Profiling the Cellular Surfaceome by Furan-Based Protein Biotinylation

The cellular surfaceome is crucial for cellular homeostasis
and
communication, with surface proteins acting as receptors, transporters,
and enzymes. However, comprehensive analysis of the surfaceome is
challenging due to the hydrophobic nature and low abundance of membrane
proteins, leading to their underrepresentation in mass spectrometry-based
proteomics data. This study exploits a novel method for profiling
the cellular surfaceome using furan chemistry. We synthesized six
furan-biotin compounds (FB1-FB6) and showed that FB1 reacts with lysine
and cysteine. Upon furan activation by oxidation, FB1 exhibited the
highest staining intensity and specificity for cell surface proteins.
Pull-downs of biotinylated proteins further confirmed the efficiency
of FB1 in enriching cell surface proteins, and FB1 was also found
to reduce nonspecific labeling of intracellular proteins compared
to biotinylation methods using N-hydroxysuccinimide
esters. Our method thus provides a robust tool for unbiased, high-specificity
proteomic studies of cell surfaceomes.