Interaction modes and approaches to glycopeptide and glycoprotein enrichment

Abstract

Protein glycosylation has received increased attention for its critical role in cell biology and diseases. Developing new methodologies to discern phenotype-dependent glycosylation will not only elucidate the mechanistic aspects of cell signaling cascades but also accelerate biomarker discovery for disease diagnosis or prognosis. In the analytical pipeline, enrichment at either the protein or peptide level is the most critical prerequisite for analyzing heterogeneous glycan composition, linkage, site occupancy and carrier proteins. Because the critical factor for choosing a suitable enrichment method is primarily a particular technique's selectivity and affinity towards target glycoproteins/glycopeptides, it is important to fully understand the working principles for the different approaches. For mechanistic insight into the enrichment protocol, we focused on the fundamental chemical and physical processes for the commonly used approaches based on: (a) glycan/peptide physicochemical properties (hydrophilic interactions, chelation/coordination chemistry) and (b) glycan-specific recognition (lectin-based affinity, covalent bond formation by hydrazide/boronic acid). Various interaction modes, such as hydrogen bonding, van der Waals interaction, multivalency, and metal- or water-mediated stabilization, are discussed in detail. In addition, we will review the design of and modifications to such methods, hyphenated approaches, and glycoproteomic applications. Finally, we will outline challenges to existing strategies and offer novel proposals for glycoproteome enrichment.

Carbohydrates on Proteins: Site-Specific Glycosylation Analysis by Mass Spectrometry

“…Similar to normal-phase liquid chromatography, stationary phases with relatively high polarity such as cellulose, sepharose, and silica bonded with amine, cyanide or amide groups have been used in HILIC (36). However, the mobile phases in HILIC are similar to those utilized in reversed-phase liquid chromatography (RPLC), which are moderately polar solvents (e.g., a mixture of acetonitrile and water) (37).…”

“…Boronic acid reacts with cis-diolcontaining glycans to form boronic esters, and the reaction can be reversed under acidic pH conditions (36,42). Consequently, materials functionalized with boronic acid groups have been used to enrich glycopeptides from human serum (43).…”

Sialic Acid-Responsive Polymeric Interface Material: From Molecular Recognition to Macroscopic Property Switching

“…However, each enrichment strategy displays its distinctive merits while usually also suffers from some inherent limitations, which has been well summarized in several excellent reviews25505152. Since the determination of great significance on SGs for immune disease diagnosis and cancer biomarkers discovery535455, researchers have never ceased to pursue high-efficiency approaches for SGs enrichment.…”

A Rapid Approach for Fabricating Boronic Acid-Functionalized Plates for On-Probe Detection of Glycoprotein and Glycopeptide

“…e decreased enrichment e ciency of glycopeptides in more complex peptide digests has also been reported by other groups. 1,13) is observation arise a concern about the inherent limited ability of boronic acid to enrichment of glycopeptides. Recently, we have made a plate coated with hydrophilic interaction chromatography (HILIC) beads, which successfully enrich 10 HRP glycopeptides in a complex peptides samples.…”