1. Efficient Preparation and Bioactivity Evaluation of Glycan-Defined Glycoproteins
- Author
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Sanggil Kim, Chang Hee Lee, Ji Young Hyun, Injae Shin, and Hyun Soo Lee
- Subjects
Azides ,Glycan ,Glycosylation ,THP-1 Cells ,media_common.quotation_subject ,G(M2) Ganglioside ,Biochemistry ,Polysaccharides ,Lectins ,Humans ,Receptor ,Internalization ,Glycoproteins ,media_common ,chemistry.chemical_classification ,biology ,General Medicine ,Fibroblasts ,Genetic code ,beta-N-Acetylhexosaminidases ,Amino acid ,chemistry ,Homogeneous ,Alkynes ,Mutation ,Biocatalysis ,Click chemistry ,biology.protein ,Molecular Medicine ,Click Chemistry ,Lysosomes ,Glycoprotein ,Protein Processing, Post-Translational - Abstract
Owing to the generation of heterogeneous glycoproteins in cells, it is highly difficult to study glycoprotein-mediated biological events and to develop biomedical agents. Thus, general and efficient methods to prepare homogeneous glycoproteins are in high demand. Herein, we report a general method for the efficient preparation of homogeneous glycoproteins that utilizes a combination of genetic code expansion and chemoselective ligation techniques. In the protocol to produce glycan-defined glycoproteins, an alkyne tag-containing protein, generated by genetic encoding of an alkynylated unnatural amino acid, was quantitatively coupled via click chemistry to versatile azide-appended glycans. The glycoproteins produced by the present strategy were found to recognize mammalian cell-surface lectins and enter the cells through lectin-mediated internalization. Also, cell studies exhibited that the glycoprotein containing multiple mannose-6-phosphate residues enters diseased cells lacking specific lysosomal glycosidases by binding to the cell-surface M6P receptor, and subsequently migrates to lysosomes for efficient degradation of stored glycosphingolipids.
- Published
- 2020