THE ROLE OF N-TERMINAL GLYCOSYLATION IN THE HUMAN OXYTOCIN RECEPTOR

Citation
T. Kimura et al., THE ROLE OF N-TERMINAL GLYCOSYLATION IN THE HUMAN OXYTOCIN RECEPTOR, Molecular human reproduction, 3(11), 1997, pp. 957-963
Citations number
33
Language
INGLESE
art.tipo
Article
ISSN journal
1360-9947
Volume
3
Issue
11
Year of publication
1997
Pages
957 - 963
Database
ISI
SICI code
1360-9947(1997)3:11<957:TRONGI>2.0.ZU;2-O
Abstract
The human oxytocin receptor includes three IV-glycosylation sites in i ts extracellular N-terminal domain. We have established permanent cell -lines in which the gene for the human oxytocin receptor (OTR) has bee n introduced into HeLa cells. These cells differ by the disruption of one or more of the N-terminal N-glycosylation sites by site-directed m utagenesis of the transfected OTR constructs. The binding capacity of each transfectant, calculated per mg membrane protein, was 5-17 times higher than that of human term myometrium. The pharmacological charact eristics of the transfected wild-type OTR are very similar to those of native myometrial OTR. The mutation of N-glycosylation sites (Asn-X-S er/Thr), namely OTR-D8N15N26 (Asn(8)-->Asp(8)), -N8D15N26(Asn(15)-->As p(15)), -N8D15D26(Asn(15)-->Asp(15), Asn(26)-->Asp(26)) and -D8N15D26 (Asn(8)-->Asp(8), Asn(26)-->Asp(26)) appear to affect neither their di ssociation constant (Kd) nor the affinities for various oxytocin relat ed ligands. As a high level of cell surface binding was retained for e ach clone, receptor trafficking appears to be normal. This suggests th at the full glycosylation of OTR observed in vivo is not essential for its activity. These results indicate also that these cell lines may p rove very useful for pharmacological screening of oxytocin related pro ducts.