Structural study of the N-linked oligosaccharides of hepatocyte growth factor by two-dimensional sugar mapping.
The structures of the N-linked oligosaccharides on recombinant human hepatocyte growth factor (rh-HGF) expressed by Chinese hamster ovary (CHO) cells were studied by two-dimensional sugar mapping. The oligosaccharides released from the glycopeptides by peptide: N-glycosidase F (PNGase F) treatment were tagged with 2-aminopyridine at the reducing ends. The alpha-chain was linked by biantennary, triantennary, and tetraantennary oligosaccharides, but the dominant oligosaccharides linking the beta-chain were biantennary (> 85%). There was no significant difference in oligosaccharide structures between the two glycosylation sites on each chain, that is, Asn263 and Asn371 on the alpha-chain, and Asn535 and Asn622 on the beta-chain. The linkage of sialic acid to the non-reducing terminal galactose was identified as NeuAc alpha(2-3) by 1H-NMR spectrometry. The structures of the N-linked oligosaccharides from rat HGF were also studied. Triantennary oligosaccharides were obtained from the alpha-chain and a biantennary oligosaccharide was obtained from the beta-chain. This result indicates that the alpha-chain is also linked by higher branched oligosaccharides than the beta-chain in rat HGF.
Hara
H
H
Pharmaceuticals Research Center of Toyobo Co., Ltd., Otsu.
Nakae
Y
Y
Sogabe
T
T
Ihara
I
I
Ueno
S
S
Sakai
H
H
Inoue
H
H
Shimizu
S
S
Nakamura
T
T
Shimizu
N
N
D000631:Aminopyridines / Q000737:chemistry; D000818:Animals; D016466:CHO Cells; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D006224:Cricetinae; D003412:Cricetulus; D017228:Hepatocyte Growth Factor / Q000737:chemistry*; D008099:Liver / Q000737:chemistry*; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000737:chemistry*; D051381:Rats; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence
D000631:Aminopyridines; D009844:Oligosaccharides; D011994:Recombinant Proteins; D017228:Hepatocyte Growth Factor; C032439:alpha-aminopyridine