| 研究生: |
鄭皓蔚 Hao-Wei Cheng |
|---|---|
| 論文名稱: |
以色胺酸及FlAsH組成之凋亡蛋白酶3螢光感測多肽 FlAsH Labeled Peptide with Tryptophan Residues as a Caspase-3 Sensor |
| 指導教授: |
李賢明
Hsein-Ming Lee 謝發坤 Fa-Kuen Shieh |
| 口試委員: | |
| 學位類別: |
碩士 Master |
| 系所名稱: |
理學院 - 化學學系 Department of Chemistry |
| 論文出版年: | 2015 |
| 畢業學年度: | 103 |
| 語文別: | 中文 |
| 論文頁數: | 116 |
| 中文關鍵詞: | 感測器 、凋亡蛋白酶3 、螢光 |
| 相關次數: | 點閱:11 下載:0 |
| 分享至: |
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本論文是以色胺酸對螢光團螢光淬滅的特性為基礎,設計並合成蛋白酶活性
感測器 FlAsH-3W2F-TCǶ利用一系列 FlAsH 螢光染料標示胜肽的螢光性質進行研究,發現帶有芳香環側鏈的胺基酸中,相較於阻胺酸與苯Ҙ胺酸,色胺酸對FlAsH 螢光染料的螢光有最好的淬滅效果;胜肽上的色胺酸數量越多對螢光的淬滅效果也越好Ƕ色胺酸對於螢光染料的淬滅是根據光誘發電子轉移,透過螢光分子與色胺酸之間碰撞所形成暫時的分子軌域,提供電子轉移的通道,使螢光團激發的能量得到鬆弛Ƕ在以 Trypsin 為模型的蛋白酶活性實驗中,感測器水解前後有將近 9 倍的螢光強度增加;在 Caspase-3 目標蛋白酶活性實驗中,也觀測到明顯的螢光增強Ƕ以色胺酸做為淬滅團在螢光感測器上的應用是可行的。
We synthesized a Caspase-3 sensor, FlAsH-3W2F-TC, whose fluorescence was quenched by tryptophan inside the peptidyl sensor backbone. By studying a series of FlAsH-labeled peptide sequences, tryptophan containing peptide sequence showed higher quenching ability than histidine and phenylalanine peptide sequences; the more tryptophan the peptide sequence contains, the higher quenching efficiency we get. The quenching phenomena is based on photo-induced electron transfer(PET). The excited fluorophore relax by conducting an electron through temporary collisional molecular orbital. In trypsin model experiment, our sensor showed ~9-fold fluorescence increasing after hydrolysis; in Caspase-3 experiment, our sensor also showed significant fluorescence increasing. Finally, the application of tryptophan as a natural quencher in peptidyl fluorescent sensor is feasible and potential.
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參考文獻:
(1) Elmore, S. Toxicologic Pathology. 2007, 35, 495.
(2) Peterson, Q. P.; Goode, D. R.; West, D. C.; Botham, R. C.; Hergenrother, P.
J. Nature Protocols. 2010, 5, 294.
(3) Majno, G.; Joris, I. American Journal of Pathology. 1995, 146, 3.
(4) Janicke, R. U.; Sprengart, M. L.; Wati, M. R.; Porter, A. G. Journal of
Biological Chemistry 1998, 273, 9357.
(5) Cohen, G. M. Biochemical Journal. 1997, 326, 1.
(6) Fadok, V. A.; Savill, J. S.; Haslett, C.; Bratton, D. L.; Doherty, D. E.;
Campbell, P. A.; Henson, P. M. Journal of Immunology 1992, 149, 4029.
(7) Smith, C. A.; Farrah, T.; Goodwin, R. G. Cell 1994, 76, 959.
(8) Ashkenazi, A.; Dixit, V. M. Science 1998, 281, 1305.
(9) Green, D. R.; Reed, J. C. Science 1998, 281, 1309.
(10) Fulda, S.; Debatin, K. M. Oncogene 2006, 25, 4798.
(11) Liu, X. S.; Kim, C. N.; Yang, J.; Jemmerson, R.; Wang, X. D. Cell 1996, 86,
147.
(12) Zou, H.; Henzel, W. J.; Liu, X. S.; Lutschg, A.; Wang, X. D. Cell 1997, 90,
405.
(13) Yang, J.; Liu, X. S.; Bhalla, K.; Kim, C. N.; Ibrado, A. M.; Cai, J. Y.; Peng,
T. I.; Jones, D. P.; Wang, X. D. Science 1997, 275, 1129.
(14) Aas, T.; Borresen, A. L.; Geisler, S.; SmithSorensen, B.; Johnsen, H.;
Varhaug, J. E.; Akslen, L. A.; Lonning, P. E. Nature Medicine 1996, 2, 811.
(15) Wang, S. W.; Konorev, E. A.; Kotamraju, S.; Joseph, J.; Kalivendi, S.;
Kalyanaraman, B. Journal of Biological Chemistry 2004, 279, 25535.
(16) Nicolier, M.; Decrion-Barthod, A.-Z.; Launay, S.; Pretet, J.-L.; Mougin, C.
Biology of the Cell 2009, 101, 455. 66
(17) Belmokhtar, C. A.; Hillion, J.; Segal-Bendirdjian, E. Oncogene 2001, 20,
3354.
(18) Chae, H.-J.; Kang, J.-S.; Byun, J.-O.; Han, K.-S.; Kim, D.-U.; Oh, S.-M.;
Kim, H.-M.; Chae, S.-W.; Kim, H.-R. Pharmacological Research 2000, 42, 373.
(19) Pop, C.; Salvesen, G. S. Journal of Biological Chemistry 2009, 284, 21777.
(20) Johansson, M. K.; Cook, R. M. Chemistry-a European Journal. 2003, 9,
3466.
(21) Ward, M. D. Chemical Society Reviews. 1997, 26, 365.
(22) Torimura, M.; Kurata, S.; Yamada, K.; Yokomaku, T.; Kamagata, Y.;
Kanagawa, T.; Kurane, R. Analytical sciences : the international journal of the Japan
Society for Analytical Chemistry 2001, 17, 155.
(23) de Silva, A. P.; Moody, T. S.; Wright, G. D. Analyst 2009, 134, 2385.
(24) Doose, S.; Neuweiler, H.; Sauer, M. ChemPhysChem 2005, 6, 2277.
(25) Doose, S.; Neuweiler, H.; Sauer, M. ChemPhysChem 2009, 10, 1389.
(26) Zhang, J.; Campbell, R. E.; Ting, A. Y.; Tsien, R. Y. Nature reviews.
Molecular cell biology 2002, 3, 906.
(27) Hoffmann, C.; Gaietta, G.; Bunemann, M.; Adams, S. R.; Oberdorff-Maass,
S.; Behr, B.; Vilardaga, J. P.; Tsien, R. Y.; Eisman, M. H.; Lohse, M. J. Nature
Methods. 2005, 2, 171.
(28) Thornberry, N. A.; Bull, H. G.; Calaycay, J. R.; Chapman, K. T.; Howard, A.
D.; Kostura, M. J.; Miller, D. K.; Molineaux, S. M.; Weidner, J. R.; Aunins, J.;
Elliston, K. O.; Ayala, J. M.; Casano, F. J.; Chin, J.; Ding, G. J. F.; Egger, L. A.;
Gaffney, E. P.; Limjuco, G.; Palyha, O. C.; Raju, S. M.; Rolando, A. M.; Salley, J. P.;
Yamin, T. T.; Lee, T. D.; Shively, J. E.; Maccross, M.; Mumford, R. A.; Schmidt, J. A.;
Tocci, M. J. Nature 1992, 356, 768.
(29) Rotonda, J.; Nicholson, D. W.; Fazil, K. M.; Gallant, M.; Gareau, Y.; 67
Labelle, M.; Peterson, E. P.; Rasper, D. M.; Ruel, R.; Vaillancourt, J. P.; Thornberry, N.
A.; Becker, J. W. Nature structural biology 1996, 3, 619.
(30) Han, Z. Y.; Hendrickson, E. A.; Bremner, T. A.; Wyche, J. H. Journal of
Biological Chemistry 1997, 272, 13432.
(31) Stennicke, H. R.; Renatus, M.; Meldal, M.; Salvesen, G. S. The Biochemical
journal 2000, 350 Pt 2, 563.
(32) Tewari, M.; Quan, L. T.; Orourke, K.; Desnoyers, S.; Zeng, Z.; Beidler, D.
R.; Poirier, G. G.; Salvesen, G. S.; Dixit, V. M. Cell 1995, 81, 801.
(33) Roy, A. C.; Saha, N.; Tan, S. M.; Kamarul, F. Z.; Ratnam, S. S.
Electrophoresis 1992, 13, 396.
(34) Gurtu, V.; Kain, S. R.; Zhang, G. Analytical Biochemistry. 1997, 251, 98.
(35) Liu, J.; Bhalgat, M.; Zhang, C.; Diwu, Z.; Hoyland, B.; Klaubert, D. H.
Bioorganic & Medicinal Chemistry Letters. 1999, 9, 3231.
(36) Wang, Z.-Q.; Liao, J.; Diwu, Z. Bioorganic & Medicinal Chemistry Letters
2005, 15, 2335.
(37) Cai, S. X.; Zhang, H.-Z.; Guastella, J.; Drewe, J.; Yang, W.; Weber, E.
Bioorganic & Medicinal Chemistry Letters 2001, 11, 39.
(38) Bullok, K.; Piwnica-Worms, D. Journal of Medicinal Chemistry. 2005, 48,
5404.
(39) Bullok, K. E.; Maxwell, D.; Kesarwala, A. H.; Gammon, S.; Prior, J. L.;
Snow, M.; Stanley, S.; Piwnica-Worms, D. Biochemistry 2007, 46, 4055.
(40) Maxwell, D.; Chang, Q.; Zhang, X.; Barnett, E. M.; Piwnica-Worms, D.
Bioconjugate Chem. 2009, 20, 702.
(41) Boeneman, K.; Mei, B. C.; Dennis, A. M.; Bao, G.; Deschamps, J. R.;
Mattoussi, H.; Medintz, I. L. Journal of the American Chemical Society 2009, 131,
3828. 68
(42) Prasuhn, D. E.; Feltz, A.; Blanco-Canosa, J. B.; Susumu, K.; Stewart, M. H.;
Mei, B. C.; Yakovlev, A. V.; Loukov, C.; Mallet, J.-M.; Oheim, M.; Dawson, P. E.;
Medintz, I. L. Acs Nano 2010, 4, 5487.
(43) Kihara, T.; Nakamura, C.; Suzuki, M.; Han, S. W.; Fukazawa, K.; Ishihara,
K.; Miyake, J. Biosensors & Bioelectronics. 2009, 25, 22.
(44) Shi, Y.; Yi, C.; Zhang, Z.; Zhang, H.; Li, M.; Yang, M.; Jiang, Q. ACS
applied materials & interfaces 2013, 5, 6494.
(45) Xu, X.; Gerard, A. L. V.; Huang, B. C. B.; Anderson, D. C.; Payan, D. G.;
Luo, Y. Nucleic Acids Research. 1998, 26, 2034.
(46) Nagai, T.; Miyawaki, A. Biochemical and Biophysical Research
Communications 2004, 319, 72.
(47) Elphick, L. M.; Meinander, A.; Mikhailov, A.; Richard, M.; Toms, N. J.;
Eriksson, J. E.; Kass, G. E. N. Analytical Biochemistry. 2006, 349, 148.
(48) Sakamoto, S.; Terauchi, M.; Hugo, A.; Kim, T.; Araki, Y.; Wada, T.
Chemical Communications. 2013, 49, 10323.
(49) Hong, Y. N.; Lam, J. W. Y.; Tang, B. Z. Chemical Communications.. 2009,
4332.
(50) Shi, H.; Kwok, R. T. K.; Liu, J.; Xing, B.; Tang, B. Z.; Liu, B. Journal of
the American Chemical Society 2012, 134, 17972.
(51) Shi, H. B.; Zhao, N.; Ding, D.; Liang, J.; Tang, B. Z.; Liu, B. Organic &
Biomolecular Chemistry 2013, 11, 7289.
(52) Ding, D.; Liang, J.; Shi, H. B.; Kwok, R. T. K.; Gao, M.; Feng, G. X.; Yuan,
Y. Y.; Tang, B. Z.; Liu, B. Journal of Materials Chemistry B 2014, 2, 231.
(53) Marme, N.; Knemeyer, J. P.; Sauer, M.; Wolfrum, J. Bioconjugate
Chemistry. 2003, 14, 1133.
(54) Marme, N.; Knemeyer, J. P.; Wolfrum, J.; Sauer, M. Angewandte Chemie-International Edition. 2004, 43, 3798.
(55) Togashi, D. M.; Szczupak, B.; Ryder, A. G.; Calvet, A.; O'Loughlin, M.
Journal of Physical Chemistry A. 2009, 113, 2757.
(56) Albert Griffin, B.; Adams, S. R.; Jones, J.; Tsien, R. Y. In Methods in
Enzymology; Jeremy Thorner, S. D. E., John, N. A., Eds.; Academic Press: 2000; Vol.
Volume 327, p 565.