Abstract:

Protein glycosylation is a ubiquitous and important biological process involved in various molecular functions and biological pathways. It also yields important biomarkers for clinical diagnoses. However, glycopeptide analysis is challenging due to low abundance, low ionization efficiency, and glycan heterogeneity. In the present study, a method based on hydrophilic interaction liquid chromatography (HILIC) was developed for the selective enrichment of glycopeptides using a novel metal-organic framework (MOF) nanocomposite (AuGC/ZIF-8). Dual functionalization with glutathione and cysteine has resulted in an ultra-hydrophilic MOF, with synergistic effects and lower steric hindrance, providing more affinity sites for the glycopeptide enrichment. Horseradish peroxidase (HRP) was used as a model glycoprotein, and AuGC/ZIF-8 was used to enrich glycopeptides prior to analysis by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). AuGC/ZIF-8 displayed outstanding performance at enriching HRP glycopeptides, with high enrichment capacity (250 μg/mg), high selectivity in mixtures containing bovine serum albumin (BSA) (HRP-BSA (1∶200, mass ratio)), and high sensitivity at very low content (0.3 ng/μL). Thus this MOF holds promise for in-depth, comprehensive glycoproteomic and related analysis.

Key words: metal-organic framework (MOF), hydrophilic interaction liquid chromatography (HILIC), matrix-assisted laser desorption ionization time-of-flight-mass spectrometry (MALDI-TOF MS), selective enrichment, post-translational modification (PTM), glycopeptide