• Journal of Microbiology and Biotechnology
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• 제9권1호
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• pp.122-125
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• 1999

Gcn4p, a transcriptional activator protein of the yeast, Sacchromyces cerevisiae, binds to the specific sequence in the promoters of many amino acid biosynthetic genes for general control. The serine residue (Ser 242) of Gcn4p directly contacts the DNA. Here, for inspecting the DNA binding properties and the level of transcriptional activation of Gcn4p, we introduced a polymerase chain reaction (PCR) site-directed saturation mutation library into the Ser 242 site using 2 outside primers and 2 oligonucleotides with its codons fully degenerated. The sequencing analysis of 146 samples revealed the even nucleotide distribution within the experimental error showing 23, 26, 25, and 26％ frequency of U, C, A, and G bases, respectively. This method turned out to be a simple, fast, and economical method for constructing a library of all 20 amino acids at specific codon.

• 한국식품영양과학회지
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• 제40권9호
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• pp.1321-1327
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• 2011

전화당(invert sugar)을 생산하기 위하여 포도주로부터 분리한 효모가 생산하는 invertase의 특성을 조사한 결과는 다음과 같다. 포도주로부터 분리한 효모는 지방산 분석을 통하여 Saccharomyces cerevisiae JS59로 잠정 동정되었다. 본 균주가 생성하는 invertase를 ammonium sulfate 침전, DEAE-Sephadex A-50, Sephadex G-200 column chromatography 법으로 정제하였을 때 단일성을 보였으며, specific activity가 7620.9 unit/mg, 최종 회수율은 13.9로 약 14배 정제된 효소를 얻었다. 본 효소의 $K_m$ 값은 11.5 mM이었다. SDS-PAGE로부터 분자량은 38.5 kDa으로 나타났다. 정제된 invertase의 최적 pH는 5였고, pH 4에서도 94%의 높은 효소활성을 나타냈으며 4에서 6까지의 pH영역에서 안정하였고, $55^{\circ}C$에서 최적 활성을 나타내었으며 $50^{\circ}C$까지는 안정하였다. $Ag^{2+}$와 $Hg^{2+}$에 의해서 저해를 받았고, $Co^{2+}$, $Mn^{2+}$에 의해서는 효소활성이 증가되었으며, 기질과 효소 반응물을 thin layer chromatography로 분석한 결과, 본 효소는 기질인 sucrose를 완전히 분해하여 환원당을 생성함이 확인되었다.

• Journal of Microbiology and Biotechnology
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• 제15권5호
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• pp.965-970
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• 2005

In our previous study [14], we found that heat-shock exposure did not stimulate the neutral trehalase activity in Sacchromyces cerevisiae KNU5377, but did in ATCC24858. Consequently, the trehalose content in KNU5377 became 2.6 times higher than that in ATCC24858. Because trehalose has been shown to stabilize the structure and function of some macromolecules, the present work was focused to elucidate the relationship between trehalose content of these strains and thermal stabilities of whole cells, through differential scanning calorimetry (DSC), and to predict critical targets calculated from the hyperthermic cell killing rates. These analyses showed that the prominent DSC transition of both strains gave identical $T_m$ (transition temperature) values in exponentially growing cells, and that the $T_m$ values of critical targets was about $3^{\circ}C$ higher in KNU5377 than in ATCC24858. Both heat-shocked KNU5377 and ATCC24858 cells displayed similar shifts in their DSC transition profiles. On the other hand, the $T_m$ value of the critical target of KNU5377 was decreased by $2.1^{\circ}C$, which was still higher than ATCC24858 showing no changes. In view of these results, the intrinsic thermotolerance of KNU5377 did not appear to result from the stability of entire cellular components, but rather possibly from that of particular macromolecules, including critical targets, even though it should be investigated in more details. Although the trehalose levels in heat-shocked cells are significantly different, as described in our previous study [14], the overall pattern of thermal stabilities and their predicted critical targets in two heat-shocked strains seemed to be identical. These data suggest that the trehalose levels examined before and after heat shock of exponentially growing cells are not closely correlated with the stabilities of whole cells and/or critical targets in both yeast strains.

• BMB Reports
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• 제30권3호
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• pp.167-172
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• 1997

The yeast Saccharomyces cerevisiae, mutant CH117, shows a drug-hypersensitivity (dhs) to cycloheximide, bleomycin, actinomycin D, 5-fluorouracil. nystatin, nigericin and several other antibiotics. CH 117 was also temperature-sensitive (ts). being unable to grow at $37^{\circ}C$ and secreted more invertase and acid phosphatase into the medium than the parent yeast. CH117 grows very slowly and the cell shape is somewhat larger and more sensitive to zymolyase than the wild type cells. Light microscopic and electron microscopic observation also revealed abnormality of the mutant cell wall. These characteristics indicate that CH117 has a defect in an essential component of the cell surface and that the cell wall which performs barrier functions has become leaky in the mutant. We screened a genomic library of wild type yeast for clones that can complement the mutation of CH117. A plasmid, pCHX1, with an insert of 3.6 kilobases (kbs) could complement the dhs and ts of CH117. Deletion and subcloning of the 3.6 kb insert showed that a gene for the complementation of mutant phenotypes was located in 1.9 kbs Puvll-Hindlll fragment.

• 자연과학논문집
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• 제15권1호
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• pp.79-88
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• 2005

효모에는 여러 가지 종류의 thiol peroxid se 동위 효소들인 cytoplasmic TPx I, cTPx II, cTPx III, mitochodrial TPx (mTPx), 및 nuclear TPx (nTPx)가 존재하고 있다. 특히 cTPx II는 다른 효모TPx와 비교해 볼 때 매우 낮은 peroxidase 활성을 보이나, cTPx II를 제거한 cTPx II mutant균주는 심하게 성장이 저해되는 특징을 보인다. 본 연구에서는 효모에서의 cTPx II의 생리학적 기능을 밝히는 연구의 첫 과정으로 cTPx II와 상호 작용하는 단백질을 탐색하였다. Sacchromyces cerevisiae genomic DNA library에서 yeast two-hybrid system을 이용하여 cTPx II와 상호 결합하는 단백질을 탐색하여 그 단백질들의 기능을 연구하여, 궁극적으로 cTPx II의 생리기능을 밝히는데 이 연구의 목적을 두었다.

• 대한본초학회지
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• 제23권3호
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• pp.103-112
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• 2008

Objectives : This research aimed to study the cytotoxicity and immuno modulating activity of fermented Artemisia argyi Lev. et Vant.(Compositae). Methods : Effect of fermented Artemisiae Argyi Folium extracts, which were fermented by Sacchromyces cerevisiae STV89(AFS), on cell viability, generation of ROS within cells, generation of NO and the level of cytokines($TNF-{\alpha}$ and IL-6) was measured using mouse macrophage RAW 264.7 cell. Results : 1. Result of MTT assay conducted to verify the cytotoxicity of fermented Artemisiae argyi folium extract illustrated that, when fermented Artemisiae argyi folium extract was processed for each concentration, there was no excessive induction of cytoxicity in the RAW 264.7 cell. 2. Fermented Artemisiae Argyi Folium extract increased the generation of H2O2 within RAW 264.7 cell as well as significantly increased inhibition of generation of H2O2 in macrophage induced by LPS. 3. Fermented Artemisiae Argyi Folium extract inhibited generation of NO in RAW 264.7 cell, and significantly inhibited increase in generation of NO of macrophage induced by LPS. 4. Fermented Artemisiae Argyi Folium extract, AFS has significantly reduced the increase in the generation of $TNF-{\alpha}$ above 10 ${\mu}g/mL$. 5. Fermented Artemisiae Argyi Folium extract, AFS has significantly reduced the increase in generation of IL-6 above 50 ${\mu}g/mL$. Conclusions : AFS fermented extract produced from Artemisiae Argyi Flium, have increased generation of ROS and reduced generation of NO in RAW 264.7 cell without excessively inducing cytotoxicity of RAW 264.7 cell. In addition, they displayed significant immuno modulating activities including inhibition of generation of $TNF-{\alpha}$ and IL-6 in macrophage, induced by LPS.

• 대한본초학회지
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• 제28권3호
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• pp.107-113
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• 2013

Objective : The purpose of this study was to investigate the effect of fermented Artemisiae Argyi Folium(AAF) on some activities of human hepatoma cell, HepG2. Method : To investigate the effect of fermented Artemisiae Argyi Folium(AAF) activity on the human hepatoma cells, AAF extracts was fermented by Lactobacillus pentosus K34(AFL) and Sacchromyces cerevisiae STV89(AFS). And the effects of AFL or AFS on the activities of HepG2 cell, such as cell viability, nitric oxide(NO) production and reactive oxygen species(ROS) production, were tested. Result : Human Hepatoma Cells were incubated each for 3 hours and 24 hours. Human Hepatoma Cells treated with the extract was measured with MTT assay. Then AFL was found to be non-toxic at concentrations of 10 ug/mL(3h), 100 ug/mL(24h) or more. AFS was the same result at concentrations of more than 10 ug/mL. The extract increased ROS generation in Human Hepatoma Cells. AFL increased at concentrations of 100 ug/mL more (3h, also 10 ug/mL more) and 50 ug/mL(24h) and AFS increased both 50 ug/mL. In point of NO generation, AFL inhibited at concentrations of 10 ug/mL(3h) and 100 ug/mL(24h) more (3h, also 10 ug/mL more) and AFS also inhibited 50 ug/mL or more. Conclusion : AFL and AFS, obtained from Artemisiae Argyi Folium extracts by fermentation, reduced the NO production and increased ROS production in HepG2 cell, without cytotoxicity on HepG2 cell. The results suggested that AFL and AFS increased the immunological effects of Artemisiae Argyi Folium extracts.

• 대한한방부인과학회지
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• 제22권2호
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• pp.79-93
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• 2009

Purpose: This research aimed to study the effect of FAE(Ferment Artemisiae Argyi Folium and Epimedii Herba) on the mouse macrophage cell activity. Methods: Effect of FAE, which was fermented by Sacchromyces cerevisiae STV89, on cell viability, amount of $H_2O_2$ within cells, amount of NO was measured and compaperd by using mouse macrophage cells. Results: 1. Result of MTT assay conducted to observe the effect of FAE on the survival rate of mouse macrophage cells illustrated that, when FAE was proccessed for each concentration, there was no significant decrease of the survival rate. 2. FAE increased the amount of $H_2O_2$ within macrophage cells and increased inhibition of amount of $H_2O_2$ in macrophage induced by LPS. 3. FAE inhibited amount of NO in macrophage cells, and significantly inhibited increase of amount of NO in mcacrophage induced by LPS. Conclusion: FAE produced by Artemisiae Argyi Folium and Epimedii Herba did not induce the decrease of macrophage cell survival rate, increased amount of $H_2O_2$ within cells, and reduced amount of NO. FAE significantly increase by LPS, reduced the increase of amount of NO in macrophage induced by LPS. These results signify FAE has significant effect on immuno modulating activity of macrophage.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 2005년도 2005 Annual Meeting & International Symposium
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• pp.154-164
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• 2005

Saccharomyces cerevisiae KNU5377 is a thermotolerant strain, which can ferment ethanol from wasted papers and starch at 40$^{\circ}C$ with the almost same rate as at 30$^{\circ}C$. This strain showed alcohol fermentation ability to convert wasted papers 200 g (w/v) to ethanol 8.4% (v/v) at 40$^{\circ}C$, meaning that 8.4% ethanol is acceptable enough to ferment in the industrial economy. As well, all kinds of starch that are using in the industry were converted into ethanol at 40$^{\circ}C$ with the almost same rate as at 30$^{\circ}C$. Hyperthermic cell killing kinetics and differential scanning calorimetry (DSC) revealed that exponentially growing cells of this yeast strain KNU5377 were more thermotolerant than those of S. cerevisiae ATCC24858 used as a control. This intrinsic thermotolernace did not result from the stability of entire cellular components but possibly from that of a particular target. Heat shock induced similar results in whole cell DSC profiles of both strains and the accumulation of trehalose in the cells of both strains, but the trehalose contents in the strain KNU5377 were 2.6 fold higher than that in the control strain. On the contrary to the trehalose level, the neutral trehalase activity in the KNU5377 cells was not changed after the heat shock. This result made a conclusion that though the trehalose may stabilize cellular components, the surplus of trehalose in KNU5377 strain was not essential for stabilization of whole cellular components. A constitutively thermotolerant yeast, S. cerevisiae KNU5377, was compared with a relatively thermosensitive control, S. cerevisiae ATCC24858, by assaying the fluidity and proton ATPase on the plasma membrane. Anisotropic values (r) of both strains were slightly increased by elevating the incubation temperatures from 25$^{\circ}C$ to 37$^{\circ}C$ when they were aerobically cultured for 12 hours in the YPD media, implying the membrane fluidity was decreased. While the temperature was elevated up to 40$^{\circ}C$, the fluidity was not changed in the KNU5377 cell, but rather increased in the control. This result implies that the plasma membrane of the KNU5377 cell can be characterized into the more stabilized state than control. Besides, heat shock decreased the fluidity in the control strain, but not in the KNU5377 strain. This means also there's a stabilization of the plasma membrane in the KNU5377 cell. Furthermore, the proton ATPase assay indicated the KNU5377 cell kept a relatively more stabilized glucose metabolism at high temperature than the control cell. Therefore, the results were concluded that the stabilization of plasma membrane and growth at high temperature for the KNU5377 cell. Genome wide transcription analysis showed that the heat shock responses were very complex and combinatory in the KNU5377 cell. Induced by the heat shock, a number of genes were related with the ubiquitin mediated proteolysis, metallothionein (prevent ROS production from copper), hsp27 (88-fold induced remarkably, preventing the protein aggregation and denaturation), oxidative stress response (to remove the hydrogen peroxide), and etc.

• 한국가금학회지
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• 제35권2호
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• pp.123-129
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• 2008

본 연구의 목적은 고에너지 사료 내 항생제와 효모제의 첨가가 육계의 생산성, 혈액 성상 및 도체 특성에 미치는 영향을 알아보기 위하여 실시하였다. 2일령 Arbor Acre broiler (male, female) 480수를 공시하였고, 사양 시험은 5주간 실시하였다. 시험사료는 옥수수-대두박 위주의 사료인 대조구(CON; basal diet), 고에너지 사료 처리구(HED; high energy diet), 효모제 사료 처리구(YD; HED without virginiamycin + 0.2% Saccharomyces cerevisiae, $15{\times}10^{10}$)로 3개 처리를 하여 처리당 8반복, 반복당 20수씩 완전 임의 배치하였다. 전체 사양 시험 기간동안 증체량은 고에너지 사료를 급여한 HED 처리구가 다른 처리구들에 비해 높은 경향을 보였으나, 처리구간에 유의적인 차이는 없었다. 사료 섭취량은 효모제 사료를 급여한 YD 처리구가 다른 처리구들에 비해 높은 경향을 보였으나, 처리구간에 유의적인 차이는 없었다. 사료 요구율에서는 고 에너지 사료를 급여한 HED 처리구가 다른 처리구들에 비해 유의적으로 낮게 나타났다(P<0.05). RBC(red blood cell), WBC (white blood cell), lymphocyte 모두 처리구간에 유의적인 차이는 없었으며, 간 무게 및 생체중에 대한 간 무게 비율, 허벅지육 무게 및 생체중에 대한 허벅지육 비율, 가슴육 무게 및 생체중에 대한 가슴육 무게 비율, 복강 지방 무게 및 생체중에 대한 복강 지방 무게 비율 모두에서 유의적인 차이가 없었으나, 종료시 체중에서는 HED 처리구가 유의적으로 가장 높았으며, 다음으로 YD, CON 순이었다(P<0.05). 결론적으로, 고에너지 사료 처리구가 사료 요구율에서 유의적으로 가장 영향을 미쳤으며, 효모제 처리구는 CON 처리구와 많은 차이를 보이지 않았다.