Human bradykinin B2 receptor sialylation and N-glycosylation participate with disulfide bonding in surface receptor dimerization.
G-Protein-coupled receptors (GPCRs) act on the cell surface where they recognize and convert external stimuli to modulate cellular activity and are regulated by agonist and various partner molecules. We here studied the cell surface post-translationally modified forms of a GPCR, the human bradykinin B2 receptor. This was by means of detailed molecular analysis of the cell surface forms of N-glycosylation site mutant and wild-type receptors that were treated with glycosidases, neuraminidase, and/or the reducing agent dithiothreitol or not treated before Western blotting. We found that the receptor undergoes similar glycosylation processes and similar cell surface organization in CHO-K1 and HEK 293 cells, used for stable and transient receptor expression, respectively. The receptor is present as dimers and monomers on the cell surface. The dimers result from heterologous association of differently glycosylated mature receptor molecules. Importantly, receptor sialylation and N-glycosylation participate with disulfide bonding in the stabilization of the cell surface human B2 receptor dimers.
INSERM U652, 17, rue du fer à Moulin, 75005 Paris, France. michinea@fer-a-moulin.inserm.fr
Stéphanie
S
Michineau
François
F
Alhenc-Gelas
Rabary M
RM
Rajerison
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D006224:Cricetinae; D019281:Dimerization; D004220:Disulfides / Q000378:metabolism*; D004795:Enzyme Stability; D006031:Glycosylation; D006801:Humans; D019158:N-Acetylneuraminic Acid / Q000378:metabolism*; D009844:Oligosaccharides / Q000378:metabolism; D043782:Receptor, Bradykinin B2 / Q000235:genetics / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D015203:Reproducibility of Results; D015398:Signal Transduction; D014162:Transfection
D004220:Disulfides; D009844:Oligosaccharides; D043782:Receptor, Bradykinin B2; D011956:Receptors, Cell Surface; D019158:N-Acetylneuraminic Acid