Purification and reconstitution of mu-opioid receptors in liposome.
Opioid receptors solubilized from rat brain membranes with digitonin were partially purified with a newly prepared affinity resin, AF-Amino Toyopearl, coupled with a mu-antagonist Tyr-Pro-Tyr-Tyr at the C-terminus of the peptide. The purified materials were reconstituted with an inhibitory GTP-binding protein (Gi) in liposome. From displacement analyses, two binding states, with a high and a low affinities for the mu-agonist [D-Ala2,Me-Phe4,Gly-ol5]enkephalin, were observed in the reconstituted system with Gi, only a low-affinity state was observed in the reconstituted system without Gi. The results suggested that the purified materials contained the mu-opioid receptors and could functionally couple with Gi as observed in the cell membranes.
Fujioka
T
T
Faculty of Pharmaceutical Sciences, Mukogawa Women's University, Nishinomiya, Japan.
Inoue
F
F
Sumita
S
S
Kuriyama
M
M
Hanawa
T
T
Katagi
T
T
D000595:Amino Acid Sequence; D000818:Animals; D001923:Brain Chemistry; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004745:Enkephalins / Q000737:chemistry; D008081:Liposomes; D008969:Molecular Sequence Data; D011869:Radioligand Assay; D051381:Rats; D011957:Receptors, Opioid / Q000302:isolation & purification*; D017450:Receptors, Opioid, mu; D012116:Resins, Plant
D004745:Enkephalins; D008081:Liposomes; D011957:Receptors, Opioid; D017450:Receptors, Opioid, mu; D012116:Resins, Plant