Abstract:

The protein content, site-specific glycosylation, and site occupancy of glycan and so on for serum amyloid A(SAA), α-1-antitrypsin(A1AT), α-2-HS-glycoprotein(A2HSG) and apolipoprotein C-Ⅲ(Apo C3) bound to high density lipoproteins(HDL) were investigated by mass spectrometry. Four kinds of trypsin digested protein standard samples were fragmented in liquid phase-quadrupole time-of-flight mass spectrometer. Some new glycosylation sites and new glycoforms were found by analyzing and matching fragment ions. For A2HSG, one O-glycosylation site and four glycoforms at that site were found, as well as 11 glycoforms at the original three sites. For Apo C3, 9 glycoforms at the original site were founded. Meanwhile, a unique peptide that used to quantify the protein were adjusted to make the quantification more accurate. Moreover, the multiple reaction monitor(MRM) behavior of peptides and glycopeptides in four proteins were studied and the site occupancy rate of each glycoform were recalculated. It is expected that our new findings in the process of exploration of site-specific glycosylation of HDL bound SAA, A1AT, A2HSG and Apo C3 will provide valuable informations in search of new HDL biomarks and will probably be practically applied in clinical detection of diseases related to HDL.

Key words: Mass spectrometry, Binding proteins to high density lipoprotein, Site-specific glycosylation, New glycoform, Multiple reaction monitoring