Advancing Ultraviolet Photodissociation Mass Spectrometry for Precision Mapping of Protein Glycosylation

PROJECT SUMMARY Glycosylation plays a structural and functional role in all fundamental features of proteins in cells.

Despite substantial evidence suggesting O-glycosylation is vital in cellular processes, the nature of O-linked glycans and the specific locations of O-glycans are not well characterized.

To a large extent, the consensus sequence motif N-X-S/T enables reliable prediction of N-glycosylation sites, but our knowledge of O-glycosylation is hampered by a lack of simple consensus motifs. As a result, our current understanding of the impact of O-glycosylation is incomplete.

A major obstacle to comprehensive characterization of O-glycosylation by mass spectrometry-based proteomic workflows is the prominent neutral loss of labile O-linked glycans using conventional collisional activated dissociation, hampering precise localization.

Moreover, no global enzyme exists that can cleave every possible O-glycan ?-O-glycosidic linkage, thwarting enzymatic glycomic analysis.

Mass spectrometry coupled with ultraviolet photodissociation (UVPD) is positioned to be an important tool in glycoproteomics by enabling residue-level resolution of PTMs of proteins implicated in human health.

UVPD can be harnessed to provide simultaneously high sequence coverage of peptides and retention of labile modifications, thus allowing O-glycan mapping and structural characterization.

My development of innovative LC-UVPD-MS strategies suitable for both targeted and global glycoproteomic applications will provide new insight into the correlation of glycosylation with disease pathways and drive new biological questions.