A new functional analysis of intracellular ion-transporting proteins

细胞内离子转运蛋白的新功能分析

• 批准号: 24659114
• 负责人: KANEKO Shuji
• 金额: $ 2.5万
• 依托单位: Kyoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Challenging Exploratory Research
• 财政年份: 2012
• 资助国家: 日本
• 起止时间: 2012-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-24659114/
• 关键词: イオン輸送; トランスポータ; 細胞内小器官; シナプス小胞; 自己組織化単分子層; 電気生理学

A novel electrophysiological assay technique based on solid supported membranes (SSM) has been shown to be useful for analysis of channels and transporters. We tried to apply this technique to synaptic vesicles (SVs) and lysosomal membranes (LMs), purified from rat brains and mouse livers, respectively. Application of ATP to the sensors with SVs triggered current responses dependent on Mg2+ and sensitive to bafilomycin A1 (Baf), which are thought to be V-ATPase specific currents. On the other hand, though LMs are also embedded with V-ATPases, the sensors evoked current responses with different characteristics. They were independent of Mg2+, only partially sensitive to Baf and even triggered by application of ADP. These features suggest that the currents are not attributed to V-ATPases but any other lysosomal protein. The present study shows the utility and potential of SSM-based electrophysiology especially for analyzing native organellar membrane proteins.

已证明一种基于固体支持的膜（SSM）的新型电生理测定技术可用于分析通道和转运蛋白。我们试图分别从大鼠大脑和小鼠肝脏纯化的突触囊泡（SVS）和溶酶体膜（LMS）应用于突触囊泡（SVS）和溶酶体膜（LMS）。将ATP应用于具有SVS的传感器，触发了电流响应，取决于MG2+和对Bafilycomin A1（BAF）敏感，这被认为是V-ATPase特异性电流。另一方面，尽管LMS还嵌入了V-ATPases，但传感器诱发了具有不同特征的电流响应。它们独立于MG2+，仅对BAF部分敏感，甚至通过ADP的应用触发。这些特征表明，这些电流不是归因于V-ATPases，而归因于任何其他溶酶体蛋白。本研究表明了基于SSM的电生理学的实用性和潜力，尤其是用于分析天然细胞器膜蛋白。

项目成果

トランスポータ活性を電気的に測定する自動化システム

用于电子测量转运体活动的自动化系统

• 发表时间: 2012
• 作者: Ishiwata R;Yokoyama U;Arakawa N;Nomura A;Oshima T;Minamisawa S;Ishikawa Y;金子周司
• 通讯作者: 金子周司

Application of a novel electrophysiological assay technique to native lysosomal membranes

新型电生理测定技术在天然溶酶体膜中的应用

• 发表时间: 2013
• 期刊: 京都大学薬学部卒業研究論文集
• 作者: 横山詩子;石渡遼;大島登志男;南沢享;石川義弘;Bai Y;Ichikawa Y;Yokota T;横山 詩子;横山詩子;横山詩子;Yokoyama U;Yokoyama U;Yu Sakamoto
• 通讯作者: Yu Sakamoto