Facile Fabrication of Biomimetic Chitosan Membrane with Honeycomb-Like Structure for Enrichment of Glycosylated Peptides

Abstract

In the study of glycoproteomics with mass spectrometry, certain pretreatments of samples are required for eliminating the interference of nonglycopeptides and improving the efficiency of glycopeptides detection. Although hydrophilic interaction chromatography (HILIC) has been developed for enrichment of glycosylated peptides, a plethora of hydrophilic materials always suffered from large steric hindrance, great cost, and difficulty with modifications of high-density hydrophilic groups. In this work, a 1 mm thick biomimetic honeycomb chitosan membrane (BHCM) with honeycomb-like accessible macropores was directly prepared by the freeze-casting method as an adsorbent for HILIC. The N-glycopeptides from trypsin digests of immunoglobulin G (IgG), mixture of IgG and bovine serum albumin (BSA), and serum proteins were enriched using this material and compared with a commercial material ZIC-HILIC. The biomimetic membrane could identify as many as 32 Nglycopeptides from the IgG digest, exhibiting high sensitivity (about 50 fmol) and a wide scope for glycopeptide enrichment. A molar ratio of IgG trypsin digest to bovine serum albumin trypsin digest as low as 1/500 verified the outstanding specificity and efficiency for glycopeptide enrichment. In addition, 270 unique N-glycosylation sites of 400 unique glycopeptides from 146 glycosylated proteins were identified from the triplicate analysis of 2 μL human serum. Furthermore, 48 unique O-glycosylation sites of 278 unique O-glycopeptides were identified from the triplicate analysis of 30 μg deglycosylated fetuin digest. These results indicated that the chitosan-based membrane prepared in this work had great potential for pretreatment of samples in glycoproteomics.

Freeze Casting: From Low‐Dimensional Building Blocks to Aligned Porous Structures—A Review of Novel Materials, Methods, and Applications

“…Of further note, in addition to monolithic freeze‐cast scaffolds, macrostructures such as beads/microspheres, [ 29 ] fibers, [ 8f,30 ] meshes, [ 10b,31 ] films, [ 32 ] membranes [ 10c,33 ] and other complex morphologies [ 34 ] with aligned porous microstructures can be achieved by combining freeze‐casting with other materials processing or shaping routes such as electrospraying, [ 29e ] electrospinning, [ 30b,35 ] extrusion, [ 8f ] tape‐casting, [ 36 ] or additive manufacturing [ 37 ] during the solidification process. As the adoption of freeze casting techniques progresses, these forming techniques are the subject of growing interest towards diverse applications as discussed in a subsequent section of this review.…”

Enrichment of glycopeptides using environmentally friendly wood materials

“…In addition, different functional groups, such as amino groups, carbonyl groups or metal nanoparticles, have further been coupled to the surface of the synthesized materials to improve the hydrophilicity. [16][17][18][19][20][21][22][23] Although intensively studied, the use of the materials are limited by high cost, high chemical consumption, environmentally unfriendly starting materials, complex and time consuming procedures for both material preparation and glycopeptide enrichment, and low repeatability and reproducibility. Wood is one of the most abundant naturally available, cheap and green hydrophilic materials.…”

Comprehensive O-glycosylation analysis of the SARS-CoV-2 spike protein

“…O-linked glycopeptides can be enriched with hydrazide chemistry 18 , hydrophilic interaction chromatography 19 , and lectin affinity chromatography methods 20 . However, the oxidation step in hydrazide chemistry method destroys the intact glycan structure; the hydrophilic interaction chromatography method cannot enrich efficiently the O-linked glycopeptides with small glycans 21 .…”