Fabrication of a reusable bifunctional biomimetic Ti4+-phosphorylated cellulose monolith with a coral-like structure for enrichment of phosphorylated and glycosylated peptides

Abstract

Protein phosphorylation and glycosylation, as two of the most important post-translational modifications of proteins, are of great significance to humans. The development of materials that can selectively enrich phosphopeptides and glycopeptides from complex biological samples has considerable value. In this work, an environmentally friendly bionic cellulose-derived monolithic material (TiPCM) with a coral-like structure was prepared using a thermally induced phase separation (TIPS) method and exhibited good hydrophilicity, a satisfactory porous structure, and a large quantity of titanium ions allowing it to simultaneously enrich both phosphopeptides and glycopeptides or enrich phosphopeptides alone from biological samples. The prepared TiPCM material could enrich phosphopeptides and glycopeptides, respectively, from a BSA/β-casein (molar ratio, 5000/1) and BSA/IgG (molar ratio, 1000/1) mixture demonstrating that it has excellent selectivity for phosphopeptides and glycopeptides. Meanwhile, it is worth mentioning that TiPCM materials show reusability in the enrichment of phosphopeptides, which has not been studied much in the literature. Moreover, three replicate analyses from 200 μg of tryptic digest of milk identified 119 unique phosphopeptides from 38 phosphoproteins, while identifying 180 glycopeptides from 93 unique glycoproteins and their corresponding 148 independent glycosylation sites. These results demonstrate the potential of TiPCM materials in the analysis of phosphorylated and glycosylated proteomics.