N-linked glycosylation is one of the most important post-translational modification of proteins. Fucosylated N-glycans are related to many biological processes and they are commonly used as biomarkers. However, determining fucose linkages in N-glycans remains challenging. Most of the fucose linkages, particularly the linkages in the antennas of N-glycans, remain unidentified. In this work, a simple multi-stage tandem mass spectrometry based on dissociation mechanisms, namely logically derived sequence tandem mass spectrometry, was developed for the de novo determination of fucose linkages in N-glycans. Using the N-glycans extracted from pine nuts, human milk, and bee venom, we demonstrated that fucose linkages in intact N-glycans, including the 1→3 or 1→6 linkage in the core and the linkage in the α(1→3) or α(1→6) antenna were identified. The procedure does not require the N-glycan standards, and the entire process is summarized in a flowchart for automation. The CID mass spectra acquired with low resolution linear ion trap mass spectrometer (peak width 0.3 Da) are very similar to those obtained with high resolution Orbitrap mass spectrometer (R = 100,000 FWHM at m/z 400), indicating that the choice of mass analyzer does not affect carbohydrate structural elucidation.
Chen et al. (Tue,) studied this question.