Boronic acid lectin affinity chromatography (BLAC). 2. Affinity micropartitioning-mediated comparative glycosylation profiling

Abstract

As a continuation of our work on boronic acid lectin affinity chromatography (BLAC), in this paper we introduce an automated affinity micropartitioning approach using combined boronic acid and concanavalin A (BLAC/Con A) resin-filled micropipette tips to isolate and enrich human serum glycoproteins. The N-linked oligosaccharides of the partitioned glycoproteins were removed by PNGase F enzyme digestion, followed by 8-aminopyrene-1,3,6-trisulfonic acid labeling. Capillary gel electrophoresis with blue LED-induced fluorescence detection was applied in a multiplexed format for comparative glycan profiling. The efficiency of BLAC affinity micropartitioning was compared with that of the individual lectin and pseudolectin affinity enrichment. Finally, we report on our findings in glycosylation differences in human serum samples from healthy and prostate cancer patients by applying BLAC/Con A micropipette tip-based enrichment and comparative multicapillary gel electrophoresis analysis of the released and labeled glycans.

N-Linked Glycoproteome Profiling of Seedling Leaf inBrachypodium distachyonL.

“…N-Glycosylation is an important posttranslational modification in all eukaryotes, involved in cellular localization, protein stability and solubility, signal recognition, intermolecular interactions, and the development of complex multicellular organisms. , It is commonly accepted that glycoproteins participate in protein secretory pathways via the endoplasmic reticulum (ER), Golgi apparatus, vacuole, plasma membrane, and extracellular compartments in plants. To date, various techniques, including lectin affinity chromatography, − zwitterionic–hydrophilic interaction liquid chromatography (ZIC-HILIC), − and filter-aided sample preparation (FASP) methods, − have been developed for specific enrichment of glycopeptides and glycoproteins. Through these enrichment techniques, coupled to high-accuracy mass spectrometry (MS) and related quantitative or qualitative analyses, many large-scale glycoproteome analyses have been performed in different plant species, such as Arabidopsis thaliana , , tomato, Vitis vinifera , Lotus japonicus , Hordeum vulgare , wheat, and cotton .…”

Fine-tuning the specificity of boronate affinity monoliths toward glycoproteins through pH manipulation

“…Glycoproteins can be captured in a variety of ways, such as solid-phase hydrazide capture, 5 chemoenzymatic approach, 6 hydrophilic interaction chromatography (HILIC), [7][8][9] and lectin affinity chromatography [10][11][12][13] , which is one of the most popular techniques for the isolation of glycoproteins. Recently, boronate affinity chromatography (BAC) [14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30][31][32] is becoming noteworthy for it serves as an effective extraction platform in the eld of glycoproteomics. Particularly, boronate affinity functionalized materials (predominantly in microbead, 14,21,22,26 nanoparticle 16,19,[23][24][25][27][28][29]32 and monolith 17,18,31 format) are generally used for the extraction of glycoproteins and glycopeptides, and a powerful platform was built through their combination with MALDI-TOF MS. …”

A Theoretical Model to Study the Interaction Between Boronic Acids and Insulin

“…For example, many studies have been performed to design a boronic acid containing hydrogel that releases insulin in response to an increased blood glucose concentration. 1,[3][4][5]7,[17][18][19][20][21][22][23][24][25] Insulin is a popular natural hormone produced by the pancreas of humans and being in use since last 100 years and commonly used therapeutic protein. [26][27][28][29][30][31] It has attracted the researchers from academics or industries.…”