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Off-line hyphenation of boronate affinity monolith-based extraction with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry for efficient analysis of glycoproteins/glycopeptides.

Analytica chimica acta (2014-06-15)
Zijun Bie, Yang Chen, Hengye Li, Ronghu Wu, Zhen Liu
ABSTRACT

Boronate affinity materials have attracted increasing attentions as sample enrichment platforms for glycoproteomic analysis in recent years. However, most of the boronate affinity materials that have already employed for proteomic analysis are suffering from apparent disadvantages, such as alkaline pH for binding, weak affinity, and relatively poor selectivity. Benzoboroxoles are a unique class of boronic acids which have showed excellent binding properties for the recognition of cis-diol-containing compounds. Recently, a 3-carboxy-benzoboroxole-functionalized monolithic column had been reported and it had exhibited the best selectivity and affinity as well as the lowest binding pH among all reported boronate affinity monolithic columns. In this study, an off-line hyphenation of this boronate affinity monolithic column-based extraction with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was developed and the powerfulness of this hyphenated approach in the analysis of glycoproteins and glycopeptides in complex samples was investigated. The approach was first applied to the analysis of glycopeptides in the tryptic digest of horseradish peroxidase (HRP). Totally 22 glycopeptides were identified. To the best of our knowledge, this is the best performance among all the boronic acid-functionalized materials. We further employed this approach to the analysis of intact proteins in human saliva. Totally 6 intact glycoproteins were successfully identified. As comparison, when the samples were analyzed without extraction, only a few glycopeptides were identified from the tryptic digest of HRP while no glycoproteins were found from the saliva samples.

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