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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 孔繁璐(Fan-Lu Kung) | |
| dc.contributor.author | Ke-Ming Huang | en |
| dc.contributor.author | 黃可名 | zh_TW |
| dc.date.accessioned | 2021-06-15T02:57:08Z | - |
| dc.date.available | 2014-10-07 | |
| dc.date.copyright | 2011-10-07 | |
| dc.date.issued | 2011 | |
| dc.date.submitted | 2011-08-16 | |
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(2006) Solution conformation and heparin-induced dimerization of the full-length extracellular domain of the human amyloid precursor protein, J Mol Biol 357, 493-508. 21. Chattopadhyay, A., and London, E. (1988) Spectroscopic and ionization properties of N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-labeled lipids in model membranes, Biochim Biophys Acta 938, 24-34. 22. Lin, S., and Struve, W. S. (1991) Time-resolved fluorescence of nitrobenzoxadiazole-aminohexanoic acid: effect of intermolecular hydrogen-bonding on non-radiative decay, Photochem Photobiol 54, 361-365. 23. Lee, M. S., Kao, S. C., Lemere, C. A., Xia, W., Tseng, H. C., Zhou, Y., Neve, R., Ahlijanian, M. K., and Tsai, L. H. (2003) APP processing is regulated by cytoplasmic phosphorylation, J Cell Biol 163, 83-95. 24. Koshino, I., and Takakuwa, Y. (2009) Disruption of lipid rafts by lidocaine inhibits erythrocyte invasion by Plasmodium falciparum, Exp Parasitol 123, 381-383. 25. Kamiya, Y., Ohta, K., and Kaneko, Y. (2005) Lidocaine-induced apoptosis and necrosis in U937 cells depending on its dosage, Biomed Res 26, 231-239. 26. Ehehalt, R., Keller, P., Haass, C., Thiele, C., and Simons, K. (2003) Amyloidogenic processing of the Alzheimer beta-amyloid precursor protein depends on lipid rafts, J Cell Biol 160, 113-123. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/44430 | - |
| dc.description.abstract | 在阿茲海默氏症(Alzheimer’s disease, AD)的研究領域中，膽固醇與b-amyloid precursor protein (APP)蛋白之間的關係一直是個懸而未決之謎。膽固醇可能借由與APP蛋白有直接的交互作用，而促使APP蛋白走向產生導致阿茲海默氏症的Ab的水解途徑。為了要進一步探討膽固醇與APP蛋白之間的交互作用如何影響APP的水解，我們利用點突變的方式將APP蛋白上與膽固醇做辨認的cholesterol recognition amino acid consensus (CRAC)序列做突變，試著藉由突變產生無法與膽固醇有良好交互作用的APP突變型蛋白。實驗結果發現，APP突變型與lipid raft的標誌蛋白flotillin-1 共分層的程度比野生型來得少；APPY422L與APPS622L在經常用來評估引發致病水解途徑的C99/C83比值上，也較野生型來得低；更進一步的研究發現在去除了膜上的膽固醇後，和僅僅破壞膜上的raft構造，但膜上的膽固醇含量卻仍維持不變時相比，APP蛋白的分層情況並不相同。這些結果指出膽固醇與APP蛋白之間的交互作用力可能造成APP蛋白更容易散布進入lipid raft中，使得APP蛋白更有機會被BACE1水解酶水解，產生Ab片段。 | zh_TW |
| dc.description.abstract | The relationship between cholesterol and b-amyloid precursor protein (APP) processing remains an unsolved mystery in the Alzheimer’s disease (AD) research field. It has been hypothesized that there is a direct interaction between cholesterol and APP, which might facilitate APP being processed into Ab in its pathogenic pathway. To assess the importance of the APP-cholesterol in the regulating of regulating APP processing, we generated several cholesterol-binding-deficient APP mutant proteins via introducing mutations in the domains with the putative cholesterol recognition amino acid consensus (CRAC) sequence of APP. Our results revealed that a smaller portion of APP mutants was co-fractionated with the lipid raft marker flotillin-1 compared to wild-type APP. The C99/C83 ratio, which is usually used in evaluating pathogenesis processing level, is also reduced by Y422L and S622L mutations. Further study shows APP pattern in cholesterol-depleted cells was different from raft-disrupted cells in which the membrane cholesterol content remained unchanged. These data suggested that the interaction between cholesterol and APP could be a driving force for distributing APP into lipid raft, which makes APP more accessible to the pathogenically critical protein BACE1. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-15T02:57:08Z (GMT). No. of bitstreams: 1 ntu-100-R97423028-1.pdf: 2684258 bytes, checksum: 0c13505be8a98ecde8fb9c511add2e12 (MD5) Previous issue date: 2011 | en |
| dc.description.tableofcontents | 口試委員會審定書…………………………………………………… I
誌謝…………………………………………………………………… II 中文摘要……………………………………………………………… III 英文摘要……………………………………………………………… IV Abbreviations ……………………………………………………… V Contents ……………………………………………………………… VII Introduction …………………………………………………………… 1 Material and Methods ……………………………………………… 4 Site-directed mutagenesis ………………………………………… 4 GST-APP fusion protein purification ………………………… 5 NBD-cholesterol binding assay ………………………………… 6 HEK293T cell transfection ……………………………………… 7 Protein lysate extraction ……………………………………… 8 Western blot ………………………………………………………… 8 Sucrose gradient fractionation ………………………………… 9 Silver stain ………………………………………………………… 9 Cholesterol assay …………………………………………………10 MTT assay ……………………………………………………………… 10 Lipid extraction …………………………………………………… 11 Results ………………………………………………………………… 12 APP mutants …………………………………………………………… 12 NBD-cholesterol binding assay ……………………………………13 Sucrose gradient fractionation ………………………………… 14 C99/C83 ratio …………………………………………………………16 MβCD and lidocaine treatment ……………………………………17 Discussion …………………………………………………………… 19 Cholesterol recognition amino acid consensus (CRAC) and APP mutants …19 NBD-cholesterol binding assay ……………………………………… 21 Sucrose gradient experiment and C99/C83 ratio assay ……… 23 MβCD and lidocaine treatment …………………………………… 24 Figures ……………………………………………………………… 26 Tables ……………………………………………………………… 44 References …………………………………………………………… 45 | |
| dc.language.iso | en | |
| dc.title | 膽固醇在APP蛋白質水解反應中所扮演的角色 | zh_TW |
| dc.title | The Role of Cholesterol in Aβ Precursor Protein Processing | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 99-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 李財坤(Tsai-Kun Li),兵岳忻(Yueh-Hsin Ping),顧記華(Jih-Hwa Guh) | |
| dc.subject.keyword | 膽固醇,APP蛋白,CRAC序列,lipid raft,C99/C83比值, | zh_TW |
| dc.subject.keyword | cholesterol,APP protein,CRAC sequence,lipid raft,C99/C83 ratio, | en |
| dc.relation.page | 47 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2011-08-16 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 藥學研究所 | zh_TW |
| 顯示於系所單位： | 藥學系 | |
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