Exo-Enzymatic Addition of Diazirine-Modified Sialic Acid to Cell Surfaces Enables Photocrosslinking of Glycoproteins
| Autor: | Nageswari Yarravarapu, Rohit Sai Reddy Konada, Narek Darabedian, Nichole J. Pedowitz, Soumya N. Krishnamurthy, Matthew R. Pratt, Jennifer J. Kohler |
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| Rok vydání: | 2022 |
| Předmět: | |
| Zdroj: | Bioconjug Chem |
| ISSN: | 1520-4812 1043-1802 |
| DOI: | 10.1021/acs.bioconjchem.2c00037 |
| Popis: | Glycan binding often mediates extracellular macromolecular recognition events. Accurate characterization of these binding interactions can be difficult because of dissociation and scrambling that occur during purification and analysis steps. Use of photocrosslinking methods has been pursued to covalently capture glycan-dependent interactions in situ however use of metabolic glycan engineering methods to incorporate photocrosslinking sugar analogs is limited to certain cell types. Here we report an exo-enzymatic labeling method to add a diazirine-modified sialic acid (SiaDAz) to cell surface glycoconjugates. The method involves chemoenzymatic synthesis of diazirine-modified CMP-sialic acid (CMP-SiaDAz), followed by sialyltransferase-catalyzed addition of SiaDAz to desialylated cell surfaces. Cell surface SiaDAz-ylation is compatible with multiple cell types and is facilitated by endogenous extracellular sialyltransferase activity present in Daudi B cells. This method for extracellular addition of α2-6-linked SiaDAz enables UV-induced crosslinking of CD22, demonstrating the utility for covalent capture of glycan-mediated binding interactions. |
| Databáze: | OpenAIRE |
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