Inhibition assay system for on-chip phosphorylation of peptide by surface plasmon resonance imaging technique

ANYL 166

Kazuki Inamori, kazuki_inamori@bio.toyobo.co.jp1, Motoki Kyo, motoki_kyo@bio.toyobo.co.jp2, Kazuki Matsukawa, kazuki_matsukawa@bio.toyobo.co.jp1, Yusuke Inoue, yusuke-inouetcm@mbox.nc.kyushu-u.ac.jp3, Tatsuhiko Sonoda, sonotcm@mbox.nc.kyushu-u.ac.jp3, Eiji Kinoshita, kinoeiji@hiroshima-u.ac.jp4, Tohru Koike, tkoike@hiroshima-u.ac.jp4, and Yoshiki Katayama, ykatatcm@mbox.nc.kyushu-u.ac.jp3. (1) Biotechnology Frontier Project, Toyobo Co., Ltd, 10-24, Toyo-cho, Tsuruga-shi, Fukui-ken, 914-0047, Japan, (2) Biotechnology Development Department, Toyobo Co., Ltd, 2-2-8,Dojima-hama, Kita-ku, Osaka-shi, 530-8230, Japan, (3) Department of Applied Chemistry Faculty of Engineering, Kyushu University, 744, Moto-oka, Nishi-ku, Fukuoka-shi, 819-0395, Japan, (4) Department of Functional Molecular Science, Division of Medicinal Chemistry, Graduate School of Biomedical Sciences, Hiroshima University, 1-2-3 Kasumi Minami-ku, Hiroshima-shi, 734-8551
We had previously reported a detection and quantification system for on-chip phosphorylation of immobilized peptides by surface plasmon resonance (SPR) imaging using biotinylated zinc(II) complex. It is the comprehensive analysis technology of the protein kinase activity. In SPR analysis, the array which was incubated by biotynylated zinc(II) complex solution in advance was exposed with streptavidin (SA) and anti-SA antibody solution. In this study, we examined the inhibition assay using this system. The covalently immobilized peptide arrays were reacted with PKA solution containing ATP. Then, PKA inhbitor was coexisted at various concentrations. By both SPR imaging and autoradiography analysis, we found the inhibition effect by the coexistence of an inhibitor. The inhibition effect was enhanced with the concentration of inhibitor. Using other kinds of protein kinases, on-chip inhibition assay was also achieved. We propose that our system for the on-chip phosphorylation assay is very valuable for the screening of drug.
 

General Papers
7:00 PM-9:00 PM, Sunday, 10 September 2006 Moscone Center -- Hall D, Poster

Division of Analytical Chemistry

The 232nd ACS National Meeting, San Francisco, CA, September 10-14, 2006