Effective enrichment of glycated proteome using ultrasmall gold nanoclusters functionalized with boronic acid

Abstract

Glycated proteins play a crucial role in various biological pathways and the pathogenesis of human diseases. A comprehensive analysis of glycated proteins is essential for understanding their biological significance. However, their low abundance and heterogeneity in complex biological samples necessitate an enrichment procedure prior to their detection. Current enrichment strategies primarily rely on the boronic acid (BA) affinity method combined with functional nanoparticles; however, the effectiveness of these approaches is often suboptimal. In this study, a novel nanocluster (NC)-based enrichment material was synthesized for the first time, characterized as Au₂₂SG₁₈ functionalized with 24 BA groups, in which SG is glutathione. The functionalized BA established a reversible covalent bond with the cis-dihydroxy group through pH adjustment, enabling selective enrichment of glycated peptides. After the optimization of the enrichment protocol, we demonstrated highly sensitive and selective enrichment of standard glycopeptides using the NC-based enrichment material, exhibiting excellent reusability. Efficient enrichment was also demonstrated for the glycated proteome from human serum. These results highlight the potential of the atomically well-defined ultrasmall Au NCs as a powerful tool for high-throughput analysis of glycated peptides.