[HPLC/ESI MS and MALDI/TOF MS analysis of microheterogeneity of the N-linked oligosaccharides of recombinant human erythropoietin].
OBJECTIVE
To elucidate the microheterogeneity of three N-linked oligosaccharide sites of the Chinese-made recombinant human erythropoietin (rHuEPO).


METHODS
Glu-C digestion, RP-HPLC separation, online HPLC/electrospray ionization mass spectrometry and matrix-assisted laser desorption/ionization time of flight mass spectrometry.


RESULTS
The sialic acid was analyzed directly. Almost every oligosaccharide was acetylated, the acetylation of tetraantennary + 2LacNAc + 4SA and tetraantennary + 2LacNAc + 4SA were reported.


CONCLUSIONS
The acetylation of multi-antennary oligosaccharide will improve the activity of rHuEPO in vivo. The biantennary oligosaccharide was found mainly existing at N-24. For the first time, the carbohydrate structures of each N-linked glycosylated site of Chinese-made rHuEPO were reported.
Peng
J H
JH
Analytical and Computer Center, China Pharmaceutical University, Nanjing 210009, China.
Sheng
L S
LS
Xiang
B R
BR
An
D K
DK
D002851:Chromatography, High Pressure Liquid; D004921:Erythropoietin / Q000737:chemistry*; D009844:Oligosaccharides / Q000737:chemistry*; D011994:Recombinant Proteins; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D009844:Oligosaccharides; D011994:Recombinant Proteins; D004921:Erythropoietin