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Title: 多麩醯胺擴增TBP 與HMGB1 的交互作用研究
Other Titles: Interaction Analysis Between Polyglutamine-expanded TATA Box Binding Protein (TBP) and High Mobility Group Box 1 (HMGB1)
Authors: 楊淑婷
Shu-Ting Yang, Hsiang-Ming Chen, Li-Ching Lee, Shih-Han Huang, Chih-Hsin Lin, Guan-Chiun Lee, Cheng-Kang Lee, Guey-Jen Lee-Chen
Issue Date: 2013
Publisher: 國立臺灣師範大學生命科學學系
Department of Life Science, NTNU
Abstract: 第十七型脊髓小腦共濟失調症(SCA17)與轉錄起始因子TATA binding protein (TBP)基因上的多麩醯胺擴增相關。TBP 與包括high mobility group box 1 (HMGB1)在內的其他蛋白因子交互作用,來調節基因的表現。本研究藉重組的HMGB1 及TBP/Q20~61 N 端蛋白,探討TBP Q 長度是否影響其與HMGB1 的結合。首先共表現HMGB1 及TBP 於 HEK-293 細胞後,利用免疫共沉澱及GSTpull-down 試驗,確認HMGB1 與 TBP 在細胞內的結合。其次原核E. coli 表現的重組HMGB1 及TBP/Q20~61 蛋白的GST pull-down 試驗,亦顯示HMGB1 與TBP 的結合。最後即時石英晶體微量天平(QCM)試驗顯示,HMGB1-TBP 交互作用隨TBP 蛋白上polyQ 長度的增加而減弱。綜合先前報導的HMGB1-TBP 交互作用可增強TBP 結合TATA 速率及穩定性,我們的研究顯示多麩醯胺擴增TBP 與HMGB1 結合的減弱,可能與SCA17 致病機轉相關。
Spinocerebellar ataxia type 17 (SCA17) involves the expression of a polyglutamine-expanded TATA box binding protein (TBP), a general transcription initiation factor. TBP interacts with other protein factors, including high mobility group box 1 (HMGB1), to regulate gene expression. In this study, we examined the effect of TBP Q-tract length on HMGB1 binding using recombinant HMGB1 and N-terminal TBP/Q20~61 proteins. The intracellular association of HMGB1 with TBP was first confirmed by using a half-in vivo co-immunoprecipitation (co-IP) and glutathione S-transferase (GST) pull-down assays in HMGB1 and TBP co-expressed HEK-293 cells. GST pull-down assay using soluble recombinant HMGB1 and TBP/Q20~61 proteins expressed in E. coli also revealed the binding activity between recombinant HMGB1 and TBP. When the binding strength was explored by using a real time quartz crystal microbalance (QCM) assay, a Q-tract length-dependent decrease of HMGB1-TBP interaction was demonstrated. Together with the reported HMGB1-TBP interaction to increase the rate of TBP binding and the stability of the TBP/TATA interaction, our study results suggest that decrease HMGB1 binding to polyQ-expanded TBP may contribute to the pathogenesis of SCA17.
Other Identifiers: 77F82F03-4467-0C50-DBC9-176B2D7D700C
Appears in Collections:生物學報

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