title: Solubilization and reconstitution of the mu-opioid receptor expressed in human neuronal SH-SY5Y and CHO cells.
abstract: The zwitterionic detergent CHAPS was used to solubilize the human mu-opioid receptor (hMOR) from SH-SY5Y neuroblastoma cells and recombinant hMOR-CHO (CHO-T7-hMOR) and hMOR-SH-SY5Y (SH-SY5Y-T7-hMOR) cell membranes. Agonist stimulation and G-protein activation by the mu-selective opioid agonist DAMGO ([D-Ala2, N-MePhe4, Gly-ol]-enkephalin) were recovered after removing of CHAPS after polyethylene glycol (PEG) precipitation. Binding assays show that hMOR solubilized and reconstituted this way was functional and able to interact with both agonist peptides and with G-protein. The effective solubilization and reconstitution of hMOR from mammalian cells, without truncation and extensive modification, represent an essential step toward the purification of a receptor bearing important post-translational modifications.
authors.affiliation: Centre National de la Recherche Scientifique, Institut de Pharmacologie et de Biologie Structurale, F-31077 Toulouse, France; Université de Toulouse, Université Paul Sabatier, Institut de Pharmacologie et de Biologie Structurale, F-31077 Toulouse, France. Electronic address: franck.talmont@ipbs.fr.
authors.forename: Franck
authors.initials: F
authors.lastname: Talmont
authors.affiliation: Centre National de la Recherche Scientifique, Institut de Pharmacologie et de Biologie Structurale, F-31077 Toulouse, France; Université de Toulouse, Université Paul Sabatier, Institut de Pharmacologie et de Biologie Structurale, F-31077 Toulouse, France.
authors.forename: Lionel
authors.initials: L
authors.lastname: Moulédous
authors.affiliation: Centre National de la Recherche Scientifique, Institut de Pharmacologie et de Biologie Structurale, F-31077 Toulouse, France; Université de Toulouse, Université Paul Sabatier, Institut de Pharmacologie et de Biologie Structurale, F-31077 Toulouse, France.
authors.forename: Catherine
authors.initials: C
authors.lastname: Mollereau
authors.affiliation: Centre National de la Recherche Scientifique, Institut de Pharmacologie et de Biologie Structurale, F-31077 Toulouse, France; Université de Toulouse, Université Paul Sabatier, Institut de Pharmacologie et de Biologie Structurale, F-31077 Toulouse, France.
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Zajac
mesh_terms: D000818:Animals; D016466:CHO Cells; D045744:Cell Line, Tumor; D002793:Cholic Acids / Q000737:chemistry; D003412:Cricetulus; D003902:Detergents / Q000737:chemistry; D004174:Diprenorphine / Q000737:chemistry; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)- / Q000737:chemistry; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000737:chemistry; D006801:Humans; D008024:Ligands; D009292:Narcotic Antagonists / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry; D011485:Protein Binding; D058849:Protein Refolding; D017450:Receptors, Opioid, mu / Q000819:agonists / Q000096:biosynthesis / Q000737:chemistry*; D012995:Solubility
chemical_list: D002793:Cholic Acids; D003902:Detergents; D008024:Ligands; D009292:Narcotic Antagonists; D017450:Receptors, Opioid, mu; D020875:Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; D004174:Diprenorphine; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D011092:Polyethylene Glycols; C028213:3-((3-cholamidopropyl)dimethylammonium)-1-propanesulfonate
keywords: G protein-coupled receptor; GPCR; Neuropeptide; Opioid receptors; Reconstitution; Solubilization