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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 翁秀貞(Shiou-Jeng Ong) | |
| dc.contributor.author | Chung-Chi Kuo | en |
| dc.contributor.author | 郭純綺 | zh_TW |
| dc.date.accessioned | 2021-06-15T00:15:16Z | - |
| dc.date.available | 2014-10-07 | |
| dc.date.copyright | 2011-10-07 | |
| dc.date.issued | 2011 | |
| dc.date.submitted | 2011-08-15 | |
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Mann, Proteomics to study genes and genomes. Nature, 2000. 405(6788): p. 837-46. 62. Steen, H. and A. Pandey, Proteomics goes quantitative: measuring protein abundance. Trends Biotechnol, 2002. 20(9): p. 361-4. 63. De Jesus, J.B., et al., A further proteomic study on the effect of iron in the human pathogen Trichomonas vaginalis. Proteomics, 2007. 7(12): p. 1961-72. 64. Seong, H.A., R. Manoharan, and H. Ha, B-MYB positively regulates serine-threonine kinase receptor-associated protein (STRAP) activity through direct interaction. J Biol Chem, 2011. 286(9): p. 7439-56. 65. Vaysse, L., et al., Development of a self-assembling nuclear targeting vector system based on the tetracycline repressor protein. J Biol Chem, 2004. 279(7): p. 5555-64. 66. Pollard, V.W., et al., A novel receptor-mediated nuclear protein import pathway. Cell, 1996. 86(6): p. 985-94. 67. Michael, W.M., P.S. Eder, and G. 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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/41287 | - |
| dc.description.abstract | 先前的研究證實陰道滴蟲Myb2及Myb3是能調控致病因子ap65-1基因的轉錄因子，當受到外界不同訊號例如鐵或H2O2的刺激時，可能使Myb2及Myb3進行轉譯後修飾，影響其進入細胞核調控下游基因的表現。目前有關陰道滴蟲中Myb如何入核之機制仍然未明，因此本研究目的是希望釐清是否Myb2及Myb3在細胞質中會與其它蛋白質結合形成複合體，並進一步找尋與Myb2及Myb3相互作用的蛋白質，探討這些蛋白質是否參與陰道滴蟲Myb蛋白質之入核運送。
利用膠體過濾法並以西方墨點法分析，發現wild type Myb2在細胞質中大部分為單體，亦有少部分是以複合體形式存在；wild type Myb3在細胞質中則是在加鐵後會形成大的複合體。當破壞Myb2結構或刪除重要序列時，除了會明顯抑制其入核外，亦會影響複合體形成；Myb3則是在短時間的鐵刺激下會促進其入核，且培養於普通或高鐵環境中時，亦會改變複合體形成。顯示Myb2及Myb3的複合體形成與入核均有關聯性，可能皆需經由與複合體內之蛋白質結合並受其調控。 由螢光免疫染色法也發現Myb2及Myb3主要分佈於細胞核中，推測其入核應是藉由主動運輸而非擴散作用。由於Myb2及Myb3與其他分子作用時間可能很短暫，利用免疫沉澱法無法找到與其交互作用之蛋白質，為了更有效率地尋找複合體中可能與其交互作用之蛋白質，本研究建立了crosslinking－IP實驗，並且藉由蛋白質體學的方法及液相層析串聯質譜儀 (Liquid chromatography-tandem mass spectrometry) 鑑定蛋白質種類。未來將以此方法繼續尋找與調控陰道滴蟲Myb入核相關之重要蛋白質，以瞭解Myb2及Myb3入核相關之訊息傳遞路徑及其在陰道滴蟲基因調控上所扮演的角色。 | zh_TW |
| dc.description.abstract | Previous study showed that Myb2 and Myb3 transcription factors may regulate inducible transcription of an adhesion protein, ap65-1, gene in the protozoan parasite Trichomonas vaginalis, and external stimuli, such as iron and H2O2 , may affect entry of these proteins into the nucleus, presumably to regulate downstream gene expression. The purpose of this study was to clarify whether Myb2 or Myb3 may complex with other proteins in the cytoplasm.
Using the gel filtration and Western blot assay, the data showed that Myb2 and Myb3 could form complexes in the cytoplasm. Myb2 and Myb3 were mainly localized to the nucleus by an immuneflouorescence assay (IFA), suggesting that they may be actively translocated into the nucleus. I explored a cross-linking coupled immunoprecipitation (IP) approach to purify Myb-interacting proteins, and used liquid chromatography-tandem mass spectrometry for protein identification. Although the protocol needs to be further refined, this work may provide tool for further investigation the roles of Myb proteins on inducible-gene regulation. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-15T00:15:16Z (GMT). No. of bitstreams: 1 ntu-100-R97445204-1.pdf: 2640226 bytes, checksum: d53d4e7637f50ba8cd9e480e027321a0 (MD5) Previous issue date: 2011 | en |
| dc.description.tableofcontents | 口試委員審定書................................. .......................I
誌謝..................................................................II 目錄.................................................................III 縮寫名詞對照表......................................................VIII 中文摘要............................................................IX 英文摘要.............................................................X 前言 一、陰道滴蟲簡介..........................................1 二、 Myb轉錄因子......................................................1 三、 鐵離子對於陰道滴蟲之重要性........................................3四、 ROS對於轉錄因子的影響...........................................4五、 蛋白質生成及核運輸機制............................................6 六、 蛋白質體學發展與運用.............................................9 七、 本研究之目的.....................................................11 材料與方法 一、 陰道滴蟲蟲株....................................................12 1. 細胞株培養與處理.............................................12 2. 質體構築.....................................................12 3. 陰道滴蟲轉染、選殖與單株化...................................12 4. 蟲株之保存與活化.............................................13 二、 蛋白質樣品製備..................................................13 三、 蛋白質聚丙烯醯胺膠體電泳........................................14 四、 西方墨點法(Western blot assay)......................................14 口試委員審定書................................. .......................I 誌謝..................................................................II 目錄.................................................................III 縮寫名詞對照表......................................................VIII 中文摘要............................................................IX 英文摘要.............................................................X 前言 一、陰道滴蟲簡介..........................................1 二、 Myb轉錄因子......................................................1 三、 鐵離子對於陰道滴蟲之重要性........................................3四、 ROS對於轉錄因子的影響...........................................4五、 蛋白質生成及核運輸機制............................................6 六、 蛋白質體學發展與運用.............................................9 七、 本研究之目的.....................................................11 材料與方法 一、 陰道滴蟲蟲株....................................................12 1. 細胞株培養與處理.............................................12 2. 質體構築.....................................................12 3. 陰道滴蟲轉染、選殖與單株化...................................12 4. 蟲株之保存與活化.............................................13 二、 蛋白質樣品製備..................................................13 三、 蛋白質聚丙烯醯胺膠體電泳........................................14 四、 西方墨點法(Western blot assay)......................................14 附表................................................................75 參考文獻............................................................77 附錄..................................................................85 | |
| dc.language.iso | zh-TW | |
| dc.subject | Myb2 | zh_TW |
| dc.subject | 陰道滴蟲 | zh_TW |
| dc.subject | Myb3轉錄因子 | zh_TW |
| dc.subject | 膠體過濾法 | zh_TW |
| dc.subject | 複合體形成 | zh_TW |
| dc.subject | 液相層析串聯質譜儀 | zh_TW |
| dc.subject | liquid chromatography-tandem mass spectrometry | en |
| dc.subject | Trichomonas vaginalis | en |
| dc.subject | Myb2 | en |
| dc.subject | Myb3 | en |
| dc.subject | gel filtration | en |
| dc.subject | cross-linking | en |
| dc.subject | immunoprecipitation | en |
| dc.title | 探討陰道滴蟲Myb蛋白質複合體形成及入核運輸 | zh_TW |
| dc.title | The complex formation and nuclear translocation of Myb proteins in Trichomonas vaginalis | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 99-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 戴榮湘(Jung-Hsiang Tai) | |
| dc.contributor.oralexamcommittee | 蕭信宏(Shin-Hong Shiao) | |
| dc.subject.keyword | 陰道滴蟲,Myb2,Myb3轉錄因子,膠體過濾法,複合體形成,液相層析串聯質譜儀, | zh_TW |
| dc.subject.keyword | Trichomonas vaginalis,Myb2,Myb3,gel filtration,cross-linking,immunoprecipitation,liquid chromatography-tandem mass spectrometry, | en |
| dc.relation.page | 90 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2011-08-15 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 微生物學研究所 | zh_TW |
| 顯示於系所單位： | 微生物學科所 | |
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