Esam Al-Shareffi

2nd Year Medical Student

Department: School of Medicine

Graduate Program: TBD

Advisor: Dr. Daniel Raleigh (rotating)

Abstract (rotation):

Advisor: Dr. Robert Haltiwanger, Department of Biochemistry, SBU

Title:  Elucidating and characterizing O-linked glycosylation structures on NOTCH protein using permethylation and novel mass spectrometric methods

Various methods have traditionally been employed in order to isolate and characterize O-linked glycosylations on proteins of interest, including NOTCH protein. Such methods employed the incubation of cells containing the protein of interest with radioactively labeled sugars, followed by immunoprecipitation and exoglycosidase digestion. In this work, we utilize the permethylation of reductively eliminated glycans and their subsequent analysis by ESI-MS in order to isolate and characterize O-linked glycans on proteins of interest. Once validated, this method will potentially replace more traditional methods of elucidating O-linked sugars by providing a more systematic and efficient analysis, which is not dependent on prior knowledge of the sugar to be elucidated, determining specific exoglycosidases, and will not use increasingly regulated radioactive labeling chemicals and techniques. This method is also highly specific and is compatible with protein assays in which only minute (10 pmol/uL) concentrations of the glycan are available for analysis.