Ultrafast and Chemoselective Biotinylation of Living Cell Surfaces for Time-Resolved Surfaceome Analysis

Cell surface proteins participate in many important biological
processes, such as cell-to-cell interaction, signal transduction,
cell adhesion, and protein transportation. In-depth study of the cell
surface protein group is of great significance. Nevertheless, detection
and analysis of the surfaceome remain a significant challenge due
to their low abundance and hydrophobicity. Herein, we reported an
ultrafast and chemoselective labeling method using our newly developed
trifunctional probe, the OPA-S-S-alkyne, which labeled cell surface
lysine residues, and then established a novel cell surfaceome profiling
approach. According to our experimental results, the OPA-S-S-alkyne
probe can react extremely fast with living cells, labeling cells in
only 1 min, while traditional NHS (labeling cell surface lysine with N-hydroxysuccinimide ester probe) and CSC (labeling cell
surface glycan with hydrazide biotin probe) methods normally take
longer time of more than 30 min and 1 h, respectively. Taking advantage
of this ultrafast property of the method, we highlight the utility
of this method by exploring the temporal dynamic changes of surfaceome
upon EGF stimulation in living Hela cells and reported “early”
and “late” EGF-regulated cell surface proteins, which
are difficult to be distinguished by the current cell surface profiling
approaches.