Functional Neoglycopeptides: Synthesis and Characterization of a New Class of MUC1 Glycoprotein Models Having Core 2-Based O-Glycan and Complex-Type N-Glycan Chains
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- 作者：Takahiko Matsushita ; Reiko Sadamoto ; Naoki Ohyabu ; Hideki Nakata ; Masataka Fumoto ; Naoki Fujitani ; Yasuhiro Takegawa ; Takeshi Sakamoto ; Masaki Kurogochi ; Hiroshi Hinou ; Hiroki Shimizu ; Takaomi Ito ; Ke
- 刊名：Biochemistry
- 出版年：2009
- 出版时间：November 24, 2009
- 年：2009
- 卷：48
- 期：46
- 页码：11117-11133
- 全文大小：1385K
- 年卷期：v.48,no.46(November 24, 2009)
- ISSN：1520-4995
文摘
An efficient protocol for the construction of MUC1-related glycopeptide analogues having complex O-glycan and N-glycan chains was established by integrating chemical and enzymatic approaches on the functional polymer platforms. We demonstrated the feasibility of sortase A-mediated ligation between two glycopeptide segments by tagging with signal peptides, LPKTGLR and GG, at each C- or N-terminal position. Structural analysis of the macromolecular N,O-glycopeptides was performed by means of ESI-TOFMS (MS/MS) equipped with an electron-captured dissociation device. Immunological assay using MUC1 glycopeptides synthesized in this study revealed that N-glycosylation near the antigenic O-glycosylated PDTR motif did not disturb the interaction between the anti-MUC1 monoclonal antibody and this crucial O-glycopeptide moiety. NMR study indicated that the N-terminal immunodominant region [Ala-Pro-Asp-Thr(O-glycan)-Arg] forms an inverse γ-turn-like structure, while the C-terminal region composed of N-glycopeptide and linker SrtA-peptide was proved to be an independently random structure. These results indicate that the bulky O- and N-glycan chains can function independently as disease-relevant epitopes and ligands for carbohydrate-binding proteins, when both are combined by an artificial intervening peptide having a possible effect of separating N- and C-terminal regions. The present strategy will greatly facilitate rapid synthesis of multiply functionalized complex neoglycopeptides as new types of convenient tools or models for the investigation of thhe structure−function relationship of various glycoproteins and development of novel class glycopeptide-based biopharmaceuticals, drug delivery systems, and biomedical materials.