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研究生:吳權哲
研究生(外文):Cyuan-jhe Wu
論文名稱:釀酒酵母在過氧化氫存在環境下生長時yArsA蛋白所扮演的角色
論文名稱(外文):The Role of yArsA in Saccharomyces cerevisiae during growth in the presence of hydrogen peroxide
指導教授:許清玫
指導教授(外文):Ching-mei Hsu
學位類別:碩士
校院名稱:國立中山大學
系所名稱:生物科學系研究所
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2006
畢業學年度:94
語文別:英文
論文頁數:37
中文關鍵詞:過氧化氫釀酒酵母氧化
外文關鍵詞:Saccharomyces cerevisiaehydrogen peroxideoxidative stress
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釀酒酵母Saccharomyces cerevisiae第四號染色體上ORFs YDL100c之蛋白產物yArsA為大腸桿菌ArsA之同源蛋白,ArsA具有ATPase活性,可將細胞中的重金屬砷及銻排出,但yArsA的功能尚未清楚。當酵母菌在正常生存環境下yArsA的缺失並不會造成酵母細胞生長差異。為了研究yArsA在釀酒酵母在氧化壓力下所扮演的角色,本實驗將野生株(WT)及突變株(KO)培養於含1 mM H2O2培養液,並針對細胞內的自由基清除系統、trehalose的含量做分析。初步實驗結果證明:生長於含1 mM H2O2培養液,KO菌株酵母細胞其生長情形較野生型菌株差且細胞內分子氧化程度較WT菌株為高,而其catalase的活性則較WT菌株低,ROS的增加以及catalase活性的降低可能是導致KO菌株細胞死亡的原因。以RT-PCR分析自由基清除系統以及trehalose代謝相關基因的mRNA表現量,結果指出生長於含1 mM H2O2培養液的WT和KO菌株,其GSH1、SOD1與TRR1的表現並無顯著差異而KO菌株的CTT1, TPS1, NTH1表現量則低於WT菌株,KO菌株的trehalose含量高於WT菌株。Trehalose代謝相關基因轉錄的下降造成trehalose分解效率下降,catalase活性的降低指出KO菌株在氧化壓力下調控general stress response能力的缺失。yArsA在酵母菌Saccharomyces cerevisiae中可能扮演一個對氧化傷害壓力適應的角色。
The E. coli ArsA is involved in arsenic detoxification but the role of yArsA (ArsA homologue of Saccharomyces cerevisiae, encoded by YDL100C ORF) in yeast is still undefined. Disruption of YDL100C ORF is not lethal. To study the role of yArsA in oxidative tolerance, wild type and knock out strain were grown in presence or absence of 1 mM H2O2 and assayed the expression of anti-oxidation machanisms . The results show that molecular oxidation is higher and catalase activity is lower in KO compared with WT. It suggests that increased ROS and decreased catalase activity are the cause of cell death. Further analysis of the expression of ROS defense mechanisms by RT-PCR show that there is no significant difference in TRR1, GSH1, and SOD1 expression in WT and KO grown in presence of 1 mM H2O2 but the CTT1, TPS1, NTH1 expression in KO are less than WT grown under oxidative stress. GSH contents is consistent with the result of RT-PCR, and trehalose contents is higher in KO strain under oxidative stress. Loss of catalase activity and decreased efficiency of degrading trehalose suggest that the deficiency in activation of general stress response in KO when grown in the presence of H2O2. Therefore, yArsA would be involved in expressing the general stress response in oxidative tolerance.
Abstract------------------------------------------1
Introduction--------------------------------------3
Materials and Methods-----------------------------8
Results------------------------------------------13
Discussion---------------------------------------16
Figures------------------------------------------18
Tables-------------------------------------------28
References---------------------------------------30
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