• Preventive Nutrition and Food Science
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• v.8 no.2
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• pp.113-118
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• 2003

We recently extracted lignans such as matairesinol and 2-hydroxyarctigenin from safflower seeds and found that they exhibit a potent cytotoxic effect on human promyleocytic leukemia HL-60 cells. In this study, we investigated whether mechanisms of the matairesinol-induced cell death are associated with the programmed cell death, apoptosis. Matairesinol dose-dependently reduced viability of HL-60 cells with an IC/sun 50/ value of 60 $\mu$M. Staining of cells with Hoechst 33342 revealed distinct morphological features of apoptosis, such as the nuclei broken into chromatin containing fragments of various sizes in the cells exposed to 100 $\mu$M matairesinol for 24 hr. Agarose gel electrophoresis of DNA from the cells treated with matairesinol showed internucleosomal DNA degradation into oligonucleosomal sizes. DNA ladder like patterns were easily detected after treatment with matairesinol concentrations ranging from 10 to 100 $\mu$M after 24 hr. In cells treated with 100 $\mu$M matairesinol for differing time periods, the DNA ladder was detectable from 6 hr onward. A time course histogram of the DNA content analyzed by flow cytometry revealed a rapid increase in subdiploid cells and a concomitant decrease in diploid cells exposed to 100 $\mu$M matairesinol. These results indicate that matairesinol-induced HL-60 cell death was due to the DNA damage and apoptosis.

• Journal of Ginseng Research
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• v.36 no.4
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• pp.418-424
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• 2012

This study focused on the enzymatic biotransformation of the major ginsenoside Rb1 into Rd for the mass production of minor ginsenosides using a novel recombinant ${\beta}$-glucosidase from Flavobacterium johnsoniae. The gene (bglF3) consisting of 2,235 bp (744 amino acid residues) was cloned and the recombinant enzyme overexpressed in Escherichia coli BL21(DE3) was characterized. This enzyme could transform ginsenoside Rb1 and gypenoside XVII to the ginsenosides Rd and F2, respectively. The glutathione S-transferase (GST) fused BglF3 was purified with GST-bind agarose resin and characterized. The kinetic parameters for ${\beta}$-glucosidase had apparent $K_m$ values of $0.91{\pm}0.02$ and $2.84{\pm}0.05$ mM and $V_{max}$ values of $5.75{\pm}0.12$ and $0.71{\pm}0.01{\mu}mol{\cdot}min^{-1}{\cdot}mg$ of $protein^{-1}$ against p-nitrophenyl-${\beta}$-D-glucopyranoside and Rb1, respectively. At optimal conditions of pH 6.0 and $37^{\circ}C$, BglF3 could only hydrolyze the outer glucose moiety of ginsenoside Rb1 and gypenoside XVII at the C-20 position of aglycon into ginsenosides Rd and F2, respectively. These results indicate that the recombinant BglF3 could be useful for the mass production of ginsenosides Rd and F2 in the pharmaceutical or cosmetic industry.

• Proceedings of the Botanical Society of Korea Conference
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• 1996.06a
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• pp.68-78
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• 1996

Two types of immunoloigcal probes, anti-ABBP Abs, have been developed. The purified ABBP from ABA-C1-BSA-sepharose 4B column was identified by PAGE and appeared in one band of about 56KD, as well as showed a specific binding ability and a high affinity for ABA (Kd2.0$\times$10-9 mol/L). Unexpectedly, the existence of rRNA with a length of around 300 nucleotides could be found, when the ABBP was digested with proteinase K and identified by eletrophorsis on an agarose gel (1%). As a result, about 120 cDNA clones coding maize 17s RNA and only one cDNA clone coding ABBP (24cDNA) were obtained from 200,000 seperated phage plaques by the anti-ABBP pAbs. 24cDNA had 1075bp and contained an open reading frame coding 254 amino acids. The anti-idiotypic Ab raised against an ABA MAb showed the ability of either mimicking ABA or competing with ABA. The localization of ABBPs in plant cell was investigated.

• YAKHAK HOEJI
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• v.47 no.6
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• pp.410-416
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• 2003

Autotaxin(ATX) was originally purified from conditioned media of A2058 human melanoma cells and shown to be a potent cell motility-stimulating factor, possessing a type II nucleotide pyrophosphatase/phosphodiesterase (NPP2) activity. Recombinant ATX has recently demonstrated that human plasma lysophosholipase D is identical to ATX and uses lysophosphatidylcholine as a substrate to mediate various biological functions including tumor cell growth and motility through G-protein coupled receptor. However, despite pivotal roles of ATX on physiological or pathophysiological states, the production of ATX is solely depends on complicated purification method which employs multiple column steps, but resulted in very poor yield. This limited the use of ATX for extensive analysis. We, therefore, expressed six histidine-tagged recombinant human ATX(His-ATX) in High Five TM insect cells to improve the generation of ATX and to make simple the purification of ATX. The signal sequence of the human ATX gene was truncated and replaced with sequence of insect cell secretion signal within expression vector. In addition, codons for six histidines were added to the C-termini of 120kDa ATX cDNA construct. A simple purification scheme utilizing two-step affinity column chromatography was designed to purify His-ATX to homogeneity from the culture supernatant of transfected insect cells. Homogenous His-ATX was detected and isolated from the concentrated insect cell medium using concanavalin A agarose and nickel affinity chromatography. Purified His-ATX was in full length with ATX capacity. A combination of this expression system and purification scheme would be useful for production and purification of high-quality functional ATX for research and practical application of multiple functional motogen, ATX/NPP-2.

• Korean Journal of Food Science and Technology
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• v.21 no.5
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• pp.606-612
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• 1989

A New lectin from baby clam, Tapes japonica, was isolated and purified through the following procedures; acetone powder, 0.15M NaCl extraction, ammonium sulfate fractionation, N-acetyl-D-galactosamine-agarose affinity column, and ion exchange Mono Q of FPLC. This lectin nonspecifically agglutinated human erythrocytes but didn't agglutinate mouse and rabbit erythrocytes. And the lectin neither stimulated human lymphocytes nor agglutinated Sarcoma 180 cells. On polyacrylamide gel electrophoresis, the lectin migrated as a major single band indicating homogeneous. A molecular weight was estimated to be about 131,000 daltons by Biogel P-300 and 125,000 daltons by SDS-PAGE without ${\beta}-mercaptoethanol$. This lectin is supposed to be a tetramer composed of heterogeneous subunits, about 30,000 and 33,000 daltons. Baby clam lectin was inhibited by EDTA and recovered agglutinating activity by $Ca^{++}\;and\;Mn^{++}$. This lectin is revealed as glycoprotein that contained about 4.2% neutral sugar.

• Journal of the Korean Society of Food Culture
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• v.19 no.2
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• pp.129-138
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• 2004

The changes in DNA damage were investigated during storage after irradiation. Beef, pork and chicken were irradiated at 1.0, 3.0 and 5.0 kGy and stored for 6 months at $-20^{\circ}C$. The comet assay was applied to the sample muscles at the beginning of irradiation and at the end of storage. Muscles were isolated, sliced, and the suspended cells were embedded in an agarose layer. After lysis of the cells, they were electrophoresed for 2 min. and then stained. DNA fragmentation in tissues caused by irradiation was quantified as tail length and tail moment (tail length ${\times}$ % DNA in tail) by comet image analyzing system. Right after irradiation, the differences in tail length between unirradiated and irradiated muscles were significant(p<0.05) in beef, pork and chicken. With increasing the increasing doses, statistically significant longer extension of the DNA from the nucleus toward anode was observed. Similarly even 6 months after irradiation, all the irradiated muscles significantly showed longer tail length than the unirradiated controls. The results represented as tail moment showed similar tendency to those of tail length, but the latter parameter was more sensitive than the former. These results indicate that the comet assay could be one of the simple methods of detecting irradiated muscles. Moreover, this method suggest that using comet assay, we were able to detect DNA damage differences even after 6 months after irradiation.

• Korean Journal of Microbiology
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• v.25 no.1
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• pp.9-16
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• 1987

Three strains of bacteria utilizing salicylate, KU801(pKU5, pKU8), KU803(pKU6, pKU9), and KU806(pKU7, pKU10), were selected from the isolates and identified as Pseudomonas putida. By agarose gel electrophoresis, it was found that the strains had two plasmids each. All three strains were resistant to antibiotics such as ampicillin, tetracyclin, and chloramphenicol, and did not utilize other aromatic and aliphatic hydrocarbons examined except salicylate. The plasmids (pKU5, pKU6, and pKU7) of larger molecular weight were cured by treatment with mitomycin C and frequencies of curing were 0.4%, 1.67%, and 0.75%, respectively. Cured strains did not degrade salicylate and still had antibiotic resistances, which were identical with wild strains. The genes for salicylate degradation were proved to be enclded on thier plasmids. The molecular weights of pKU5 and pKU6 were estimated as 103.5Md, and that of pKU 7 as 101 Md. The new SAL plasmids, pKU5, pKU6, and pKU7 were transferred to P. putida and P. aeruginosa, but not to E. coli.

• The Korean Journal of Ecology
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• v.11 no.1
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• pp.29-36
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• 1988

This present study has been carried out to examine the correlation between the distribution of the coliform bacterial plasmids and the viability test against heavy metals in the upper stream of Han river(Gapyung; clean water) and the its basin (Anyang Cheon; polluted water). And the distribution of plasmids were examined to be used as trophic indicator for analysis between the clean and polluted waters. 1. A total of 110 isolates were analyzed for the presence of plasmids by means of the boiling method and agarose gel techniques. Plasmids were significantly more frequent in the strains which had been isolated from the clean water (14.3%). Also, there were much higher multiplicity of plasmids at the polluted water(41.0%), compared with the clean water (33.0%). By the comparision between molecular weight of bacterial plasmids in the clean water and those of polluted water, there were no significant differences of the clean water from the polluted water, to such extent as 30.0%, 28.6%, respectively, in frequency for occurrence of high molecular weight plasmids iver 35.8% Mdal. 2. Each isolate was carried out the resistance test for mercury(10-5M), nickel(10-3M) and arsenic (0.1%). At the polluted site, the survival ability of the plasmid-carrying straings(Hg, 31.0%l Ni, 5.7%; As, 65.7%) was higher than that of the non plasmid-carrying strains(Hg, 12.1%; Ni, 3.0%; As, 54.6%). This trend was more remarkable in the clean site (plasmid-carrying strains: 16.7%,-,-). As a result it is suggested that plasmids could be used as an indicator of a certain types of water pollution. In addition, heavy metals might have inflyenced, some extent, to the distribution of plasmids in the environment which has been surveyed in the present study.

• Journal of Pharmacopuncture
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• v.10 no.2 s.23
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• pp.73-79
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• 2007

Objectives : Tumor necrosis factor-${\alpha}{\cdot}$(TNF-${\alpha}{\cdot}$) is a proinflammatory cytokine involved in the pathogenesis of rheumatoid arthritis. This study was designed to investigate the relation between TNF-${\alpha}{\cdot}$ gene polymorphism and rheumatoid arthritis in Korean population. Methods : This study was carried out on 103 rheumatoid arthritis patients who fulfilled the American College of Rheumatology 1987 revised criteria for rheumatoid arthritis and 208 healthy control subjects. Blood samples from all subjects were obtained for DNA extraction. The extracted DNA was amplified by polymerse chain reaction(PCR). PCR products were visualized by 2% agarose gel electrophoresis. We investigated the genotyping of TNF-${\alpha}{\cdot}$ by using Pyrosequencing. Results : The genotypes of TNF-${\alpha}{\cdot}$ gene were GG, AG and AA. While the distribution of TNF-${\alpha}{\cdot}$ polymorphism in control subjects was 92.31%, 7.21%, 0.48% respectively, in rheumatoid arthritis patients was 93.20%, 6.80%, 0.00%(GG, AG. AA). There was no statistical significant allelic frequency difference between control and rheumatoid arthritis groups. Conclusion : We concluded that there was no significant association between TNF-${\alpha}{\cdot}$ gene polymorphism and rheumatoid arthritis. However, the findings of this study need to be confirmed in more patients and further studies.

• Journal of Microbiology and Biotechnology
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• v.21 no.11
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• pp.1116-1122
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• 2011

In this study, site-directed mutagenesis was performed on the ${\beta}$-agarase AgaA gene from Zobellia galactanivorans to improve its catalytic activity and thermostability. The activities of three mutant enzymes, S63K, C253I, and S63K-C253I, were 126% (1,757.78 U/mg), 2.4% (33.47 U/mg), and 0.57% (8.01 U/mg), respectively, relative to the wild-type ${\beta}$-agarase AgaA (1,392.61 U/mg) at $40^{\circ}C$. The stability of the mutant S63K enzyme was 125% of the wild-type up to $45^{\circ}C$, where agar is in a sol state. The mutant S63K enzyme produced 166%, 257%, and 220% more neoagarohexaose, and 230%, 427%, and 350% more neoagarotetraose than the wild-type in sol, gel, and nonmelted powder agar, respectively, at $45^{\circ}C$ over 24 h. The mutant S63K enzyme produced 50% more neoagarooligosaccharides from agar than the wild-type ${\beta}$-agarase AgaA from agarose under the same conditions. Thus, mutant S63K ${\beta}$-agarase AgaA may be useful for the production of functional neoagarooligosaccharides.