Resin and Magnetic Nanoparticle-Based Free Radical Probes for Glycan Capture, Isolation, and Structural Characterization

Abstract

By combing the merits of solid supports and free radical activated glycan sequencing (FRAGS) reagents, we develop a multi-functional solid-supported free radical probe (SS-FRAGS), which enables glycan enrichment and characterization. SS-FRAGS comprises a solid support, free radical precursor, disulfide bond, pyridyl, and hydrazine moieties. Thioactivated resin and magnetic nanoparticles (MNPs) are chosen as the solid support to selectively capture free glycans via the hydrazine moiety, allowing for their enrichment and isolation. The disulfide bond acts as a temporary covalent linkage between the solid support and the captured glycan, allowing the release of glycans via the cleavage of the disulfide bond by dithiothreitol. The basic pyridyl functional group provides a site for the formation of a fixed charge, enabling detection by mass spectrometry and avoiding glycan rearrangement during collisional activation. The free radical precursor generates a nascent free radical upon collisional activation and thus simultaneously induces systematic and predictable fragmentation for glycan structure elucidation. A radical-driven glycan deconstruction diagram (R-DECON) is developed to visually summarize the MS 2 results and thus allow for the assembly of the glycan skeleton, making the differentiation of isobaric glycan isomers unambiguous. For application to a real-world sample, we demonstrate the efficacy of the SS-FRAGS by analyzing glycan structures enzymatically cleaved from RNase-B.

On-line enrichment of N-glycans by immobilized metal-affinity monolith for capillary electrophoresis analysis

“…However, these chromatographic supports exhibitquite low selectivities. More specific sorbents like lectins-[20], boronate-[21], hydrazine-based materials [22] or those relying on lanthanides [23] have also been described as interesting alternatives for Nglycan enrichment, most frequently before derivatization. Recently, however, a few groups reported strategies involving enrichment after derivatization and relying on the interaction between the derivatization agent attached to the glycans and the solid support [24][25].…”

Preparation of a boronate affinity-functionalized metal–organic framework material for selective recognition and separation of glycoproteins at physiological pH

“…The main specific enrichment methods for glycoproteins are the hydrazine chemical method, 4 hydrophilic interaction liquid chromatography (HILIC), 5,6 and the boronate affinity method. [7][8][9] Among them, the boronate affinity method is simple and has good ability for selective enrichment.…”

Fast and Sensitive Detection of Oligosaccharides Using Desalting Paper Spray Mass Spectrometry (DPS-MS)

“…Traditionally, electrospray ionization mass spectrometry (ESI-MS) and matrix-assisted laser desorption-ionization mass spectrometry (MALDI-MS) are often used to characterize N-linked glycans − and plant cell wall polymers . Accurate molecular mass, along with tandem mass analysis, provides unprecedented capability for glycan structure characterization and sugar sequence assignment. − However, because of signal suppression by more readily ionized compounds (e.g., salts and other solutes), it is still challenging to detect and analyze oligosaccharides in complex biological mixtures . Thus, efficient ionization and detection of these carbohydrates require enrichment and desalting steps.…”