Live-Cell Labeling of Specific Protein Glycoforms by Proximity-Enhanced Bioorthogonal Ligation.
Reagents for detecting post-translational modifications in the context of their protein scaffold are powerful tools, but are challenging to develop for glycosylated epitopes. We describe a strategy for detecting protein-specific glycosylation through the use of cyclooctyne-aptamer conjugates. These molecules selectively ligate to azidosugar-labeled glycans exclusively on a target protein on live cells. We characterized aptamer conjugates against two different cell surface glycoproteins and show that these reagents are amenable to detecting protein sialoforms by mass spectrometry, Western blotting, and flow cytometry. Given the abundance of aptamers that bind cell surface targets, we expect this technology will be a useful platform for investigating the roles of protein-specific glycosylation in various cellular contexts.
Departments of Chemistry and Molecular and Cell Biology and the Howard Hughes Medical Institute, University of California , Berkeley, California 94720, United States.
Peter V
PV
Robinson
Departments of Chemistry and Molecular and Cell Biology and the Howard Hughes Medical Institute, University of California , Berkeley, California 94720, United States.
Gabriela
G
de Almeida-Escobedo
Departments of Chemistry and Molecular and Cell Biology and the Howard Hughes Medical Institute, University of California , Berkeley, California 94720, United States.
Amber E
AE
de Groot
Departments of Chemistry and Molecular and Cell Biology and the Howard Hughes Medical Institute, University of California , Berkeley, California 94720, United States.
Julia L
JL
McKechnie
Departments of Chemistry and Molecular and Cell Biology and the Howard Hughes Medical Institute, University of California , Berkeley, California 94720, United States.
Carolyn R
CR
Bertozzi
D052157:Aptamers, Nucleotide / Q000378:metabolism; D002470:Cell Survival; D006023:Glycoproteins / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D013194:Staining and Labeling / Q000379:methods*
D052157:Aptamers, Nucleotide; D006023:Glycoproteins