Functional Expression of Prestin, the Outer Hair Cell Motor Protein, Using the Baculovirus/Insect Cell System
スポンサーリンク
概要
- 論文の詳細を見る
A source of the electromotility of the outer hair cells (OHCs), which enables the high sensitivity of mammalian hearing, is believed to be voltage-dependent conformational change of the motor protein prestin embedded in the lateral wall of the OHCs. As a result of extensive studies on this unique motor protein using prestin-expressing cells, knowledge about its structure and function has been gradually accumulated. To obtain further knowledge about prestin, research using purified prestin molecules is necessary, and thus a method for efficiently obtaining prestin molecules is required. In the present study, construction of an expression system for prestin using the baculovirus/Sf9 insect cell system was attempted. The expression and localization in the plasma membrane of prestin in infected Sf9 cells were confirmed by Western blotting and immunofluorescence analysis, and prestin's functional activity was determined by patch-clamp measurements. Furthermore, to obtain prestin from the cells efficiently, culture conditions of the cells were examined, and it was clarified that cells should be harvested around 72 hours after infection. Quantitative Western blotting revealed that the maximal amount of prestin per liter of culture medium was around 800 μg. The results of this study indicate that the expression system which was constructed is potentially useful for purification of prestin due to the rapidity of its construction and its high yield.
- 一般社団法人 日本機械学会の論文
著者
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Kobayashi Toshimitsu
Department Of Cardiovascular Surgery Tohoku University Graduate School Of Medicine
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Tsumoto Kouhei
Department Of Biomolecular Engineering Graduate School Of Engineering Tohoku University
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Kumagai Izumi
Department of Biomolecular Engineering, Tohoku University
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Kumagai Izumi
Department Of Biochemistry And Engineering Graduate School Of Engineering Tohoku University
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Iida Koji
Department Of Applied Physics National Defence Academy
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Kumano Shun
Department Of Bioengineering And Robotics Tohoku University
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Ikeda Katsuhisa
Department Of Otolaryngology
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Murakoshi Michio
Department Of Bioengineering And Robotics Tohoku University
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Wada Hiroshi
Department Of Bioengineering And Robotics Tohoku University
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Wada Hiroshi
Department of Bioengineering and Robotics, Tohoku University
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TADENUMA Takashi
Department of Bioengineering and Robotics, Tohoku University
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