Reconstitution of rate brain /mu/ opioid receptors with purified guanine nucleotide-binding regulatory proteins, G/sub i/ and G/sub o/

Journal Article · Thu Sep 01 04:00:00 EDT 1988 · Proc. Natl. Acad. Sci. USA; (United States)

DOI:https://doi.org/10.1073/pnas.85.18.7013· OSTI ID:5603725

Ueda, Hiroshi; Harada, Hitoshi; Nozaki, Masakatsu; Katada, Toshiaki; Ui, Michio; Satoh, Masamichi; Takagi, Hiroshi

Reconstitution of purified /mu/ opioid receptors with purified guanine nucleotide-binding regulatory proteins (G proteins) was investigated. The purified /mu/ opioid receptor (pI 5.6) migrated as a single M/sub r/ 58,000 polypeptide by NaDodSO/sub 4//PAGE, a value identical to that obtained by affinity cross-linking purified /mu/ receptors. When purified /mu/ receptors were reconstituted with purified G/sub i/, the G protein that mediates the inhibition of adenylate cyclase, the displacement of (/sup 3/H)naloxone binding by (D-Ala/sup 2/,MePhe/sup 4/,Gly-ol/sup 5/)enkephalin was increased 215-fold; this increase was abolished by adding 100 /mu/M guanosine 5'-(/gamma/-thio)triphosphate. Similar increases in agonist displacement of (/sup 3/H)naloxone binding (33-fold) and its abolition by guanosine 5'-(/gamma/-thio)triphosphate were observed with G/sub o/, the G protein of unknown function, but not with the v-Ki-ras protein p.21. The stoichiometry was such that the stimulation of 1 mol of /mu/ receptor led to the binding of (/sup 3/H)guanosine 5'-(/beta/,/gamma/-imido)triphosphate to 2.5 mol of G/sub i/ or to 1.37 mol of G/sub o/. These results suggest that the purified /mu/ opioid receptor is functionally coupled to G/sub i/ and G/sub o/ in the reconstituted phospholipid vesicles.

• Research Organization: Kyoto Univ. (Japan)
• OSTI ID: 5603725
• Journal Information: Proc. Natl. Acad. Sci. USA; (United States), Journal Name: Proc. Natl. Acad. Sci. USA; (United States) Vol. 85:18; ISSN PNASA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AROMATICS
AZAARENES
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
BODY
BRAIN
CENTRAL NERVOUS SYSTEM
CENTRAL NERVOUS SYSTEM DEPRESSANTS
CHEMICAL REACTIONS
CROSS-LINKING
DAYS LIVING RADIOISOTOPES
DOSE-RESPONSE RELATIONSHIPS
DRUGS
ESTERS
GUANOSINE
HETEROCYCLIC COMPOUNDS
ISOTOPES
KINETICS
LABELLED COMPOUNDS
LIGHT NUCLEI
LIPIDS
MEMBRANE PROTEINS
MEMBRANES
MOLECULAR STRUCTURE
NARCOTICS
NERVOUS SYSTEM
NUCLEI
NUCLEOSIDES
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANS
PHOSPHOLIPIDS
PHOSPHORUS 32
PHOSPHORUS ISOTOPES
POLYMERIZATION
PROTEINS
PURIFICATION
PURINES
RADIOISOTOPES
REACTION KINETICS
RECEPTORS
RIBOSIDES
STOICHIOMETRY
TRITIUM COMPOUNDS