Profiling of N- and O-glycopeptides of erythropoietin by capillary zwitterionic type of hydrophilic interaction chromatography/electrospray ionization mass spectrometry.
Capillary zwitterionic-type hydrophilic interaction chromatography (ZIC-HILIC)/ESI-MS has been applied to the Glu-C digest of recombinant human erythropoietin (rhEPO) expressed in Chinese hamster ovary (CHO) cells. N-Glycopeptides (105) and O-glycopeptides (8) were detected in a single run of the capillary ZIC-HILIC/ESI-MS analysis. Among them, N-acetyl-neuraminic acids (Neu5Ac) of N- and O-glycans were partially acetylated and some were replaced with N-glycoyl-neuraminic acid (Neu5Gc). Their retentions in the ZIC-HILIC separation can be explained to some extent with the degree of acetylation and N-glycoylation, both of which influence the hydrophilicity/hydrophobicity of the N- and O-glycan moieties of glycopeptides.
Takegawa
Yasuhiro
Y
Graduate School of Advanced Life Science, Hokkaido University, Sapporo, Japan.
Ito
Hiroki
H
Keira
Takuro
T
Deguchi
Kisaburo
K
Nakagawa
Hiroaki
H
Nishimura
Shin-Ichiro
S
D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D002853:Chromatography, Liquid / Q000379:methods*; D006224:Cricetinae; D003412:Cricetulus; D004563:Electrochemistry; D004921:Erythropoietin / Q000737:chemistry* / Q000235:genetics; D006020:Glycopeptides / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D006031:Glycosylation; D006801:Humans; D008969:Molecular Sequence Data; D015394:Molecular Structure; D011994:Recombinant Proteins; D012697:Serine Endopeptidases; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
D006020:Glycopeptides; D011994:Recombinant Proteins; D004921:Erythropoietin; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase