Functional reconstitution of prostaglandin E receptor from bovine adrenal medulla with guanine nucleotide binding proteins
Prostaglandin E/sub 2/ (PEG/sub 2/) was found to bind specifically to a 100,000 x g pellet prepared from bovine adrenal medulla. The PGE receptor was associated with a GTP-binding protein (G-protein) and could be covalently cross-linked with this G-protein by dithiobis(succinimidyl propionate) in the 100,000 x g pellet. In order to characterize the G-protein associated with the PGE receptor and reconstitute these proteins in phospholipid vesicles, the authors purified the G-protein to apparent homogeneity from the 100,000 x g pellet. The G-protein served as a substrate of pertussis toxin but differed in its ..cap alpha.. subunit from two known pertussis toxin substrate G-proteins (G/sub i/ and G/sub 0/) purified from bovine brain. The molecular weight of the ..cap alpha.. subunit was 40,000, which is between those of G/sub i/ and G/sub 0/. The purified protein was also distinguished immunologically from G/sub i/ and G/sub 0/ and was referred to as G/sub am/. Reconstitution of the PGE receptor with pure C/sub am/, G/sub i/, or G/sub 0/ in phospholipid vesicles resulted in a remarkable restoration of (/sup 3/H)PGE/sub 2/ binding activity in a GTP-dependent manner. The efficiency of these three G-proteins in this capacity was roughly equal. When pertussis toxin- or N-ethylmaleimide-treated G-proteins, instead of the native ones, were reconstituted into vesicles, the restoration of binding activity was no longer observed. These results indicate that the PGE receptor can couple functionally with G/sub am/, G/sub i/, or G/sub 0/ in phospholipid vesicles and suggest that G/sub am/ may be involved in signal transduction of the PGE receptor in bovine adrenal medulla.
- Research Organization:
- Research Development Corporation of Japan, Kyoto (USA)
- OSTI ID:
- 6771889
- Journal Information:
- J. Biol. Chem.; (United States), Vol. 263:14
- Country of Publication:
- United States
- Language:
- English
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PROSTAGLANDINS
CROSS-LINKING
RECEPTORS
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TRITIUM COMPOUNDS
ADRENAL GLANDS
CATTLE
NUCLEOTIDES
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PURIFICATION
SULFUR 35
ANIMALS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BODY
CHEMICAL REACTIONS
DAYS LIVING RADIOISOTOPES
DOMESTIC ANIMALS
ENDOCRINE GLANDS
EVEN-ODD NUCLEI
GLANDS
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MEMBRANE PROTEINS
NUCLEI
ORGANIC COMPOUNDS
ORGANS
POLYMERIZATION
RADIOISOTOPES
RUMINANTS
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VERTEBRATES
550201* - Biochemistry- Tracer Techniques