TY - JOUR
T1 - Alteration of N-linked oligosaccharide structures of human chorionic gonadotropin β-subunit by disruption of disulfide bonds
AU - Moriwaki, Takayuki
AU - Suganuma, Nobuhiko
AU - Furuhashi, Madoka
AU - Kikkawa, Fumitaka
AU - Tomoda, Yutaka
AU - Boime, Irving
AU - Nakata, Munehiro
AU - Mizuochi, Tsuguo
N1 - Funding Information:
This study was supported in part by Grants-in-Aid from the Ministry of Education, Science, Sports and Culture and the Ministry of Health and Welfare of Japan and from the Proposal-Based Advanced Industrial Technology R&D Program of New Energy and Industrial Technology Development Organization (NEDO) of Japan.
PY - 1997
Y1 - 1997
N2 - The human chorionic gonadotropin and subunit (hCGβ) is a glycoprotein in which 12 cysteine residues pair to form six intramolecular disulfide bonds. In order to elucidate the effect of each disulfide bond on glycosylation of the molecule, we analysed structures of asparagine-linked oligosaccharides of various recombinant hCGβ produced in Chinese hamster ovary (CHO) cells: wild-type hCGβ (βWT) and mutants in which any one of the six intramolecular disulfide bonds had been disrupted by site-directed mutagenesis. SDS-PAGE analysis of βWT and these mutants before and after digestion with endoglycosidase F and H revealed structural changes in the oligosaccharide moieties of some mutants. In addition, structural analysis of oligosaccharides obtained from metabolically labeled βWT and a mutant showed that the mutant contained additional high mannose type oligosaccharides. These results suggest that elimination of a specific disulfide bond, resulting in a change in the protein conformation, disturbs the normal assembly of the mature complex type oligosaccharides in the hCGβ molecule.
AB - The human chorionic gonadotropin and subunit (hCGβ) is a glycoprotein in which 12 cysteine residues pair to form six intramolecular disulfide bonds. In order to elucidate the effect of each disulfide bond on glycosylation of the molecule, we analysed structures of asparagine-linked oligosaccharides of various recombinant hCGβ produced in Chinese hamster ovary (CHO) cells: wild-type hCGβ (βWT) and mutants in which any one of the six intramolecular disulfide bonds had been disrupted by site-directed mutagenesis. SDS-PAGE analysis of βWT and these mutants before and after digestion with endoglycosidase F and H revealed structural changes in the oligosaccharide moieties of some mutants. In addition, structural analysis of oligosaccharides obtained from metabolically labeled βWT and a mutant showed that the mutant contained additional high mannose type oligosaccharides. These results suggest that elimination of a specific disulfide bond, resulting in a change in the protein conformation, disturbs the normal assembly of the mature complex type oligosaccharides in the hCGβ molecule.
KW - Disulfide bond
KW - Human chorionic gonadotrophin
KW - N-linked oligosaccharide processing
KW - hCGβ-subunit
UR - http://www.scopus.com/inward/record.url?scp=0030972640&partnerID=8YFLogxK
U2 - 10.1023/A:1018593805890
DO - 10.1023/A:1018593805890
M3 - Article
C2 - 9111139
AN - SCOPUS:0030972640
SN - 0282-0080
VL - 14
SP - 225
EP - 229
JO - Glycoconjugate Journal
JF - Glycoconjugate Journal
IS - 2
ER -