小分子物質與離子對蛋白質行使功能扮演重要的角色,包括對於細胞訊息傳導相當重要的Ca2+或是與神經衝動傳遞有關的Na2+與K+。為了可以更容易偵測小分子物質與離子的移動,本研究使用一簡易電化裝置來測量蛋白質行使功能時所造成的帶電小分子或離子的移動。電化裝置中由6至8 kDa的半透膜隔成sample cell與reference cell,並利用可以透過光線與導電的氧化銦錫(indium tin oxide, ITO)玻璃作為電極,在不對系統施加偏壓的狀況下測量訊號。在本研究中利用此ITO電化裝置成功偵測光驅動離子幫浦(Light-driven ion pump) Bacteriorhodopsin(BR)與Halorhodopsin(HR)受光刺激後運送離子所造成的電流訊號,也成功偵測到單純加入帶電小分子物質後引起的電流訊號,更進一步用來偵測點突變蛋白質,用以鑑定特定胺基酸在蛋白質行使功能時所扮演的角色。綜合以上結果,此一ITO電化裝置可以用於研究小分子物質或離子的移動,與蛋白質行使功能之間的關係。
Small charged molecules and ions play important roles in functioning proteins, including the cell signal transduction messenger Ca2+ and Na2+ and K+ for nerve impulse. In order to detect the movement of small charged molecules and ions in functioning proteins, an ITO (Indium tin oxide)-coated electrochemical device was adopted as one of the components when we designed a system to serve this need. In this bias-voltage free ITO-coated electrochemical system, we used a 6 to 8 kDa semi permeable membrane to separate the chamber intosample cell and reference cell, while the ITO-coated slides served as electrodes.The results showed we successfully employed ITO-coated electrochemical device to detect the light-driven proton and chloride signals of two light-driven ion pumps, bacteriorhodopsin and halorhodopsin, respectively. We also successfully detected the current signals caused by small charged molecules. We further used this device to investigate the relationship between mutated proteins and their functions. According to above results, this ITO-coated electrochemical device can be used to investigate the movement of small charged molecules and ions in functioning proteins in a quantive way.
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