Protease-protection strategy combined with the SERS tags for detection of O-GlcNAc transferase activity

Protein glycosylation plays a key role in the process of life, and the related O-GlcNAc transferase (OGT) has become a sweet new target for cancer diagnosis and therapy. Herein, based on protease-protection strategy combining with the well-designed surface-enhanced Raman scattering (SERS) tags, a sensitive SERS assay was developed for specifically OGT activity determination. In the assay, a peptide containing both glycosylation and protease sites was designed. The two ends of the peptide are biotin and cysteine residues. Therefore, the SERS tags and the magnetic bead can be connected to the two ends of the peptide through Ag-S bond and affinity interaction, respectively. Peptide glycosylation occurs in the presence of OGT, which will protect the peptide from the next cleavage by proteinase K, and more SERS tags left in the surface of magnetic beads will contribute a high SERS signal. The SERS intensity was identified to be correlate with the activity of OGT during the measurement, and the minimum detection concentration of OGT down to 0.1 nmol/L was achieved with a wide linearity range from 10-10 to 10-7 mol/L. The inhibition assay and serum sample detection confirmed the feasibility of the SERS assay in practical application, which is essential for drug development and clinical diagnosis.

Automated summary

Protein glycosylation and the O-GlcNAc transferase (OGT) play vital roles in cancer diagnosis and therapy. A sensitive SERS assay was developed based on protease-protection strategy and well-designed tags, which showed high correlation with OGT activity and achieved a low detection concentration of 0.1 nmol/L. Inhibition assay and serum sample detection validated the feasibility of the assay in practical applications for drug development and clinical diagnosis.