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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 陳振中(Chun-Chung Chan) | |
| dc.contributor.author | Yu-Ting Chang | en |
| dc.contributor.author | 張宇廷 | zh_TW |
| dc.date.accessioned | 2021-06-17T07:23:26Z | - |
| dc.date.available | 2021-07-15 | |
| dc.date.copyright | 2019-07-15 | |
| dc.date.issued | 2019 | |
| dc.date.submitted | 2019-07-01 | |
| dc.identifier.citation | 1. Lodish, H.; Berk, A.; Kaiser, C.; Krieger, M.; Bretscher, A.; Ploegh, H.; Amon, A.; Martin, K., Molecular Cell Biology. W.H. Freeman 2016.
2. Wallin, E.; von Heijne, G., Genome-wide analysis of integral membrane proteins from eubacterial, archaean, and eukaryotic organisms. Protein Sci 1998, 7 (4), 1029-1038. 3. Yildirim, M. A.; Goh, K. I.; Cusick, M. E.; Barabasi, A. L.; Vidal, M., Drug-target network. Nature biotechnology 2007, 25 (10), 1119-26. 4. Berman, H. M.; Westbrook, J.; Feng, Z.; Gilliland, G.; Bhat, T. N.; Weissig, H.; Shindyalov, I. N.; Bourne, P. E., The Protein Data Bank. Nucleic Acids Res 2000, 28 (1), 235-242. 5. Levy, S.; Todd, S. C.; Maecker, H. T., CD81 (TAPA-1): a molecule involved in signal transduction and cell adhesion in the immune system. Annual review of immunology 1998, 16, 89-109. 6. Bracey, M. H.; Cravatt, B. F.; Stevens, R. C., Structural commonalities among integral membrane enzymes. FEBS letters 2004, 567 (2-3), 159-65. 7. Popot, J. L., MEMBRANE PROTEINS IN AQUEOUS SOLUTIONS: from Detergents to Amphipols. SPRINGER 2019. 8. Dahl, J. L.; Hokin, L. E., The sodium-potassium adenosinetriphosphatase. Annu Rev Biochem 1974, 43 (0), 327-56. 9. Wickstrand, C.; Dods, R.; Royant, A.; Neutze, R., Bacteriorhodopsin: Would the real structural intermediates please stand up? Biochimica et biophysica acta 2015, 1850 (3), 536-53. 10. Emma, P.; Akre, R.; Arthur, J.; Bionta, R.; Bostedt, C.; Bozek, J.; Brachmann, A.; Bucksbaum, P.; Coffee, R.; Decker, F. J.; Ding, Y.; Dowell, D.; Edstrom, S.; Fisher, A.; Frisch, J.; Gilevich, S.; Hastings, J.; Hays, G.; Hering, P.; Huang, Z.; Iverson, R.; Loos, H.; Messerschmidt, M.; Miahnahri, A.; Moeller, S.; Nuhn, H. D.; Pile, G.; Ratner, D.; Rzepiela, J.; Schultz, D.; Smith, T.; Stefan, P.; Tompkins, H.; Turner, J.; Welch, J.; White, W.; Wu, J.; Yocky, G.; Galayda, J., First lasing and operation of an angstrom-wavelength free-electron laser. Nat Photonics 2010, 4 (9), 641-647. 11. Neutze, R.; Branden, G.; Schertler, G. F., Membrane protein structural biology using X-ray free electron lasers. Current opinion in structural biology 2015, 33, 115-25. 12. Wickstrand, C.; Nogly, P.; Nango, E.; Iwata, S.; Standfuss, J.; Neutze, R., Bacteriorhodopsin: structural insights revealed using X-ray lasers and synchrotron radiation. Annual review of biochemistry 2019, 88. 13. Andersson, M.; Malmerberg, E.; Westenhoff, S.; Katona, G.; Cammarata, M.; Wohri, A. B.; Johansson, L. C.; Ewald, F.; Eklund, M.; Wulff, M.; Davidsson, J.; Neutze, R., Structural Dynamics of Light-Driven Proton Pumps. Biophysical journal 2010, 98 (3), 226a-226a. 14. Koch, M. H. J.; Dencher, N. A.; Oesterhelt, D.; Plohn, H. J.; Rapp, G.; Buldt, G., Time-Resolved X-Ray-Diffraction Study of Structural-Changes Associated with the Photocycle of Bacteriorhodopsin. Embo Journal 1991, 10 (3), 521-526. 15. Kamikubo, H.; Oka, T.; Imamoto, Y.; Tokunaga, F.; Lanyi, J. K.; Kataoka, M., The last phase of the reprotonation switch in bacteriorhodopsin: The transition between the M-type and the N-type protein conformation depends on hydration. Biochemistry 1997, 36 (40), 12282-12287. 16. Oka, T.; Kamikubo, H.; Tokunaga, F.; Lanyi, J. K.; Needleman, R.; Kataoka, M., Conformational change of helix G in the bacteriorhodopsin photocycle: Investigation with heavy atom labeling and X-ray diffraction. Biophysical journal 1999, 76 (2), 1018-1023. 17. Dencher, N. A.; Dresselhaus, D.; Zaccai, G.; Buldt, G., Structural-Changes in Bacteriorhodopsin during Proton Translocation Revealed by Neutron-Diffraction. Proceedings of the National Academy of Sciences of the United States of America 1989, 86 (20), 7876-7879. 18. Subramaniam, S.; Gerstein, M.; Oesterhelt, D.; Henderson, R., Electron-Diffraction Analysis of Structural-Changes in the Photocycle of Bacteriorhodopsin. Embo Journal 1993, 12 (1), 1-8. 19. Oesterhelt, D.; Stoeckenius, W., Functions of a new photoreceptor membrane. Proceedings of the National Academy of Sciences of the United States of America 1973, 70 (10), 2853-7. 20. Brzezinski, P.; Gennis, R. B., Cytochrome c oxidase: exciting progress and remaining mysteries. Journal of bioenergetics and biomembranes 2008, 40 (5), 521-31. 21. Walker, J. E., The ATP synthase: the understood, the uncertain and the unknown. Biochemical Society transactions 2013, 41 (1), 1-16. 22. Saidijam, M.; Bettaney, K. E.; Szakonyi, G.; Psakis, G.; Shibayama, K.; Suzuki, S.; Clough, J. L.; Blessie, V.; Abu-Bakr, A.; Baumberg, S.; Meuller, J.; Hoyle, C. K.; Palmer, S. L.; Butaye, P.; Walravens, K.; Patching, S. G.; O'Reilly, J.; Rutherford, N. G.; Bill, R. M.; Roper, D. I.; Phillips-Jones, M. K.; Henderson, P. J., Active membrane transport and receptor proteins from bacteria. Biochemical Society transactions 2005, 33 (Pt 4), 867-72. 23. Mizobe, T.; Maze, M.; Lam, V.; Suryanarayana, S.; Kobilka, B. K., Arrangement of transmembrane domains in adrenergic receptors - Similarity to bacteriorhodopsin. Journal of Biological Chemistry 1996, 271 (5), 2387-2389. 24. Henderson, R.; Unwin, P. N. T., Three-dimensional model of purple membrane obtained by electron microscopy. Nature 1975, 257 (5521), 28. 25. Henderson, R.; Baldwin, J. M.; Ceska, T. A.; Zemlin, F.; Beckmann, E.; Downing, K. H., Model for the structure of bacteriorhodopsin based on high-resolution electron cryo-microscopy. Journal of molecular biology 1990, 213 (4), 899-929. 26. Landau, E. M.; Rosenbusch, J. P., Lipidic cubic phases: A novel concept for the crystallization of membrane proteins. Proceedings of the National Academy of Sciences of the United States of America 1996, 93 (25), 14532-14535. 27. Edman, K.; Nollert, P.; Royant, A.; Belrhali, H.; Pebay-Peyroula, E.; Hajdu, J.; Neutze, R.; Landau, E. M., High-resolution X-ray structure of an early intermediate in the bacteriorhodopsin photocycle. Nature 1999, 401 (6755), 822-6. 28. Royant, A.; Edman, K.; Ursby, T.; Pebay-Peyroula, E.; Landau, E. M.; Neutze, R., Helix deformation is coupled to vectorial proton transport in the photocycle of bacteriorhodopsin. Nature 2000, 406 (6796), 645-648. 29. Luecke, H.; Schobert, B.; Richter, H. T.; Cartailler, J. P.; Lanyi, J. K., Structural changes in bacteriorhodopsin during ion transport at 2 Angstrom resolution. Science 1999, 286 (5438), 255-260. 30. Luecke, H.; Schobert, B.; Cartailler, J. P.; Richter, H. T.; Rosengarth, A.; Needleman, R.; Lanyi, J. K., Coupling photoisomerization of retinal to directional transport in bacteriorhodopsin. Journal of molecular biology 2000, 300 (5), 1237-1255. 31. Sass, H. J.; Buldt, G.; Gessenich, R.; Hehn, D.; Neff, D.; Schlesinger, R.; Berendzen, J.; Ormos, P., Structural alterations for proton translocation in the M state of wild-type bacteriorhodopsin. Nature 2000, 406 (6796), 649-53. 32. Vonck, J., Structure of the bacteriorhodopsin mutant F219L N intermediate revealed by electron crystallography. The EMBO journal 2000, 19 (10), 2152-60. 33. Kühlbrandt, W., Bacteriorhodopsin—the movie. Nature 2000, 406 (6796), 569. 34. Ohno, K.; Takeuchi, Y.; Yoshida, M., Effect of light-adaptation on the photoreaction of bacteriorhodopsin from Halobacterium halobium. Biochimica et Biophysica Acta (BBA)-Bioenergetics 1977, 462 (3), 575-582. 35. Subramaniam, S.; Henderson, R., Molecular mechanism of vectorial proton translocation by bacteriorhodopsin. Nature 2000, 406 (6796), 653-7. 36. Ginzburg, M., Ion Metabolism in a Halobacterium: I. Influence of age of culture on intracellular concentrations. The Journal of General Physiology 1970, 55 (2), 187-207. 37. Baliga, N. S.; Bonneau, R.; Facciotti, M. T.; Pan, M.; Glusman, G.; Deutsch, E. W.; Shannon, P.; Chiu, Y. L.; Gan, R. R.; Hung, P. L.; Date, S. V.; Marcotte, E.; Hood, L.; Ng, W. V., Genome sequence of Haloarcula marismortui: A halophilic archaeon from the Dead Sea. Genome research 2004, 14 (11), 2221-2234. 38. Hsu, M. F.; Yu, T. F.; Chou, C. C.; Fu, H. Y.; Yang, C. S.; Wang, A. H., Using Haloarcula marismortui bacteriorhodopsin as a fusion tag for enhancing and visible expression of integral membrane proteins in Escherichia coli. PloS one 2013, 8 (2), e56363. 39. Shevchenko, V.; Gushchin, I.; Polovinkin, V.; Round, E.; Borshchevskiy, V.; Utrobin, P.; Popov, A.; Balandin, T.; Buldt, G.; Gordeliy, V., Crystal structure of Escherichia coli-expressed Haloarcula marismortui bacteriorhodopsin I in the trimeric form. PloS one 2014, 9 (12), e112873. 40. Tsai, F. K.; Fu, H. Y.; Yang, C. S.; Chu, L. K., Photochemistry of a dual-bacteriorhodopsin system in Haloarcula marismortui: HmbRI and HmbRII. The journal of physical chemistry. B 2014, 118 (26), 7290-301. 41. Yeh, V., Study of Bacteriorhodopsin in a Controlled Lipid Environment. Springer: 2018. 42. Cai, M.; Huang, Y.; Sakaguchi, K.; Clore, G. M.; Gronenborn, A. M.; Craigie, R., An efficient and cost-effective isotope labeling protocol for proteins expressed in shape Escherichia coli. Journal of biomolecular NMR 1998, 11 (1), 97-102. 43. Kay, L.; Keifer, P.; Saarinen, T., Pure absorption gradient enhanced heteronuclear single quantum correlation spectroscopy with improved sensitivity. Journal of the American Chemical Society 1992, 114 (26), 10663-10665. 44. Delaglio, F.; Grzesiek, S.; Vuister, G. W.; Zhu, G.; Pfeifer, J.; Bax, A., NMRPipe: a multidimensional spectral processing system based on UNIX pipes. Journal of biomolecular NMR 1995, 6 (3), 277-293. 45. Dencher, N. A.; Heyn, M. P., [2] Preparation and properties of monomeric bacteriorhodopsin. In Methods in enzymology, Elsevier: 1982; Vol. 88, pp 5-10. 46. Wang, J.; Link, S.; Heyes, C. D.; El-Sayed, M. A., Comparison of the dynamics of the primary events of bacteriorhodopsin in its trimeric and monomeric states. Biophysical journal 2002, 83 (3), 1557-66. 47. Chaptal, V.; Delolme, F.; Kilburg, A.; Magnard, S.; Montigny, C.; Picard, M.; Prier, C.; Monticelli, L.; Bornert, O.; Agez, M.; Ravaud, S.; Orelle, C.; Wagner, R.; Jawhari, A.; Broutin, I.; Pebay-Peyroula, E.; Jault, J. M.; Kaback, H. R.; le Maire, M.; Falson, P., Quantification of Detergents Complexed with Membrane Proteins. Scientific reports 2017, 7, 41751. 48. Seigneuret, M.; Neumann, J.-M.; Rigaud, J.-L., Detergent delipidation and solubilization strategies for high-resolution NMR of the membrane protein bacteriorhodopsin. Journal of Biological Chemistry 1991, 266 (16), 10066-10069. 49. Müller, D. J.; Kessler, M.; Oesterhelt, F.; Möller, C.; Oesterhelt, D.; Gaub, H., Stability of bacteriorhodopsin α-helices and loops analyzed by single-molecule force spectroscopy. Biophysical journal 2002, 83 (6), 3578-3588. 50. Kelly, A. E.; Ou, H. D.; Withers, R.; Dötsch, V., Low-conductivity buffers for high-sensitivity NMR measurements. Journal of the American Chemical Society 2002, 124 (40), 12013-12019. 51. Milder, S. J.; Thorgeirsson, T. E.; Miercke, L. J.; Stroud, R. M.; Kliger, D. S., Effects of detergent environments on the photocycle of purified monomeric bacteriorhodopsin. Biochemistry 1991, 30 (7), 1751-61. 52. Fu, H. Y.; Yi, H. P.; Lu, Y. H.; Yang, C. S., Insight into a single halobacterium using a dual-bacteriorhodopsin system with different functionally optimized pH ranges to cope with periplasmic pH changes associated with continuous light illumination. Molecular microbiology 2013, 88 (3), 551-61. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/73226 | - |
| dc.description.abstract | 膜蛋白細菌視紫質在吸收綠光之後，產生一系列構形變化，將氫離子運輸至細胞膜外，並在細胞膜內外產生氫離子梯度，最終轉換為生物可利用能量三磷酸腺苷。膜蛋白的研究雖然重要，但是因為樣品製備等問題，造成研究上的困難。在降低了純化用緩衝溶液的酸鹼度以及加入了離子交換層析法之後，不只提升了純化步驟的產率，更使單體與三聚體的分離更有效率。我們更發現單體以及三聚體的比例可以透過人為的方式進行調控。經過介面活性劑與緩衝溶液種類的篩選，我們得到不錯的氫氮二維異核化學位移相關光譜。我們測量了樣品亮適應態、激發態的動力學數據以及光強度對亮適應的影響。將以上數據與四條光纖以及兩根核磁共振樣品管結合後，我們在原位液態核磁共振實驗中觀察到了亮適應態與暗適應態的差異。 | zh_TW |
| dc.description.abstract | A proton pump that builds up a proton gradient across a biological membrane is a membrane protein. A typical example is Haloarcula marismortui bacteriorhodopsin (HmbRI) that achieves its functionality as a proton pump by changing its conformation in the presence of green light. The study of the structural changes in membrane protein is crucial but difficult because of the problems such as sample preparation. In this thesis, we report that the purification of monomeric D94N HmbRI was improved by lowering the pH of the purification buffers and including an additional step of ionic-exchange chromatography, which can successfully and efficiently separate the monomeric and trimeric form of D94N HmbRI. Moreover, we found that the ratio between monomer and trimer can be manually controlled. With extensive detergent and buffer screening, we were able to obtain the 1H-15N TROSY HSQC spectrum of D94N HmbRI with good quality. By incorporating four optical fibers into a double layer NMR tube, the in-situ light coupling solution state NMR measurements were conducted. The lifetime of the light-adapted state and the M-state decay of D94N HmbRI was measured. The correlation between light intensity and the duration of light adaptation was further investigated. Using this device, the differences between the 1H-15N TROSY HSQC spectra for the dark-adapted state and the light-adapted state of D94N HmbRI were observed. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-17T07:23:26Z (GMT). No. of bitstreams: 1 ntu-108-R06223171-1.pdf: 2685455 bytes, checksum: a768cc3271444af4dc58d6a7e0022d4c (MD5) Previous issue date: 2019 | en |
| dc.description.tableofcontents | 1. Introduction 1
1.1 Membrane Protein 1 1.2 Bacteriorhodopsin 5 1.2.1 Halobacterium salinarum Bacteriorhodopsin 5 1.2.2 Haloarcula marismortui Bacteriorhodopsin 9 1.3 Motivation and Specific aim of Thesis 11 2. Experimental background 12 2.1 Size Exclusion Chromatography 12 2.2 Ionic-Exchange Chromatography 13 2.3 Steady-state UV/Vis Absorption Spectroscopy 15 2.4 Transient Absorption Spectroscopy 19 2.5 Solution-state NMR spectroscopy 20 3. Materials and Methods 26 3.1 Sample Preparation 26 3.1.1 DNA Sequence 26 3.1.2 Transformation 27 3.1.3 Protein expression 28 3.1.4 Protein purification 29 3.1.5 Detergent screening 32 3.1.6 Buffer screening 33 3.2 Sample Characterization 34 3.2.1 SDS PAGE 34 3.2.2 Matrix Assisted Laser Desorption Ionization Time of Flight (MALDI-TOF) Mass spectroscopy 34 3.2.3 Steady-state UV-VIS Absorption Spectroscopy measurement of dark adaptation 35 3.2.4 Transient Absorption Spectroscopy 35 3.2.5 Steady-state UV-VIS Absorption Spectroscopy measurement of light adaptation 36 3.2.6 Solution-state NMR 37 3.2.7 Solution-state light coupling NMR experiments 37 4. Results and Discussion 41 4.1 Sample Preparation 41 4.1.1 Protein expression and purification 41 4.1.2 Detergent screening 48 4.1.3 Buffer screening 53 4.2 The lifetime of the light-adapted state 54 4.3 M state decay 57 4.4 Light intensity influence on light adaptation 59 4.5 The 1H-15N TROSY HSQC difference between the light and dark-adapted state 61 5. Conclusions 66 6. Reference 68 | |
| dc.language.iso | en | |
| dc.subject | 膜蛋白 | zh_TW |
| dc.subject | 細菌視紫質 | zh_TW |
| dc.subject | 原位核磁共振 | zh_TW |
| dc.subject | 亮/暗適應 | zh_TW |
| dc.subject | 蛋白質構型變化 | zh_TW |
| dc.subject | protein conformational change | en |
| dc.subject | Membrane protein | en |
| dc.subject | bacteriorhodopsin | en |
| dc.subject | in-situ NMR | en |
| dc.subject | light/dark adaptation | en |
| dc.title | 以核磁共振研究細菌視紫質之暗與光適應態之結構差異 | zh_TW |
| dc.title | In-situ NMR characterization of the difference between the light and dark-adapted state of D94N HmbRI | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 107-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 余慈顏(Tsyr-Yan Yu) | |
| dc.contributor.oralexamcommittee | Daniel Nietlispach(Daniel Nietlispach),徐駿森(Chun-Hua Hsu) | |
| dc.subject.keyword | 膜蛋白,細菌視紫質,原位核磁共振,亮/暗適應,蛋白質構型變化, | zh_TW |
| dc.subject.keyword | Membrane protein,bacteriorhodopsin,in-situ NMR,light/dark adaptation,protein conformational change, | en |
| dc.relation.page | 74 | |
| dc.identifier.doi | 10.6342/NTU201901165 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2019-07-02 | |
| dc.contributor.author-college | 理學院 | zh_TW |
| dc.contributor.author-dept | 化學研究所 | zh_TW |
| 顯示於系所單位： | 化學系 | |
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