Hydrophilic Porous Magnetic Graphene Oxide/Chitosan Composites for the Selective Separation and Enrichment of N-Glyco- and Phosphopeptides

Highly selective enrichment of low abundance Nglyco- and phosphopeptides from intricate biological samples before mass spectrometry detection is a vital prerequisite for indepth proteomics. A hydrophilic porous magnetic graphene oxide/ chitosan composite (mGO/CS beads) was developed and applied to selectively enrich N-glyco- and phosphopeptides in biological samples. The obtained mGO/CS beads showed strong hydrophilicity, large pore structure, and magnetic property. Based on the multivalent hydrophilic interactions between glycan group on Nglycopeptides, phosphate on phosphopeptides, and amino and hydroxyl groups on chitosan, N-glyco- and phosphopeptides could be simultaneously captured by mGO/CS beads and sequentially released by different eluents. The mGO/CS beads exhibited high sensitivity (limit of detection: 1.0 fmol mu L-1 for N-glycopeptides and 0.5 fmol mu L-1 for phosphopeptides) and high selectivity (IgG/ BSA and ss-casein/BSA tryptic digests with a mass ratio of 1:100 and 1:200). After using mGO/CS beads treatment, a total of 83 Nglycopeptides assigned to 80 N-glycoproteins and 10 phosphopeptides were identified by mass spectrometry from human serum and nonfat milk, respectively. These experimental results indicated that mGO/CS beads have great potential for highly efficient enrichment of low-abundance N-glyco- and phosphopeptides in complex biological samples.

Automated summary

This study developed a hydrophilic porous magnetic material for highly selective enrichment of low abundance N-glyco- and phosphopeptides in biological samples. The material exhibited high sensitivity and selectivity, and was successfully applied for the identification of N-glyco- and phosphopeptides from human serum and nonfat milk.