title: Semisynthetic lectin-4-dimethylaminopyridine conjugates for labeling and profiling glycoproteins on live cell surfaces.
abstract: Glycoproteins on cell surfaces play important roles in biological processes, including cell-cell interaction/signaling, immune response, and cell differentiation. Given the diversity of the structure of glycans, labeling and imaging of selected glycoproteins are challenging, although several promising strategies have been developed recently. Here, we design and construct semisynthetic reactive lectins (sugar-binding proteins) that are able to selectively label glycoproteins. Congerin II, an animal galectin, and wheat germ agglutinin are conjugated with 4-dimethylaminopyridine (DMAP), a well-known acyl transfer catalyst by our affinity-guided DMAP method and Cu(I)-assisted click chemistry. Selective labeling of glycoproteins is facilitated by the DMAP-tethered lectin catalysts both in vitro and on living cells. Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) analysis enabled us to isolate labeled glycoproteins that are uniquely exposed on distinct cell lines. Furthermore, the combination of immunoprecipitation with mass spectrometry (MS)-fingerprinting techniques allowed us to characterize 48 glycoproteins endogenously expressed on HeLa cells, and some low-abundant glycoproteins, such as epidermal growth factor receptor (EGFR) and neuropilin-1, were successfully identified. Our results demonstrate that semisynthetic DMAP-tethered lectins provide a new tool for labeling and profiling glycoproteins on living cells.
authors.affiliation: Department of Synthetic Chemistry and Biological Chemistry, Graduate School of Engineering, Kyoto University, Katsura, Kyoto 615-8510, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Hayashi
authors.forename: Yedi
authors.initials: Y
authors.lastname: Sun
authors.forename: Tomonori
authors.initials: T
authors.lastname: Tamura
authors.forename: Keiko
authors.initials: K
authors.lastname: Kuwata
authors.forename: Zhining
authors.initials: Z
authors.lastname: Song
authors.forename: Yousuke
authors.initials: Y
authors.lastname: Takaoka
authors.forename: Itaru
authors.initials: I
authors.lastname: Hamachi
mesh_terms: D015761:4-Aminopyridine / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000737:chemistry; D002470:Cell Survival; D060326:Chemistry Techniques, Synthetic; D006023:Glycoproteins / Q000737:chemistry*; D006367:HeLa Cells; D006801:Humans; D037102:Lectins / Q000737:chemistry*; D008958:Models, Molecular; D011487:Protein Conformation; D013194:Staining and Labeling
chemical_list: D006023:Glycoproteins; D037102:Lectins; D015761:4-Aminopyridine; C003885:4-dimethylaminopyridine