Search for in vivo substrates and inhibitors of calpain related to Alzheimer's disease

• Project/Area Number: 25350917
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Multi-year Fund
• Section: 一般
• Research Field: Applied health science
• Research Institution: Institute of Physical and Chemical Research
• Principal Investigator: TSUBUKI SATOSHI 国立研究開発法人理化学研究所, 脳科学総合研究センター, 専門職研究員 (20217368)
• Project Period (FY): 2013-04-01 – 2016-03-31
• Project Status: Completed (Fiscal Year 2015)
• Budget Amount: ¥4,420,000 (Direct Cost: ¥3,400,000、Indirect Cost: ¥1,020,000); Fiscal Year 2015: ¥1,040,000 (Direct Cost: ¥800,000、Indirect Cost: ¥240,000); Fiscal Year 2014: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000); Fiscal Year 2013: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
• Keywords: アルツハイマー病 / カルパイン / カルパスタチン / 二次元電気泳動 / タンパク質同定 / 質量分析 / 遺伝子改変マウス / 初代培養細胞 / 等電点電気泳動

Outline of Final Research Achievements

I tried to identify in vivo substrates of calpain related to Alzheimer’s disease using hippocampal primary neurons of calpastatin-ko and wild-type mice. I analysed four fractions of homogenates of primary neurons by two dimensional fluorescence difference gel electrophoresis. Also I applied them to the calpain-binding column and identified the adsorbed protein as a in vivo substrate by liquid chromatography-mass spectrometry. And I tried kinetic analysis using the specific antibodies.