Carbohydrates presented on cellular surfaces take on important biological functions in signaling and intermolecular recognitions. Sequencing of carbohydrates is crucial to identify their biological functions because they modulate the biological activity of proteins through specific interactions. However, polydispersity and structural complexity caused by a non-template-driven biosynthesis render carbohydrate sequencing challenging. These challenges are chiefly related to isomers of precursors and fragment ions produced by tandem-mass spectrometry. Here, we apply our recently developed tandem trapped ion mobility spectrometer-mass spectrometer (tTIMS/MS) to characterize carbohydrate sequences. Our tTIMS/MS method allows a variety of measurement workflow that includes mobility-separation, mobility-selection, and collisional-dissociation (CID) of precursor ions in a first CID stage. Additionally, the fragment ions produced by the first CID stage can be mobility-separated, mass-selected, and subsequently collisionally-dissociated followed by TOF-mass analysis. These multiple, sequential mobility analyses enable mobility-analysis of precursor as well as of fragment ions and effective MS3 experiments. Our contribution here discusses the ability of the tTIMS/MS method to characterize carbohydrate structures based on the isomeric trisaccharide systems raffinose, melezitose, and maltotriose. Additionally, we discuss application of tTIMS/MS to sequence larger carbohydrates systems related to glycosaminoglycans including the pentasaccharide Arixtra and octasaccharides of dermatan sulfate.

Structural characterization of carbohydrate oligosaccharides using tandem-trapped ion mobility spectrometry – mass spectrometry (tTIMS/MS)

ACS Spring 2022

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