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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 楊啟伸 | |
| dc.contributor.author | Hsu-Yuan Fu | en |
| dc.contributor.author | 傅煦媛 | zh_TW |
| dc.date.accessioned | 2021-06-14T17:14:27Z | - |
| dc.date.available | 2018-07-24 | |
| dc.date.copyright | 2008-08-05 | |
| dc.date.issued | 2008 | |
| dc.date.submitted | 2008-07-25 | |
| dc.identifier.citation | 1. Spudich, W.R.B.a.J.L. Handbook of Photosensory Receptors (2005).
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Structure-function studies on bacteriorhodopsin. IV. Purification and renaturation of bacterio-opsin polypeptide expressed in Escherichia coli. J Biol Chem 262, 9271-6 (1987). 8. Mogi, T., Marti, T. & Khorana, H.G. Structure-function studies on bacteriorhodopsin. IX. Substitutions of tryptophan residues affect protein-retinal interactions in bacteriorhodopsin. J Biol Chem 264, 14197-201 (1989). 9. Hackett, N.R., Stern, L.J., Chao, B.H., Kronis, K.A. & Khorana, H.G. Structure-function studies on bacteriorhodopsin. V. Effects of amino acid substitutions in the putative helix F. J Biol Chem 262, 9277-84 (1987). 10. Mogi, T., Stern, L.J., Chao, B.H. & Khorana, H.G. Structure-function studies on bacteriorhodopsin. VIII. Substitutions of the membrane-embedded prolines 50, 91, and 186: the effects are determined by the substituting amino acids. J Biol Chem 264, 14192-6 (1989). 11. Stern, L.J. & Khorana, H.G. Structure-function studies on bacteriorhodopsin. X. Individual substitutions of arginine residues by glutamine affect chromophore formation, photocycle, and proton translocation. J Biol Chem 264, 14202-8 (1989). 12. Hohenfeld, I.P., Wegener, A.A. & Engelhard, M. Purification of histidine tagged bacteriorhodopsin, pharaonis halorhodopsin and pharaonis sensory rhodopsin II functionally expressed in Escherichia coli. FEBS Lett 442, 198-202 (1999). 13. Schmies, G., Chizhov, I. & Engelhard, M. Functional expression of His-tagged sensory rhodopsin I in Escherichia coli. FEBS Lett 466, 67-9 (2000). 14. Heymann, J., Jager, R. & Subramaniam, S. Expression of bacteriorhodopsin in Sf9 and COS-1 cells. J Bioenerg Biomembr 29, 55-9 (1997). 15. Seki, A. et al. Heterologous expression of Pharaonis halorhodopsin in Xenopus laevis oocytes and electrophysiological characterization of its light-driven Cl- pump activity. Biophys J 92, 2559-69 (2007). 16. Kalmbach, R. et al. Functional cell-free synthesis of a seven helix membrane protein: in situ insertion of bacteriorhodopsin into liposomes. J Mol Biol 371, 639-48 (2007). 17. Krebs, M.P., Mollaaghababa, R. & Khorana, H.G. Gene replacement in Halobacterium halobium and expression of bacteriorhodopsin mutants. Proc Natl Acad Sci U S A 90, 1987-91 (1993). 18. Shand, R.F. & Betlach, M.C. bop gene cluster expression in bacteriorhodopsin-overproducing mutants of Halobacterium halobium. J Bacteriol 176, 1655-60 (1994). 19. Otomo, J. & Muramatsu, T. Over-expression of a new photo-active halorhodopsin in Halobacterium salinarium. Biochim Biophys Acta 1240, 248-56 (1995). 20. Chizhov, I. & Engelhard, M. Temperature and halide dependence of the photocycle of halorhodopsin from Natronobacterium pharaonis. Biophys J 81, 1600-12 (2001). 21. Chizhov, I. et al. The photophobic receptor from Natronobacterium pharaonis: temperature and pH dependencies of the photocycle of sensory rhodopsin II. Biophys J 75, 999-1009 (1998). 22. Wang, W.W., Sineshchekov, O.A., Spudich, E.N. & Spudich, J.L. Spectroscopic and photochemical characterization of a deep ocean proteorhodopsin. J Biol Chem 278, 33985-91 (2003). 23. Sudo, Y. & Spudich, J.L. Three strategically placed hydrogen-bonding residues convert a proton pump into a sensory receptor. Proc Natl Acad Sci U S A 103, 16129-34 (2006). 24. Falke, J.J., Bass, R.B., Butler, S.L., Chervitz, S.A. & Danielson, M.A. The two-component signaling pathway of bacterial chemotaxis: a molecular view of signal transduction by receptors, kinases, and adaptation enzymes. Annu Rev Cell Dev Biol 13, 457-512 (1997). 25. Hoff, W.D., Jung, K.H. & Spudich, J.L. Molecular mechanism of photosignaling by archaeal sensory rhodopsins. Annu Rev Biophys Biomol Struct 26, 223-58 (1997). 26. Pebay-Peyroula, E., Rummel, G., Rosenbusch, J.P. & Landau, E.M. X-ray structure of bacteriorhodopsin at 2.5 angstroms from microcrystals grown in lipidic cubic phases. Science 277, 1676-81 (1997). 27. Kolbe, M., Besir, H., Essen, L.O. & Oesterhelt, D. Structure of the light-driven chloride pump halorhodopsin at 1.8 A resolution. Science 288, 1390-6 (2000). 28. Luecke, H., Schobert, B., Lanyi, J.K., Spudich, E.N. & Spudich, J.L. Crystal structure of sensory rhodopsin II at 2.4 angstroms: insights into color tuning and transducer interaction. Science 293, 1499-503 (2001). 29. Gordeliy, V.I. et al. Molecular basis of transmembrane signalling by sensory rhodopsin II-transducer complex. Nature 419, 484-7 (2002). 30. Sasaki, J. et al. Conversion of bacteriorhodopsin into a chloride ion pump. Science 269, 73-5 (1995). 31. Bogomolni, R.A. et al. Removal of transducer HtrI allows electrogenic proton translocation by sensory rhodopsin I. Proc Natl Acad Sci U S A 91, 10188-92 (1994). 32. Schmies, G., Engelhard, M., Wood, P.G., Nagel, G. & Bamberg, E. Electrophysiological characterization of specific interactions between bacterial sensory rhodopsins and their transducers. Proc Natl Acad Sci U S A 98, 1555-9 (2001). 33. Sharma, A.K., Spudich, J.L. & Doolittle, W.F. Microbial rhodopsins: functional versatility and genetic mobility. Trends Microbiol 14, 463-9 (2006). 34. Grisshammer, R. Understanding recombinant expression of membrane proteins. Curr Opin Biotechnol 17, 337-40 (2006). 35. Charlebois, R.L., Lam, W.L., Cline, S.W. & Doolittle, W.F. Characterization of pHV2 from Halobacterium volcanii and its use in demonstrating transformation of an archaebacterium. Proc Natl Acad Sci U S A 84, 8530-4 (1987). 36. Lam, W.L. & Doolittle, W.F. Shuttle vectors for the archaebacterium Halobacterium volcanii. Proc Natl Acad Sci U S A 86, 5478-82 (1989). 37. Sasaki, J. et al. Different dark conformations function in color-sensitive photosignaling by the sensory rhodopsin I-HtrI complex. Biophys J 92, 4045-53 (2007). 38. Sharma, A.K. et al. Evolution of rhodopsin ion pumps in haloarchaea. BMC Evol Biol 7, 79 (2007). 39. Kehoe, D.M. & Gutu, A. Responding to color: the regulation of complementary chromatic adaptation. Annu Rev Plant Biol 57, 127-50 (2006). 40. Hirokawa, T., Boon-Chieng, S. & Mitaku, S. SOSUI: classification and secondary structure prediction system for membrane proteins. Bioinformatics 14, 378-9 (1998). 41. Peitsch, M.C. ProMod and Swiss-Model: Internet-based tools for automated comparative protein modelling. Biochem Soc Trans 24, 274-9 (1996). 42. Luecke, H., Schobert, B., Richter, H.T., Cartailler, J.P. & Lanyi, J.K. Structure of bacteriorhodopsin at 1.55 A resolution. J Mol Biol 291, 899-911 (1999). 43. Royant, A. et al. X-ray structure of sensory rhodopsin II at 2.1-A resolution. Proc Natl Acad Sci U S A 98, 10131-6 (2001). | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/41059 | - |
| dc.description.abstract | 在微生物中，光感受體 (photoreceptor) 對環境光照刺激反應，扮演十分重要的角色。目前為止，已經有超過 100 種的 photoreceptors 被鑑定出來。科學研究將它們分成四大類：包括 bacteriorhodopsin (BR)，其功能是將細胞內的氫離子運送到細胞外；halorhodopsin (HR) 則是將細胞外的氯離子運送到細胞內；sensory rhodopsin I (SRI) 同時具有調控正趨光及負趨光性；以及對於接近 UV 波長之光源有負趨光性之 sensory rhodopsin II (SRII)。
根據 2004 年完成的 Haloarcula marismortui 的基因體計畫，在其基因註解(annotation) 中，具有六個 photoreceptors，是目前所知在單一嗜鹽古細菌中最多的。本論文主要針對兩大問題：A) 訂出這六個預測的 photoreceptors 特徵吸收峰與其功能，以及 B) 這六個 photoreceptors 的生理角色。 本論文成功將六個 photoreceptors 選殖入 E. coli 表現系統且能順利表現，並得知：i) 這六個 photoreceptors 確實的特徵吸收峰；且 ii) 其特徵吸收峰在可見光譜上之分佈十分獨特；此外，iii) 鑑定出一個全新類型的 photoreceptor；並發現 iv) 這是第一個單一微生物體中，擁有兩個光驅動氫離子幫浦的光感受體，且其兩者之特徵吸收峰十分接近；最後，v) 照光影響基因 xop2 的蛋白質 (HmSRM) 表現量，初步證實 H. marismortui 可能具有光適應性生長的現象 (chromatic adaptation)。同時，本論文也在成功地確定這些 photoreceptors 的確具有標準 photoreceptors 特質後，有系統的為它們命名。 | zh_TW |
| dc.description.abstract | Microbial photoreceptors play important roles when responding to environmental stimuli in microbes and a total of more than one hundred microbial photoreceptors has been identified. So far, four distinct functions have been identified, including bacteriorhodopsin (BR) that functions as outward proton pump, and halorhodopsin (HR) can serve as an inward chloride pump; sensory rhodopsin I (SRI) was shown to mediate both attract and repellent signaling, and sensory rhodopsin II (SRII) that triggers repellent signaling against near-UV light.
According to the genome project of the Haloarcula marismortui, there are six predicted photoreceptors, the most numbered photoreceptors in a single archaeon. This goals of this study will be: A) Determination of the maximum absorbance and functions of those six predicted photoreceptors and, B) the potential physiological roles of them. We successful cloned and over-expressed all six photoreceptors using E. coli system, and concluded that: First, those six photoreceptors have their characteristic maximum absorbance, and second, a unique distribution pattern according to the maximum absorbance of those six photoreceptors. Third, a new type of sensory rhodopsin is identified; and fourth, two photoreceptors function as proton pumps and share the same maximum absorbance; the first time a single microbe found to have two light-driven proton pumps. Finally, chromatic adaptation is concluded in H. marismortui since different expression levels of HmSRM was observed when cells were grown under different wavelength of lights. Also, all of the photoreceptors were officially nominated after the confirmation of their characteristics as standard photoreceptors. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-14T17:14:27Z (GMT). No. of bitstreams: 1 ntu-97-R95b47206-1.pdf: 12130302 bytes, checksum: 5ff2ff61ee93ad03e36bcf7bf34de9c2 (MD5) Previous issue date: 2008 | en |
| dc.description.tableofcontents | 目錄
目錄 i 圖目錄 iii 表目錄 iv 附錄 v 摘要 vi Abstract vii 第一章 緒論 1 1.1 微生物視紫紅質 (Microbial rhodopsin) 1 1.2 Haloarcula marismortui 3 1.3國內外研究情形 4 1.3.1 基因選殖與表現 4 1.3.2 光週期 (Photocycle) 5 1.3.3 功能性分析 5 1.3.4 立體結構解析 6 1.3.5 蛋白質工程 7 1.4 實驗動機 9 1.4.1 膜蛋白質大量表現、分離及純化 9 1.4.2 特徵作用光譜分析 (Action spectrum) 10 1.4.3 光週期 (Photocycle) 10 1.4.4 光驅動離子幫浦能力測試 11 1.4.5 嗜鹽古細菌中新型光源調控反應系統 11 1.4.6 色調適應現象 (Chromatic adaptation) 12 1.5 實驗目的 13 1.5.1 基因選殖及膜蛋白質表現與純化 13 1.5.2 功能性鑑定 13 第二章 材料與方法 14 第一節 實驗材料與藥品 14 1.1 菌種 14 1.2 質體 15 1.3 藥品 15 第二節 實驗儀器與設備 16 2.1 核酸電泳設備 16 2.2 蛋白質電泳與轉印設備 16 2.3 離心機 16 2.4 其他 17 第三節 實驗方法 18 3.1 於 Haloarcula marismortui 中選殖光感蛋白質與其相關蛋白質 18 3.2 於大腸桿菌 E. coli 中表現 H. marismortui 中的光感蛋白質與其相關蛋白質 18 3.3 重組光感蛋白質與其相關蛋白質純化 19 3.4 蛋白質分析 20 第三章 結果與討論 24 第一節 生物資訊軟體分析 24 1.1 六種 photoreceptors 演化樹之建立 24 1.2 六種 photoreceptors DNA 序列分析 26 1.3 六種 photoreceptors 胺基酸序列分析 27 第二節 基因選殖、重組及表現蛋白質 30 2.1 基因選殖及重組 30 2.2 以E. coli 系統表現蛋白質 31 2.3 Photoreceptor 全光譜特徵鑑定 33 第三節 功能性鑑定：離子幫浦能力 (Ion pump activity) 35 3.1 光驅動氫離子幫浦活性測試 35 3.2 雷射光誘發光感受體的光週期 36 第四節 色調適應現象 (Chromatic adaptation) 39 4.1 生長趨勢及培養基中 pH 值改變 39 4.2 H. marismortui 全膜蛋白質可見光譜 41 第四章 總結 45 第五章 未來展望 47 參考文獻 48 圖目錄 圖 一：Haloarchaea 中四類 photoreceptors 及其功能.. 2 圖 二：F. diplosiphon 的互補色調適應現象的顏色......... 12 圖 三：H. marismortui 中六種光感受器之親緣關係......... 25 圖 四：藉由 SOSUI 預測 H. marismortui 中六種光感受器之二級結構................................................... 28 圖 五：藉由 SWISS-MODEL 預測 H. marismortui 中六種光感受器三級結構............................................ 29 圖 六：六種表現 photoreceptors 後的 E. coli 菌體顏色.....32 圖 七：六種表現的 photoreceptors 的顏色............ 33 圖 八：六種 photoreceptors 的全光譜...................34 圖 九：H. marismorui 中六個 photoreceptors 的初步 photocycle 結果................................. 37 圖 十：不同照光條件下 H. marismortui 生長趨勢 (OD600 變化) ........................................... 40 圖 十一：不同照光條件下 H. marismortui 培養液 pH 變化... 41 圖 十二：H. marismortui 全膜蛋白質在不同光照條件下可見光光譜................................................... 42 圖 十三：過去研究中已鑑定出最高特徵吸收峰的各種 photoreceptors 的可見光分 布及所屬物種........................................ 44 表目錄 表 一：六種 H. marismortui 中光感受器中之功能預測....24 表 二：六種 H. marismortui 中光感受器 DNA 序列 GC 含量..26 表 三：六種 H. marismortui 中光感受器之等電點...........27 表 四：六種 photoreceptors 之引子序列................. 31 | |
| dc.language.iso | zh-TW | |
| dc.subject | Haloarcula marismortui | zh_TW |
| dc.subject | 光感受體 | zh_TW |
| dc.subject | 基因體計畫 | zh_TW |
| dc.subject | 嗜鹽古細菌 | zh_TW |
| dc.subject | photoreceptor | en |
| dc.subject | Haloarchaea | en |
| dc.subject | Haloarcula marismortui | en |
| dc.subject | genome project | en |
| dc.title | 表現 Haloarcula marismortui 之六個光感受體揭露其獨特的感光特性 | zh_TW |
| dc.title | Expression of Six Photoreceptors in Haloarcula marismortui Unveiled Unique Photosensing Features | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 96-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 王惠鈞,吳韋訥,李平篤,許瑞祥 | |
| dc.subject.keyword | 嗜鹽古細菌,Haloarcula marismortui,基因體計畫,光感受體, | zh_TW |
| dc.subject.keyword | Haloarchaea,Haloarcula marismortui,genome project,photoreceptor, | en |
| dc.relation.page | 50 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2008-07-28 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 微生物與生化學研究所 | zh_TW |
| 顯示於系所單位： | 微生物學科所 | |
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