| 研究生: |
廖君宇 Chun-Yu Liao |
|---|---|
| 論文名稱: |
利用酵母菌蛋白質體晶片找出與前信使核糖核酸加 工因子19泛素連接酶經泛素化作用之受質 Ubiquitination subtract screening of ubiquitin ligase pre-mRNA-processing factor 19 using yeast proteome microarrays |
| 指導教授: |
陳健生
Chien-Sheng Chen |
| 口試委員: | |
| 學位類別: |
碩士 Master |
| 系所名稱: |
生醫理工學院 - 系統生物與生物資訊研究所 Graduate Institute of Systems Biology and Bioinformatics |
| 論文出版年: | 2017 |
| 畢業學年度: | 105 |
| 語文別: | 英文 |
| 論文頁數: | 65 |
| 中文關鍵詞: | 前信使核糖核酸加 工因子19 、泛素 、泛素化 、酵母菌蛋白質體晶片 、受質 |
| 外文關鍵詞: | pre-mRNA-processing factor 19, ubiquitin, Ubiquitination, yeast proteome microarrays, subtract |
| 相關次數: | 點閱:12 下載:0 |
| 分享至: |
| 查詢本校圖書館目錄 查詢臺灣博碩士論文知識加值系統 勘誤回報 |
泛素為一常見於細胞中的小蛋白質,主要用以標記蛋白質受質,並進行後續如去氧
核醣核酸修復、蛋白質水解等路徑。標記泛素的過程稱謂泛素化,其透過泛素活化酶、 泛素結合酶與泛素連接酶將特定受質接上泛素。其中,泛素連接酶負責辨識特定的受質, 扮演泛素化中的關鍵角色。前信使核糖核酸加工因子 19 為其中一種泛素連接酶,與去 氧核醣核酸修復與信使核糖核酸剪接有所關聯,並被發現在肺癌中有過度表現的現象。 我對大於五千八百種蛋白質的酵母菌蛋白質體晶片進行前信使核糖核酸加工因子 19 的 胞外泛素化實驗,篩選出其對應的四十五個受質,並透過生物資訊的方式找到這四十五 個受質共有結合序列與前信使核糖核酸加工因子 19 透過泛素化受質而進一步產生關聯 的十類生物機制、七類細胞組件、六類分子功能與十二類路徑。另外,在四十五個受質 中,四十個牽涉到七類生物機制、四類細胞組件以及兩類路徑與前信使核糖核酸加工因 子 19 有關聯。此外,有三類生物機制、六類細胞組件和三類分子功能呈現顯著富集。 還有,我找出兩組受質分別有不同的序列模組,提供與前信使核糖核酸加工因子 19 可 能的結合點。根據結果,我找到這些基因本體論與路徑可能透過受質泛素化而被前信使 核糖核酸加工因子 19 所調控,對前信使核糖核酸加工因子 19 進一步的研究方向。
Ubiquitin is a common small protein in cells, it usually served as tag conjugated to the substrate, for DNA repair, proteolysis, and others. This process is called ubiquitination, which ubiquitinates specific substrates by ubiquitin activation enzyme, ubiquitin conjugating enzyme and ubiquitin ligase. Ubiquitin ligase serves as a critical point for substrates specificity. Pre- mRNA-processing factor 19 (PRP19) is one of the ubiquitin ligases, involves in DNA repair, mRNA splicing, and has been found overexpressed in lung cancer. I found forty-five substrates of PRP19 from in vitro ubiquitination assay on yeast proteome chip which contains more than 5800 kinds of the yeast protein. I used bioinformatics analysis to find PRP19 further involving ten classes of biological processes, seven classes of cellular components, six classes of molecule functions, and twelve classes of pathways by ubiquitinates its substrates. Forty of forty-five substrates involved in seven classes of biological processes, four classes of cellular components and two pathways which relate to PRP19.In addition, there were three classes of biological processes, six classes of cellular components, three classes of molecule functions were significantly enriched. Also, I found two groups of substrates had different consensus motifs as putative binding sites with PRP19. According to the result, I found those GO and pathways might be regulated by PRP19-substrates ubiquitination, it gave the direction for
further study on PRP19.
1. Hershko, A. and A. Ciechanover, The ubiquitin system. Annu Rev Biochem, 1998. 67: p. 425-79.
2. Pickart, C.M., Mechanisms underlying ubiquitination. Annu Rev Biochem, 2001. 70: p. 503-33.
3. Pickart, C.M. and D. Fushman, Polyubiquitin chains: polymeric protein signals. Curr Opin Chem Biol, 2004. 8(6): p. 610-6.
4. Berndsen, C.E. and C. Wolberger, New insights into ubiquitin E3 ligase mechanism. Nat Struct Mol Biol, 2014. 21(4): p. 301-7.
5. Finley, D., et al., The ubiquitin-proteasome system of Saccharomyces cerevisiae. Genetics, 2012. 192(2): p. 319-60.
6. Dawson, T.M. and V.L. Dawson, Molecular pathways of neurodegeneration in Parkinson's disease. Science, 2003. 302(5646): p. 819-22.
7. Liu, Y.C., Ubiquitin ligases and the immune response. Annu Rev Immunol, 2004. 22: p. 81-127.
8. Confalonieri, S., et al., Alterations of ubiquitin ligases in human cancer and their association with the natural history of the tumor. Oncogene, 2009. 28(33): p. 2959- 68.
9. Gupta, R., et al., Ubiquitination screen using protein microarrays for comprehensive identification of Rsp5 substrates in yeast. Mol Syst Biol, 2007. 3: p. 116.
10. Loch, C.M., M.J. Eddins, and J.E. Strickler, Protein microarrays for the identification of praja1 e3 ubiquitin ligase substrates. Cell Biochem Biophys, 2011. 60(1-2): p. 127-35.
11. Andrews, P.S., et al., Identification of substrates of SMURF1 ubiquitin ligase activity utilizing protein microarrays. Assay Drug Dev Technol, 2010. 8(4): p. 471-87.
12. Marechal, A., et al., PRP19 transforms into a sensor of RPA-ssDNA after DNA
damage and drives ATR activation via a ubiquitin-mediated circuitry. Mol Cell, 2014.
53(2): p. 235-46.
13. Song, E.J., et al., The Prp19 complex and the Usp4Sart3 deubiquitinating enzyme
control reversible ubiquitination at the spliceosome. Genes Dev, 2010. 24(13): p.
1434-47.
14. Das, T., et al., USP15 regulates dynamic protein-protein interactions of the
spliceosome through deubiquitination of PRP31. Nucleic Acids Res, 2017. 45(8): p. 4866-4880.
54
15. Bellare, P., et al., A role for ubiquitin in the spliceosome assembly pathway. Nat Struct Mol Biol, 2008. 15(5): p. 444-51.
16. Zhu, H., et al., Global analysis of protein activities using proteome chips. Science, 2001. 293(5537): p. 2101-5.
17. Chen, Y.W., et al., Identification of bacterial factors involved in type 1 fimbria expression using an Escherichia coli K12 proteome chip. Mol Cell Proteomics, 2014. 13(6): p. 1485-94.
18. Lu, J.Y., et al., Functional dissection of a HECT ubiquitin E3 ligase. Mol Cell Proteomics, 2008. 7(1): p. 35-45.
19. Apweiler, R., et al., UniProt: the Universal Protein knowledgebase. Nucleic Acids Res, 2004. 32(Database issue): p. D115-9.
20. Ashburner, M., et al., Gene ontology: tool for the unification of biology. The Gene Ontology Consortium. Nat Genet, 2000. 25(1): p. 25-9.
21. Huang da, W., B.T. Sherman, and R.A. Lempicki, Systematic and integrative analysis of large gene lists using DAVID bioinformatics resources. Nat Protoc, 2009. 4(1): p. 44-57.
22. Croft, D., et al., Reactome: a database of reactions, pathways and biological processes. Nucleic Acids Res, 2011. 39(Database issue): p. D691-7.
23. Frith, M.C., et al., Discovering sequence motifs with arbitrary insertions and deletions. PLoS Comput Biol, 2008. 4(4): p. e1000071.
24. Carlson, M., GO.db: A set of annotation maps describing the entire Gene Ontology. R package version 3.4.1. 2017.
25. Binns, D., et al., QuickGO: a web-based tool for Gene Ontology searching. Bioinformatics, 2009. 25(22): p. 3045-6.
26. Norbeck, J., et al., Purification and Characterization of Two Isoenzymes of DL- Glycerol-3-phosphatase from Saccharomyces cerevisiae. Journal of Biological Chemistry, 1996. 271(23): p. 13875-13881.