• Journal of Life Science
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• v.19 no.7
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• pp.845-851
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• 2009

A vector harboring double cassettes; a heterologous gene expression cassette of pHCE-InaN-antigen and a ghost formation cassette of pAPR-cI-E lysis 37 SDM was constructed and introduced to E. coli DH5a. For the production of a bacterial ghost vaccine, bacterial ghosts from E. coli / Streptococcus iniae with four different types of antigens - enolase, GAPDH, sagA and piaA - were produced by the optimization of fermentation parameters such as a glucose concentration of 1 g/l, agitation of 300 rpm and aeration of 1 vvm. Efficiency of ghost bacteria formation was evaluated with cultures of OD$_{600}$=1.0, 2.0 and 3.0. The efficiency of the ghost bacteria formation was 99.54, 99.67, 99.99 and 99.99% with inductions at OD$_{600}$=3.0, 1.0, 2.0 and 1.0 for E. coli/S. iniae antigens enolase, piaA, GAPDH and sagA, respectively. Ghost bacteria as a vaccine was harvested by centrifugation. The antigen protein expressions were analyzed by SDS-PAGE and western blot analysis, and the molecular weights of the enolase, piaA, GAPDH and sagA were 78, 26, 67 and 26 kDa, respectively. The molecular weights of the expressed antigens were consistent with theoretical sizes obtained from the amino acid sequences.

• The Plant Pathology Journal
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• v.26 no.4
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• pp.386-393
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• 2010

In order to study disease resistance mechanisms in rice against the rice blast fungus Magnaporthe grisea, we screened fungal elicitor-responsive genes from rice suspension-cultured cells treated with fungal elicitors employing differential hybridization (DH). By DH screening, 31 distinct rice clones were isolated and a majority of them were full-length cDNAs encoding pathogenesisrelated (PR) genes. Sixteen of the 31 genes were upregulated at 4, 8, and 12 h following fungal elicitor treatment. To elucidate the effect of signal molecules and biotic elicitors on the regulation of rice defense genes, we further characterized the transcriptional expression patterns of representative isolated PR genes; OsGlu1, OsGlu2, OsTLP, OsRLK, and OsPR-10, following treatment with fungal elicitor, phytohormones, cycloheximide, and inhibitors of protein phosphorylation. Jasmonic acid (JA) induced transcriptional expression of OsGlu1, OsTLP, and OsRLK, but not of OsGlu2 and OsPR-10 at any of the tested time points. Salicylic acid (SA) and abscisic acid weakly induced the expression of OsTLP and OsRLK. SA showed an antagonistic effect with fungal elicitor and JA. Cycloheximide suppressed all these genes upon elicitor treatment, except for OsGlu2. Staurosporine only induced the expression of OsRLK. Application of calyculin A strongly induced OsRLK expression, but suppressed the expression of OsGlu2. Our study yielded a number of PR genes that play a role in defense mechanisms against the rice blast fungus, as well as contribute towards the elucidation of crosstalk between phytohormones and other modifications during defense signaling.

• Biomedical Science Letters
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• v.2 no.2
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• pp.175-185
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• 1996

The mitogen-activated protein(MAP) kinase signal transduction pathway represents an important mechanism by which mitogen, such as serum and PMA, regulate cell proliferation and differentiation. Target substrates of the MAP kinase are located within several compartments containing plasma membranes and nucleus. We now report that serum addition induces proliferation of the P388 murine leukemia cell, but PMA does not, while both serum and PMA treatment cause translocation of the MAP kinase, mainly p42$^{mapk}$ isoform, from cytosol into the nucleus, which was monitored by immunoblot analysis using polyclonal anti-ERK1 antibodies. We investigated whether the MAP kinase was capable of phosphorylating c-Jun protein and GST-fusion proteins, the P562$^{kk}$N-terminal peptides (1-77 or 1-123 domain) of the T cell tyrosine kinase, using the partially purified MAP kinase by SP-sephadex C-50, phenyl superose and Mono Q column chromatography. We found that the partially purified MAP kinase was able to phosphorylate c-Jun protein and the GST-fusion protein expressed using E.coli DH5$\alpha$ which is transformed with pGEX-3Xb plasmid vector carrying of p562$^{kk}$N-terminal peptide-encoding DNA. These results imply that tyrosine kinase receptor/Ras/Raf/MAP kinase pathway is a major mechanism for mitogen-induced cell proliferation in P388 murine leukemia cell and that the various MAP kinase isoforms may have their own target substrates located in distinct subcellular compartments.

• Journal of the Korean Applied Science and Technology
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• v.36 no.2
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• pp.676-684
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• 2019

The Pictet-Spengler reactions have widely known for the organic synthesis or biosynthesis of biologically active compounds, tetrahydro-${\beta}$-carbolines. We have developed the simple and efficient synthetic method for the synthesis of ${\beta}$-carbolines in water. Their chemical structures were characterized by nmr and UPLC/MS/QTOF. Calculated masses of compound 1 ($C_{17}H_{17}N_2$ 249.1392), 2 ($C_{17}H_{23}N_2$ 255.1861), 3 ($C_{19}H_{21}N_2O_3$ 325.1552) and 4 ($C_{19}H_{19}N_2O$ 279.1497) were almost identical with the detected masses of compound 1 (249.1315), 2 (255.1789), 3 (325.1460) and 4 (279.1364) respectively. Those synthesized four compounds showed strong antibiotic activity against the common E. coli.

• Korean Journal of Poultry Science
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• v.24 no.3
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• pp.145-151
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• 1997

This study was conducted to investigate the influence of dietary olive oil, canola oil, or sardine oil on the fatty acids composition and cholesterol contents in eggs The experimental layers, 62~65 weeks of age, were randomly allotted to one of the four treatments; a) Control (commercial feed), b) T1 (commercial feed plus 10% olive oil), c) T2 (commercial feed plus 10% sardine oil), d) m (commercial feed plus 10% sardine oil) for 3 weeks. The egg samples were stored at 4${\pm}$1˚C for a certain period (1, 3, 5, 7, and 9 days) and analyzed for the fatty acids composition and cholesterol contents. The unsaturated fatty acid contents in the egg yolk were slightly decreased as the storage periods extended. The oleic acid content in T1 was the highest in all treatments. The linoleic and linolenic acid contents in T2 were higher than in the other treatments. The eicosahexaenoic acid (EPA), docosapentaenoic acid (DPA), and docosasahexaenoic acid ( DH A) contents in eggs from m were higher than the other treatments. The ratio of egg yolk n-6 /n-3 fatty acids contents in all treatments were slightly increased as the storage periods extended. The n-3 fatty acid contents of the egg yolk from T3 was the highest in all treatments. The cholesterol contents of the eggs were significantly decreased as the storage period extended(P<0.05). There was no significant difference in egg cholesterol content among all treatments.

• Korean Journal of Microbiology
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• v.32 no.3
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• pp.186-191
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• 1994

The origin of the leading strand replication (ori) and of lagging strand replication (M-O) of R-plasmid pSBK203 was identified and its base sequence was determined. About 50 bp of ori sequence residues overlapped with the structural gene of rep. Sequence comparison reveals that pSBK-ori shares obvious identities with those of pT181 family and consists of two regions, one is conserved and the other is variable region. Of two palindrome sequence located one after another in upstream region of rep gene, palA' instead of palA which shares sequence homology with diverse family of plasmids such as pOX6, pC194, and pE194 seems to act as a signal for conversion of primarily replicated ssDNA to dsDNA (minus origin (M-O)).

• Journal of Ocean Engineering and Technology
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• v.25 no.3
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• pp.73-84
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• 2011

This is the fifth in a series of companion papers dealing with the dynamic hardening properties of various marine structural steels at intermediate strain rates. Five steps of strain rate levels (0.001, 1, 10, 100, 200/s) and three steps of temperature levels (LT ($-40^{\circ}C$), RT, and HT ($200^{\circ}C$)) were taken into account for the dynamic tensile tests of three types of marine structural steels: API 2W50 and Classifications EH36 and DH36. The total number of specimens was 180 pieces. It was seen that the effects of dynamic hardening became clearer at LT than at RT. Dynamic strain aging accompanying serrated flow stress curves was also observed from high temperature tests for all kinds of steels. The dynamic hardening factors (DHFs) at the two temperature levels of LT and RT were derived at the three plastic strain levels of 0.05, 0.10, 0.15 from dynamic tensile tests. Meanwhile, no DHFs were found for the high temperature tests because a slight negative strain rate dependency due to dynamic strain aging had occurred. A new formulation to determine material constant D in a Cowper-Symonds constitutive equation is provided as a function of the plastic strain rate, as well as the plastic strain level. The proposed formula is verified by comparing with test flow stress curves, not only at intermediate strain rate ranges but also at high strain rate ranges.

• Journal of Microbiology and Biotechnology
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• v.19 no.3
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• pp.257-264
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• 2009

A $\beta$-agarase gene, agaB34, was functionally cloned from the genomic DNA of a marine bacterium, Agarivorans albus YKW-34. The open reading frame of agaB34 consisted of 1,362 bp encoding 453 amino acids. The deduced amino acid sequence, consisting of a typical N-terminal signal peptide followed by a catalytic domain of glycoside hydrolase family 16 (GH-16) and a carbohydrate-binding module (CBM), showed 37-86% identity to those of agarases belonging to family GH-16. The recombinant enzyme (rAgaB34) with a molecular mass of 49 kDa was produced extracellularly using Escherichia coli $DH5{\alpha}$ as a host. The purified rAgaB34 was a $\beta$-agarase yielding neoagarotetraose (NA4) as the main product. It acted on neoagarohexaose to produce NA4 and neoagarobiose, but it could not further degrade NA4. The maximal activity of rAgaB34 was observed at $30^{\circ}C$ and pH 7.0. It was stable over pH 5.0-9.0 and at temperatures up to $50^{\circ}C$. Its specific activity and $k_{cat}/K_m$ value for agarose were 242 U/mg and $1.7{\times}10^6/sM$, respectively. The activity of rAgaB34 was not affected by metal ions commonly existing in seawater. It was resistant to chelating reagents (EDTA, EGTA), reducing reagents (DTT, $\beta$-mercaptoethanol), and denaturing reagents (SDS and urea). The E. coli cell harboring the pUC18-derived agarase expression vector was able to efficiently excrete agarase into the culture medium. Hence, this expression system might be used to express secretory proteins.

• Toxicological Research
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• v.18 no.3
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• pp.249-257
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• 2002

Cytochrome P450 3As such as 3A4 and 3A5 metabolize a wide range of pharmaceutical compounds. The vectors for the expression of fusion protein containing an N-terminal human P450 3A4 or P450 3A5 sequences and a C-terminal rat NADPH-cytochrome P450 reductase moiety were constructed. These plasmids were used to express the fusion protein in Escherichia coli DH5$\alpha$ cells. High levels of expression were achieved (100~200 nmol/liter) and the expressed fusion protein in E. coli membranes were catalytically active for nifedipine oxidation, a typical enzymatic activity of P450 3A4. The NADPH-P450 reductase activities of these fusion protein were also determined by measuring reduction of cytochrome c. To fine a specific Inhibitor of P450 3A4 from naturally occurring chemicals, a series of isothiocyanate compounds were evaluated for the inhibitory activity of P450 using the fusion proteins in E. coli membranes. Of the five isothiocyanates (phenethyl isothiocyanate, phenyl isothiocyanate, benzol isothiocyanate, benzoyl isothiocyanate and cyclohexyl isothiocyanate) tested, benzoyl isothiocyanate showed a strong inhibition of P450 3A4 with an $IC_{50}$value of 2.8 $\mu\textrm{M}$. Our results indicate that the self-sufficient fusion protein will be very useful tool to study the drug metabolism and benzyl isothiocyanate may be valuable for characterizing the enzymatic properties of P450 3A4.

• Food Science of Animal Resources
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• v.23 no.1
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• pp.63-69
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• 2003

This experiment was carried out to study changes in solubility and emulsifying properties of whey protein. Whey protein hydrolysates were obtained from tryptic hydrolysis of whey protein concentrate at pH 8.0 and 37$^{\circ}C$ for 6 hours. Emulsifying activity of whey protein hydrolysate was highest at 4 hours of hydroysis and at 5.50% of DH. During hydrolysis of whey protein concentrate with trypsin, ${\alpha}$-lactalbumin was not easily broken down. But ${\beta}$-lactoglobulin was hydrolysed rapidly from the early stage of hydrolysis, producing several low molecular weight peptides, which have to participate in increasing emusifying activity. The solulbility of hydyolysates tended to increase depending on hydrolysis time; however, there was a gradual decrease after 5 hours. The hydrolysate had a minimum solubility near the isoelectric point range (pH 4∼5). The more hydrolysed the whey protein concentrates, the more soluble they are near the pl. They aye also more soluble above pH 6. Emulsifying activity of hydrolysates showed similar results to solubility. Creaming stability gradually increased when hydrolysis increased, increasing rapidly above pH 8 after 4 hours of hydrolysis.