonoid glycoside is difficult due to diverse hydroxy groups in flavonoids and sugars. As such, enzymatic synthesis or biotransformation is an approach to solve this problem. In this report, we used stepwise biotransformation to synthesize two quercetin bisglycosides (quercetin 3-O-glucuronic acid 7-O-rhamnoside [Q-GR] and quercetin 3-O-arabinose 7-O-rhamnoside [Q-AR]) because quercetin O-rhamnosides contain antiviral activity. Two sequential enzymatic reactions were required to synthesize these flavonoid glycosides. We first synthesized quercetin 3-O-glucuronic acid [Q-G], and quercetin 3-O-arabinose [Q-A] from quercetin using E. coli harboring specific uridine diphopsphate glycosyltransferase (UGT) and genes for UDP-glucuronic acid and UDP-arabinose, respectively. With each quercetin 3-O-glycoside, rhamnosylation using E. coli harboring UGT and the gene for UDP-rhamnose was conducted. This approach resulted in the production of 44.8 mg/l Q-GR and 45.1 mg/l Q-AR. This stepwise synthesis could be applicable to synthesize various natural product derivatives in case that the final yield of product was low due to the multistep reaction in one cell or when sequential synthesis is necessary in order to reduce the synthesis of byproducts.

• Bulletin of the Korean Chemical Society
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• v.35 no.6
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• pp.1633-1638
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• 2014

[6]-Gingerol is known as the major bioactive constituent of ginger. In the study, it was observed to effectively protect against ${\bullet}OH$-induced DNA damage ($IC_{50}$ $328.60{\pm}24.41{\mu}M$). Antioxidant assays indicated that [6]-gingerol could efficiently scavenge various free radicals, including ${\bullet}OH$ radical ($IC_{50}$ $70.39{\pm}1.23{\mu}M$), ${\bullet}O_2{^-}$ radical ($IC_{50}$ $228.40{\pm}9.20{\mu}M$), $DPPH{\bullet}$radical ($IC_{50}$ $27.35{\pm}1.44{\mu}M$), and $ABTS{^+}{\bullet}$radical ($IC_{50}$ $2.53{\pm}0.070{\mu}M$), and reduce $Cu^{2+}$ ion ($IC_{50}$ $11.97{\pm}0.68{\mu}M$). In order to investigate the possible mechanism, the reaction product of [6]-gingerol and $DPPH{\bullet}$ radical was further measured using HPLC combined mass spectrometry. The product showed a molecular ion peak at m/z 316 $[M+Na]^+$, and diagnostic fragment loss (m/z 28) for quinone. On this basis, it can be concluded that: (i) [6]-gingerol can effectively protect against ${\bullet}OH$-induced DNA damage; (ii) a possible mechanism for [6]-gingerol to protect against oxidative damage is ${\bullet}OH$ radical scavenging; (iii) [6]-gingerol scavenges ${\bullet}OH$ radical through hydrogen atom ($H{\bullet}$) transfer (HAT) and sequential electron (e) proton transfer (SEPT) mechanisms; and (iv) both mechanisms make [6]-gingerol be oxidized to semi-quinone or quinone forms.

• Journal of Veterinary Science
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• v.22 no.6
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• pp.78.1-78.10
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• 2021

Background: Recurrent subclinical mastitis (RScM) due to resistant bacteria has low clinical and bacteriological cure rates, often requiring the culling of cows. The sequential intramammary administration of enrofloxacin hydrochloride-dihydrate (enro-C) followed by ceftiofur HCl may be useful for treating these cases. Objectives: This study assessed the bacteriological and clinical cure-efficacies of the sequentially intramammary administration of enro-C, followed by ceftiofur HCl to treat RScM in Holstein/Friesian cows. Methods: This trial was conducted in a herd with a high prevalence of RScM, and 20 Holstein/Friesian cows were included: 45% suffering subclinical mastitis and 38.9% of the mammary quarters affected. Twenty-nine bacterial isolates in vitro resistant to enro-C were obtained (coagulase-negative Staphylococcus spp, 55.2%; Staphylococcus aureus, 27.6%; Escherichia coli, 6.9%; Streptococcus uberis, 6.9%; Corynebacterium bovis, 3.4%). Polymerase chain reaction-isolated the following genes linked to enro-C resistance: chromosomal (gyrA) and plasmid (aac(6')-lb-cr). The treatments were as follows: twice-daily intramammary infusions of enro-C (300 mg/10 mL) for 5 days. Cows clinically considered treatment failures were also treated with intramammary ceftiofur (125 mg/10 mL, twice daily for 5 days. The clinical and bacteriological cure rates were carried out when completing each treatment phase and at 14 and 21 days, aided by a California mastitis test, somatic cell count, and failure to identify the initially causative bacteria. Results: Enro-C achieved 65% clinical and bacteriological cure rates, and 100% cure rates were obtained after the rescue treatment with ceftiofur HCl. Conclusions: Outstanding clinical and bacteriological cure rates in cows affected by RScM were achieved with the consecutive intramammary infusions of enro-C, followed by ceftiofur HCl.

• Journal of the Korean Geotechnical Society
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• v.15 no.5
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• pp.217-228
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• 1999

The three methods of analysis (i) REV(representative elemental volume), (ii) non-REV and (iii) joint network analysis are introduced in this paper to analyze the groundwater flow in jointed rock mass and the inflow into underground excavations. The results from those methods are compared one another to reveal their characteristics by varying the number of joints and the diameter of the opening. The pre-processor, the so-called sequential analysis, is introduced to predict the equivalent hydraulic conductivity of a jointed rock mass having a number of intersecting joints. Using the finite element mesh, joint map and sequential analysis, the equivalent hydraulic conductivities are calculated for all 445 elements. The hydraulic inhomogeneity and the determination of the representative properties of jointed rock masses are discussed. In the REV analysis where the entire rock mass is homogenized through the representative properties, the inflow is increased regularly and consistently by increasing the joint density, the opening size and the conductivity contrast value. Though the non-REV analysis showed irregular variation of the inflow due to the local inhomogeneity allowed to individual elements, the inflow approached the REV results as the characteristic length increases. The joint network analysis showed the most sensitive reaction to the joint density, the opening size and the presence of the network crossing the opening. The reliability of the network analysis depends on the geometric data of individual joints. In view of the limited field data on joint geometry and possible uncertainty the REV and non-REV methods are considered more practical and rational than the joint network analysis.

• Korean Journal of Breeding Science
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• v.40 no.4
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• pp.394-400
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• 2008

Blast resistance of one hundred and thirty-one rice cultivars bred in Korea was tested with thirty Korean isolates and twenty-two Philippines isolates using three screening methods. In the blast nursery conducted in Korea and in the Philippines, average disease index of rice cultivars were 4.6 and 2.2, respectively. Seventy-nine cultivars showed different resistance reaction in Korea and in the Philippines, and 19 cultivars showed the same resistant reaction in two locations. In the seedling test, Korean blast isolates displayed different levels of virulence. 93-093, a Korean isolate, was compatible with 90 cultivars whereas 97-057 showed a compatible reaction with 13 cultivars. Twenty-three cultivars showed high level of resistance against Korean and Philippines isolates but Chucheongbyeo, Heugnambyeo, and Manmibyeo showed susceptible reaction to all blast isolates. Through the sequential planting test in Korea and in the Philippines, Palgongbyeo and Seomjinbyeo displayed durable resistance, and Nagdongbyeo and Gihobyeo showed high level of disease infection over the planting time. These results indicate that blast isolates collected in two countries have different genetic background and number of compatible isolates should be considered in definition the durability of rice cultivar to rice blast.

• Tuberculosis and Respiratory Diseases
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• v.38 no.4
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• pp.357-371
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• 1991

To identify the pathogenetic role of reactive oxygen free radical-induced oxidation reaction in endotoxin-induced acute lung injury, we infused endotoxin into 8 domestic pigs; endotoxin only (n=3), pretreatment with dimethylthiourea (DMTU) (n=5). We observed the sequential changes in hemodynamic parameters, the concentration of plasma oxidized glutathione (GSSG) in pulmonary arterial and venous blood, and albumin content in bronchoalveolar lavage fluid (BALF). The results were as follows. 1) While cardiac output decreased, mean pulmonary arterial pressure, pulmonary vascular resistance, and alveolar-arterial oxygen difference increased over phase 1 (0-2 hr) and phase 2 (2-4.5 hr) by endotoxin, DMTU attenuated the above changes only during phase 2. 2) While the concentration of plasma GSSG increased significantly by endotoxin during phase 2, there were no significant differences between pulmonary arterial and venous GSSG contents during both phases. The increase in plasma GSSG content was attenuated by DMTU. 3) The content of BALF albumin was significantly lower in DMTU group than that of endotoxin group. These results suggest that reactive oxygen free radical-induced oxidation reaction may have an important pathogenetic role in endotoxin-induced acute lung injury in pigs, which seems to be greater during phase 2 rather than phase 1.

• Journal of Microbiology and Biotechnology
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• v.16 no.3
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• pp.355-359
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• 2006

The functional stability of mRNA is one of the crucial factors affecting the efficiency of in vitro translation. As the rapid degradation of mRNA in the cell extract (S30 extract) causes early termination of the translational reactions, extending the mRNA half-life will improve the productivity of the in vitro protein synthesis. Thus, a simple PCR-based method is introduced to increase the stability of mRNA in an S30 extract. The target genes are PCR-amplified with primers designed to make the ends of the transcribed mRNA molecule anneal to each other. When compared with normal mRNA, the mRNA with the annealing sequences resulted in an approximately 2-fold increase of protein synthesis in an in vitro translation reaction. In addition, sequential transcription and translation reactions in a single tube enabled direct protein expression from the PCR-amplified genes without any separate purification of the mRNA.