( sup 32 P)3 prime -O-(4-benzoyl)benzoyl ATP as a photoaffinity label for a phospholipase C-coupled P2Y-purinergic receptor
- Univ. of North Carolina, Chapel Hill (USA)
A 32P-labelled ATP analog, 3{prime}-O-(4-benzoyl)benzoyl ATP (BzATP) previously shown to be an agonist at P2Y-purinergic receptors, has been used as a probe for the P2Y-purinergic receptor on turkey erythrocyte plasma membranes. In the absence of light, (32P)BzATP bound to membranes with high affinity (KD approximately 5 nM), and in a saturable and reversible manner. The binding of (32P)BzATP was competitively inhibited by ATP and ADP analogs (2-methylthioadenosine 5'-triphosphate greater than adenosine 5{prime}-O-(2-thiodiphosphate) greater than BzATP greater than ATP greater than beta,gamma-methyleneadenosine 5{prime}-triphosphate greater than 5{prime}-adenylylimidodiphosphate) with pharmacological specificity consistent with that of a P2Y-purinergic receptor. Guanine nucleotides (guanosine 5{prime}-O-(3-thiotriphosphate) greater than GTP greater than guanosine 5{prime}-O-(2-thiodiphosphate) greater than GMP) noncompetitively inhibited the binding of radioligand. Photolysis of (32P) BzATP-prelabeled membranes resulted in incorporation of radiolabel into a protein of approximately 53,000 Da. Photolabeling was inhibited in a concentration-dependent manner by ATP and ADP analogs with a potency order characteristic for a P2Y-purinergic receptor and was modulated by guanine nucleotides. A protein of approximately 53,000 daltons was also labeled by (32P)BzATP in membranes from several other tissues known to express the P2Y-purinergic receptor. These results suggest that (32P)BzATP can be used to label covalently the P2Y-purinergic receptor and that this radioprobe will be a useful reagent for further characterization and purification of the P2Y-purinergic receptor.
- OSTI ID:
- 6622991
- Journal Information:
- Journal of Biological Chemistry; (USA), Vol. 265:23; ISSN 0021-9258
- Country of Publication:
- United States
- Language:
- English
Similar Records
Interaction of a radiolabeled agonist with cardiac muscarinic cholinergic receptors
Adenine and guanine nucleotide metabolism during platelet storage at 22 degree C
Related Subjects
RECEPTORS
CHEMICAL COMPOSITION
SYMPATHOMIMETICS
ADP
ATP
BIOCHEMICAL REACTION KINETICS
CELL MEMBRANES
ERYTHROCYTES
FOWL
LIGANDS
NUCLEOTIDES
PHOSPHORUS 32
TRACER TECHNIQUES
ANIMALS
AUTONOMIC NERVOUS SYSTEM AGENTS
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOLOGICAL MATERIALS
BIRDS
BLOOD
BLOOD CELLS
BODY FLUIDS
CELL CONSTITUENTS
DAYS LIVING RADIOISOTOPES
DRUGS
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MATERIALS
MEMBRANE PROTEINS
MEMBRANES
NUCLEI
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
PHOSPHORUS ISOTOPES
PROTEINS
RADIOISOTOPES
REACTION KINETICS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques