Suzuki Masafumi, Furukawa Shoichiro, Kuramori Chikanori, Sawa Chika, Kabe Yasuaki, Nakamura Mitsuhiro, Sawada Jun-Ichi, Yamaguchi Yuki, Sakamoto Satoshi, Inouye Satoshi, Handa Hiroshi
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama 226-8501, Japan.
Biochem Biophys Res Commun. 2008 Apr 11;368(3):600-5. doi: 10.1016/j.bbrc.2008.01.107. Epub 2008 Feb 1.
We developed a unique screening system that consists of combination of high photo-sensitivity of photoprotein aequorin (AQ) and our developed high-performance affinity purification system. In the present study, we demonstrated to detect the specific interaction between methotrexate (MTX) and its target dihydrofolate reductase (DHFR) fused with AQ. We succeeded to prepare highly purified AQ-fused DHFR, which showed high sensitive light emission. To test the screening system, we prepared the complex of MTX-immobilized magnetic nanobeads and AQ-fused DHFR. Bound AQ-fused DHFR with the beads was specifically released by addition of MTX. Thus, this methodology enables us to search a novel chemical that binds to target proteins without complicated processes. Furthermore, thank to the highly sensitive luminescence intensity of AQ, this methodology would be performed in very small scale with high responsibility, leading to development of high throughput screening systems.
