Exo-Enzymatic Addition of Diazirine-Modified Sialic Acid to Cell Surfaces Enables Photocrosslinking of Glycoproteins

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 coval...

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Bibliographic Details
Published in:Bioconjugate chemistry 2022-05, Vol.33 (5), p.781-787
Main Authors: Yarravarapu, Nageswari, Konada, Rohit Sai Reddy, Darabedian, Narek, Pedowitz, Nichole J., Krishnamurthy, Soumya N., Pratt, Matthew R., Kohler, Jennifer J.
Format: Article
Language:English
Subjects:
Binding
CD22 antigen
Cell surface
Crosslinking
Diazomethane - chemistry
Glycan
Glycoconjugates
Glycoproteins
Glycoproteins - chemistry
Lymphocytes B
Macromolecules
Metabolic engineering
N-Acetylneuraminic Acid - chemistry
Polysaccharides - chemistry
Sialic Acids - chemistry
Sialyltransferases - chemistry
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Summary: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 the chemoenzymatic synthesis of diazirine-modified CMP-sialic acid (CMP-SiaDAz), followed by sialyltransferase-catalyzed addition of SiaDAz to desialylated cell surfaces. Cell surface SiaDAzylation 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.
ISSN:1043-1802
1520-4812
DOI:10.1021/acs.bioconjchem.2c00037