Sulfonyl hydrazine-functionalized polymer as a specific capturer of reducing glycans from complex samples for high-throughput analysis by electrospray ionization mass spectrometry

摘要

Qualitative and quantitative studies of glycosylation patterns of various biologically important proteins represent a key field for the understanding of their complex structure-function relationships. However, the analysis of glycoprotein glycans is usually undermined by tedious sample processing steps prior to detection, including deproteination, desalting and removal of some other non-glycan impurities, which results in considerable sample loss and increased difficulty of quantitative analysis. Herein we report a facile and versatile method for the quantitative isolation of reducing glycans from complex samples using sulfonyl hydrazine-functionalized polystyrene (SHPS) beads, namely the SHPS-based glycan capturing procedure. This method allows the chemoselective and efficient condensation of the aldehyde group of reducing glycans with the active sulfonyl hydrazine group of SHPS beads under anhydrous conditions, resulting in the formation of sulfonyl hydrazone conjugates. The non-glycan components in samples, such as proteins, salts and some other impurities, can be completely removed by washing the sulfonyl hydrazone conjugates. Regeneration of the reducing glycans can be performed via mild hydrolysis of the washed sulfonyl hydrazone conjugates. This procedure is compatible with almost all the current techniques for the derivatization or detection of reducing glycans. We have obtained essential data for this method, including optimized reaction conditions, linearity and reproducibility for glycan quantitation, as well as a final glycan recovery ratio. Moreover, mass spectrometric analysis of the glycans from some complex biological samples, including milk, human blood plasma and fetal bovine serum, was achieved, demonstrating good applicability of this novel procedure.