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Enzymatic characterization of Paenibacillus amylolyticus xylanases GH10 and GH30 for xylan hydrolysis (Paenibacillus amylolyticus 유래 xylanase GH10 및 GH30의 xylan 가수분해 특성)

  • Nam, Gyeong-Hwa;Jang, Myoung-Uoon;Kim, Min-Jeong;Lee, Jung-Min;Lee, Min-Jae;Kim, Tae-Jip
    • Korean Journal of Microbiology
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    • v.52 no.4
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    • pp.463-470
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    • 2016
  • The enzymatic degradation of xylans is the most versatile way to obtain the high value-added functional compounds or the fermentable sugars for renewable energy. The endo-${\beta}$-xylanases are the major enzymes which hydrolyze the internal ${\beta}$-1,4-linkages of xylan backbones to produce the mixtures of xylooligosaccharides including xylobiose and xylotriose. Among them, glucuronoxylanase GH30 can exclusively hydrolyze the internal ${\beta}$-1,4-linkages of xylans decorated with methylglucuronic acid branches. In the present study, two xylanolytic enzyme (PaXN_10 and PaGuXN_30) genes were cloned from Paenibacillus amylolyticus KCTC 3005, and expressed in Escherichia coli, respectively. PaXN_10 (38.7 kDa) belongs to the endo-${\beta}$-xylanases GH10 family, while PaGuXN_30 (58.5 kDa) is a member of glucuronoxylanase GH30. They share the same optimal reaction conditions at $50^{\circ}C$ and pH 7.0. Enzymatic characterization proposed that P. amylolyticus can utilize the hardwood glucuronoarabinoxylans via the cooperative actions of xylanases GH10 and GH30. The extracellular PaGuXN_30 is secreted into the medium and hydrolyzes glucuronoarabinoxylans to release a series of aldouronic acid mixtures with a methylglucuronic acid branch. The resultant products being transported into the microbial cell are successively degraded into the smaller xylooligosaccharides by the intracellular PaXN_10, which will be utilized for the cellular metabolism.

Consumption and Preference for Bokbunja (Rubus coreanus Miquel) Products (복분자 제품에 대한 기호도 및 이용실태 조사)

  • Lee, Jeong-Ae;Park, Geum-Soon
    • Korean journal of food and cookery science
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    • v.27 no.3
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    • pp.11-20
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    • 2011
  • The purpose of this study was to investigate the use and perception of adults concerning bokbunja (Rubus coreanus Miquel) products. Self-administered questionnaires were collected from 502 residents in the Daegu and Gyeongbuk area. According to the survey more females in their 20s responded than males (45.2 and 54.8%, respectively). In total, 82.6% of respondents had an income of under 4,000,000 won per month and most were living in the city. The results regarding knowledge about bokbunja products showed that the bokbunja wine score was highest (3.66), and intake frequency showed that over half of the respondents had never eaten bokbunja products. The preference for bokbunja wine by males was higher than that for females, whereas the others products showed higher scores for females than for males. When asked about how they knew about bokbunja products, 37.0% of males and 46.9% of females responded a family member or neighbor. When purchasing bokbunja products, males and females answered that they considered taste, country of origin, and manufacturer, and price. Approximately 48% of the respondents answered that they purchased bokbunja products in the grocery and department store. As a result, popularization of bokbunja products was based on product development and sales promotion, and the product development factor significantly influenced preference for bokbunja products.

Antioxidant, Antiaging and Antimicrobial Effects of Ethanolic Extract and Ethyl Acetate Fraction from Eclipta prostrata (한련초 에탄올 추출물과 에틸아세테이트 분획물의 항산화, 항노화 및 항균 효과)

  • Lee, Sang Lae;Song, Ba Reum;Shin, Hyuk Soo;Lee, Yun Ju;Park, Soo Nam
    • Journal of the Society of Cosmetic Scientists of Korea
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    • v.44 no.3
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    • pp.349-362
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    • 2018
  • An annual plant, Eclipta prostrata (Linn) is a member of the Asteraceae plant family and inhabited in tropical or subtropical regions of the world. Through many previous researches, E. prostrata has been extensively studied for its hepatoprotective effect, antivenom potential against viper venom, antioxidant, hair-growth, wound-healing efficacy and so on. In this study, for better understanding of the potential of E. prostrata as skin protectant, we conducted the experiments evaluating the antioxidant and antiaging efficacy. To this end, 50% ethanolic extract of E. prostrata and its ethyl acetate fraction were prepared and investigated. For the evaluation of antioxidant capacity of the samples, $FSC_{50}$ and $OSC_{50}$ were estimated. As a result, $OSC_{50}$ of ethyl acetate fraction was 2.7 times superior to $OSC_{50}$ of L-ascorbic acid, a well known antioxidant agent. Futhermore E. prostrata showed notable reactive oxygen species (ROS) scavenging effect and protective effect against $H_2O_2$ in the celluar level as well. Especially, in the $^1O_2$ induced hemolysis test, $64{\mu}g/mL$ of ethyl acetate fraction showed greater than 6 times increased retardation effect compare to control which means E. prostrata has remarkable antioxidant capacity. To validate the antiaging effect of the samples, we conducted elastase inhibition assay using elastase solution extracted from human skin fibroblasts, Hs68. As a result, $16{\mu}g/mL$ of each sample showed 6.8% and 14.0% of elastase inhibition respectively. Finally, antimicrobial activity of E. prostrata was assessed to validate the possibility as alternative preservative. From the result, ethyl acetate fraction showed oustanding antimicrobial activity as of methyl paraben, a well known chemical preservative. In conclusion, these results suggest that E. prostrata can be used as natural skin protectant or preservative as natural ingredient in food or cosmetics industry.

The Proteasome Inhibitor MG132 Sensitizes Lung Cancer Cells to TRAIL-induced Apoptosis by Inhibiting NF-κ Activation (폐암세포주에서 NFκ 활성 억제를 통한 Proteasome 억제제 MG132의 TRAIL-유도성 Apoptosis 감작 효과)

  • Seo, Pil Won;Lee, Kye Young
    • Tuberculosis and Respiratory Diseases
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    • v.65 no.6
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    • pp.476-486
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    • 2008
  • Background: TRAIL (TNF-related apoptosis inducing ligand) is a newly identified member of the TNF gene family which appears to have tumor-selective cytotoxicity due to the distinct decoy receptor system. TRAIL has direct access to caspase machinery and induces apoptosis regardless of p53 phenotype. Therefore, TRAIL has a therapeutic potential in lung cancer which frequently harbors p53 mutation in more than 50% of cases. However, it was shown that TRAIL also could activates $NF-{\kappa}B$ in some cell lines which might inhibit TRAIL-induced apoptosis. This study was designed to investigate whether TRAIL can activate $NF-{\kappa}B$ in lung cancer cell lines relatively resistant to TRAIL-induced apoptosis and inhibition of $NF-{\kappa}B$ activation using proteasome inhibitor MG132 which blocks $I{\kappa}B{\alpha}$ degradation can sensitize lung cancer cells to TRAIL-induced apoptosis. Methods: A549 (wt p53) and NCI-H1299 (null p53) lung cancer cells were used and cell viability test was done by MTT assay. Apoptosis was confirmed with Annexin V assay followed by FACS analysis. To study $NF-{\kappa}B$-dependent transcriptional activation, a luciferase reporter gene assay was used after making A549 and NCI-H1299 cells stably transfected with IgG ${\kappa}-NF-{\kappa}B$ luciferase construct. To investigate DNA binding of $NF-{\kappa}B$ activated by TRAIL, electromobility shift assay was used and supershift assay was done using anti-p65 antibody. Western blot was done for the study of $I{\kappa}B{\alpha}$ degradation. Results: A549 and NCI-H1299 cells were relatively resistant to TRAIL-induced apoptosis showing only 20~30% cell death even at the concentration 100 ng/ml, but MG132 ($3{\mu}M$) pre-treatment 1 hour prior to TRAIL addition greatly increased cell death more than 80%. Luciferase assay showed TRAIL-induced $NF-{\kappa}B$ transcriptional activity in both cell lines. Electromobility shift assay demonstrated DNA binding complex of $NF-{\kappa}B$ activated by TRAIL and supershift with p65 antibody. $I{\kappa}B{\alpha}$ degradation was proven by western blot. MG132 completely blocked both TRAIL-induced $NF-{\kappa}B$ dependent luciferase activity and DNA binding of $NF-{\kappa}B$. Conclusion: This results suggest that inhibition of $NF-{\kappa}B$ can be a potentially useful strategy to enhance TRAIL-induced tumor cell killing in lung cancer.

Effects of Myelophycus Simplex Papenfuss Methanol Extract on Adipocyte Differentiation and Adipogenesis in 3T3-L1 Preadipocytes (바위수염 메탄올 추출물이 3T3-L1 지방전구세포의 분화에 미치는 영향)

  • Kim, Hyang Suk;Kwon, Da Hye;Cheon, Ji Min;Choi, Eun Ok;Kim, Ji Hyun;Han, Min Ho;Choi, Yung Hyun;Kim, Byung Woo;Hwang, Hye Jin
    • Journal of Life Science
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    • v.25 no.1
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    • pp.62-67
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    • 2015
  • Myelophycus simplex Papenfuss is distributed over the northern Pacific and southern coast of Korea, and is a member of the brown algae family. The objective of this study was to investigate the effect of M. simplex methanol extract on adipocyte differentiation and adipogenesis in 3T3-L1 preadipocytes. Treatment with M. simplex methanol extract significantly suppressed terminal differentiation of 3T3-L1 preadipocytes in a dose-dependent manner, as confirmed by a decrease in lipid droplet content observed by Oil Red O staining. Also, the M. simplex methanol extract significantly suppressed the triglyceride content of 3T3-L1 preadipocytes in a dose-dependent manner. Treatment with 300 and $500{\mu}g/ml$ of M. simplex methanol extract caused a 42% and 76% reduction in lipid droplet content, respectively. In order to understand the anti-adipogenic effects of M. simplex methanol extract, the changes in the expression of several adipogenic transcription factors, including peroxisome proliferator-activated receptor (PPAR) ${\gamma}$-cytidine-cytidine-adenosine-adenosine-thymidine (CCAAT)/enhancer binding protein (C/EBP) ${\alpha}$ and ${\beta}$, were investigated using immunoblotting. M. simplex suppressed the expression of $PPAR{\gamma}$, $C/EBP{\alpha}$, and $C/EBP{\beta}$ proteins compared with control. Therefore, the results of this study suggest that M. simplex methanol extract inhibits adipocyte differentiation and thus may have applications as a potential source for an anti-obesity functional food agent.

Extracts of Allium fistulosum Attenuates Pro-Inflammatory Action in the Lipopolysaccharide-Stimulated BV2 Microglia Cells (Lipopolysaccharide에 의한 BV2 세포의 염증 반응에 대한 파 추출물의 저해 활성)

  • Park, Shin-Hyoung;Kim, Jung-In;Jeong, Yong-Kee;Choi, Yung-Hyun
    • Journal of Life Science
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    • v.21 no.6
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    • pp.796-804
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    • 2011
  • Microglia are central nervous system (CNS)-resident professional macrophages that function as the principal immune cells responding to pathological stimulations in the CNS. Activation of microglia, induced by various pathogens, protects neurons and maintains homeostasis in the CNS, but severe activation causes inflammatory responses secreting various neurotoxic molecules such as nitric oxide (NO), prostaglandin $E_2$ ($PGE_2$) and pro-inflammatory cytokines. Allium fistulosum, a member of the onion family, is mainly cultivated for consumption, as well as medicinal use in Oriental medicine. It has been reported that A. fistulosum has various biological effects such as anti-oxidant, anti-platelet aggregation, anti-fungus and anti-cholesterol synthesis, however there has been no research about the anti-inflammatory effects of A. fistulosum extracts. In this study, it was undertaken to explore the functions of A. fistulosum as a suppressor of neuronal inflammation by using BV2 microglia cells. As a result, it was found that four kinds of extracts of A. fistulosum effectively reduced the expressions of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2) at both mRNA and protein levels, and also attenuated pro-inflammatory cytokines such as tumor necrosis alpha (TNF-${\alpha}$), interleukin-$1{\beta}$ (IL-$1{\beta}$) and interleukin-6 (IL-6) at the mRNA level in BV2 stimulated by lipopolysaccharide (LPS). In addition, the extracts of A. fistulosum attenuated the release of NO markedly, as well as resulting in slight decreases of TNF-${\alpha}$ and IL-6 production, the effects of which were most significant when treated with ethyl alcohol extract from the whole A. fistulosum. In conclusion, the data indicated that the anti-inflammatory actions of A. fistulosum against BV2 microglia cells is through the down-regulation of iNOS, COX2 and pro-inflammatory cytokines such as TNF-${\alpha}$ and IL-6, and these effects are expected to help in the protection of nerve tissues by suppressions of neuronal inflammation in various neurodegenerative diseases.

Lymphotoxin β Receptor Stimulation Is Linked to MLCK Activity and Suppresses Stress Fiber Formation in Agonistic Anti-LTβR Antibody-stimulated Fibroblastic Reticular Cells (FRC에서 agonistic anti-LTβR antibody의 LTβR 자극은 MLCK 연관성 및 stress fiber 형성에 대한 강력한 억제 작용)

  • Kim, Min Hwan;Lee, Jong-Hwan
    • Journal of Life Science
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    • v.27 no.10
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    • pp.1199-1206
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    • 2017
  • The lymphotoxin ${\beta}$ receptor ($LT{\beta}R$), a member of the tumor necrosis factor receptor family, plays an important role in lymphoid tissue's architecture and organogenesis. We found that $LT{\beta}R$ stimulation induced changes in stress fibers (SFs) in fibroblastic reticular cells (FRCs). MLCK and ROCK play critical roles in the regulation of SF formation in cells. The present study was performed to investigate the antifibrotic effects on SF regulation of $LT{\beta}R$ signaling, with a focus on MLCK inhibition. The effect of $LT{\beta}R$ on the SF change was analyzed using immunoblot and fluorescence assays and agonistic $anti-LT{\beta}R$ antibody-treated FRCs. In addition, we checked the level of Rho-guanosine diphosphate (GDP)/guanosine triphosphate (GTP) exchange activity with FRC lysate. Phospho-ezrin proteins acting as membrane-cytoskeleton linkers completely de-phosphorylated in agonistic $anti-LT{\beta}R$ antibody-treated FRCs. The actin bundles rearranged into SFs, where phospho-myosin light chain (p-MLC) co-localized in FRCs. ML7-treated FRCs completely blocked SFs and showed retraction and shrinkage processes comparable to those observed in agonistic $anti-LT{\beta}R$ antibody-treated cells. Inhibition of ROCK activity induced changes in the actin cytoskeleton organization; however, some SFs remained in the cells, while they were completely disrupted by MLCK inhibition with ML7. We showed that the phosphorylation of MLC was completely abolished with $LT{\beta}R$ stimulation in FRCs. When $LT{\beta}R$ was stimulated with the agonistic $anti-LT{\beta}R$ antibody, the Rho-GDP/GTP exchange activity was reduced, however, the activity was not completely abolished. Collectively, the results illustrated that MLCK was potently responsible for the SF regulation triggered via $LT{\beta}R$ signaling in FRCs.

Development and Evaluation of the Home Economics-Relevant Korean Culture Experience Program for Adolescents with Multicultural Backgrounds - for the Improvement of Self-Identity and Social Adaptation - (가정교과가 연계된 다문화가정 청소년의 한국문화 체험 프로그램의 개발과 평가 - 자아정체감 및 사회적응력 향상을 위하여 -)

  • Lee, Shin Sook;Kim, So Ra
    • Journal of Korean Home Economics Education Association
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    • v.25 no.4
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    • pp.63-77
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    • 2013
  • The purpose of this study is to develop the home economics-relevant program on Korean traditional culture for adolescents from multicultural families for them for experience it directly, set up ego-identity and improve social adaptability. Further, we checked the effectiveness and efficiency of the developed program. First we developed the program composed of a total of 12 sessions based on experiencing Korean culture. It consists of largely 5 madang(parts) centered on themes such as Eowool madang, Hangeul madang, Play madang, Cooking madang and hostory madang. We ran the developed program for 3 months targeting a total of 18 persons composed of the adolescents from multicultural families and their mothers residing in S city of eastern area of Jeonnam province. We measured the participants' ego-identity, social adaptibility and participation satisfaction to examine the effectiveness of the program. The results of pre-test, post-test and satisfaction which are evaluation instruments used in this program are as follows. First, it appeared that the self-identity of the adolescent of multicultural families inceased by 0.30 from average score of 3.03 before the participation in the program to 3.33 after the participation. Social adaptability increased by 0.30 from average 3.11 before the participation to 3.41 after the participation. Second, in the program participation satisfaction, it appeared that first the middle satisfaction marked average 3.30 which exceeded median value at 3.00, meaning a little high satisfaction, whereas the final satisfaction had an averagescore of 4.03, which means a big satisfaction concerning the program participation. The results indicated that the home economics-relevant program was effective in improving the ego-identity and social adaptability of adolescents from multicultural families and supporting a role of as a member of Korean family.

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Characterization and Control of Vascellum curtisii (Berkeley)Kreisel Causing the Fairy Ring Arcs in the Golf Course in Korea (골프코스에서 페어리링의 원인이되는 Vascellum curtisii의 특징과 방계)

  • Choi, Dae-Hong;Lee, Jung-Han;Kim, Hee-Kyu
    • Asian Journal of Turfgrass Science
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    • v.22 no.2
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    • pp.171-178
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    • 2008
  • We have found the clusters of tiny spiny puffball-like mushrooms growing gregariously in fairy ring (arcs) rimmed by a zone of darker green grass in the golf courses. Macroscopic as well as microscopic characters were examined for the morphology of fruiting body. Exoperidium is thin and densely spiny with minute fibrillae at early stage. The connivent spines were soft and quite persistent. In age, the fibrillae scrumble away with a powdery coating, which leaves white endoperidium becoming pale brown. It's interior was white and fleshy at first, but turns into an olive-colored dust as the gleba, the spore-producing tissue, develops to maturity and loaded with olive-brown spore mass. Then, distinct apical pore developed on the endoperidium. Rudimentary subgleba(sterile base) was narrow, chambered, delineated from the gleba by a membrane in young material. These characters suggested this fungus is a Vascellum, a member of the family Lycoperdaceae. The shapes of the spores were globose, echinulate, $3{\sim}3.5{\mu}m$ in diameter, thick-walled, and olive brown. Capillitial threads were $8-9{\mu}m$ wide, mostly colorless in KOH solution and thin-walled, which designated as "paracapillitium". This is an another character that distinguishes this mushroom from Lycoperdon spp. The spines developed on exoperidium were characteristically connivent; their apices joined together in a point, leaving a space below, which gives the appearance of vault to each group of usually 5 to 6 fibrillae. Based on the above characters, this fungus is identified as Vascellum curtisii (Berkeley). The characters distinguishable this from Lycoperdon pulcherrimum, and Vascellum pretense are discussed in detail. Control trial was also attempted. Strong vertical raking(SVR) followed by applying 500x detergent solution (Spark, Aekyung Co. Seoul) resulted in excellent control over any other treatments. In this plot, fruiting body was not developed throughout the end of mushroom growing season.

Cis-acting Replication Element Variation of the Foot-and-mouth Disease Virus is Associated with the Determination of Host Susceptibility (구제역바이러스의 숙주 특이성 결정에 연관되어있는 구제역바이러스 cis-acting replication element 변이 분석 연구)

  • Kang, Hyo Rin;Seong, Mi So;Ku, Bok Kyung;Cheong, JaeHun
    • Journal of Life Science
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    • v.30 no.11
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    • pp.947-955
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    • 2020
  • The foot-and-mouth disease virus (FMDV), a member of the Aphthovirus genus in the Picornaviridae family, affects wild and domesticated ruminants and pigs. During replication of the FMDV RNA (ribonucleic acid) genome, FMDV-encoding RNA polymerase 3D acts in a highly location-specific manner. This suggests that specific RNA structures recognized by 3D polymerase within non-coding regions of the FMDV genome assist with binding during replication. One such region is the cis-acting replication element (CRE), which functions as a template for RNA replication. The FMDV CRE adopts a stem-loop conformation with an extended duplex stem, supporting a novel 15-17 nucleotide loop that derives stability from base-stacking interactions, with the exact RNA nucleotide sequence of the CRE producing different RNA secondary structures. Here, we show that CRE sequences of FMDVs isolated in Korea from 2010 to 2017 exhibit A and O genotypes. Interestingly, variations in the RNA secondary structure of the Korean FMDVs are consistent with the phylogenetic relationships between these viruses and reveal the specificity of FMDV infections for particular host species. Therefore, we conclude that each genetic clade of Korean FMDV is characterized by a unique functional CRE and that the evolutionary success of new genetic lineages may be associated with the invention of a novel CRE motif. Therefore, we propose that the specific RNA structure of a CRE is an additional criterion for FMDV classification dependent on the host species. These findings will help correctly analyze CRE sequences and indicate the specificity of host species for future FMDV epidemics.