網羅的グライコプロテオミクスのためのシステム開発
詳細
In order to elucidate precise functions of glycans attached to extensive proteins, it is necessary to develop a fundamental database of glycoproteins, which is annotated as regards not only genes that encode core proteins and glycosylation sites, but also glycan structures. Various techniques have now been attempted to achieve this with the aid of advanced procedures of mass spectrometry. However, it has not yet been achieved in a fully satisfactory manner. In the course of the NEDO project "Structural Glycomics" (2002.3-2005.3), we have started developing a total system for glycoproteomics by collaboration of GL Sciences on the basis of a recently established procedure named "glyco-catch", which enables selective capture of a set of glycopeptides by lectin columns followed by a large scale of identification of glycoprotein genes and Asn-linked glycosylation sites by an improved proteomic MS/MS procedure (1,2). To further profile glycan structures, however, the above strategy must be much improved as regards in particular 1) manipulation of a number of glycopeptides separated by a 2D-LC system, each of which is subjected to peptide-N-glycanse treatment to liberate glycans, and 2) selective recovery of peptides and glycans for further structural analysis by either MS or lectin profiling. In the session, the core strategy is described with a few experiments using a model glycoprotein and actual samples of mouse glycoproteins captured by a ConA-column.
(1) Hirabayashi, J., Kaji, H., Isobe, T., and Kasai K. J. Biochem (Tokyo) 132, 103 - 114, 2002
(2) Kaji, H., Saito, H., Yamauchi, Y., Shinkawa, T., Taoka, M., Hirabayashi, J., Kasai, K., Takahashi, N., Isobe, T. Nat Biotechnol. 21, 667-672, 2003
