Immunological characterization of adenosine A2A receptors in human and porcine cardiovascular tissues.
Antipeptide antibody was raised in rabbit against the sequence (361-390) of RDC-8, the presumed adenosine A2A receptor cDNA from canine. The antibody titer was estimated by solid phase radioimmunoassay. Western blot analysis under reducing conditions identified a major 45 +/- 1 kDa protein in bovine striatal membranes. This immunoreactive band was competed in the presence of excess peptide. Furthermore, the antibody recognized a single 45-kDa immunoreactive band in membranes from cells transfected with the recombinant human adenosine A2A receptors, whereas, fail to cross-react with membranes from cells transfected with recombinant rat A1 and human A3 receptors. Membranes from human and porcine coronary artery, ventricle, atria and platelets (human only) showed a major immunoreactive band at 45 +/- 1 kDa size. Under nonreducing conditions, the migration patterns of the immunoreactive bands were not altered indicating the absence of interchain disulfide bond. The 45-kDa immunoreactive band co-migrated with 2-[4-(2-¿2-[(4-aminophenyl)methylcarbonylamino]ethyl-aminocarbo nyl¿et hyl)phenyl]ethylamino-5'-Nethylcarboxamidoadenosine photoaffinity labeled A2A adenosine receptor using SANPAH as the photoaffinity cross-linker. We provide immunological evidence for the presence of A2A adenosine receptor in human cardiovascular tissues that exists as a 45-kDa monomeric protein. This study also presents evidence for the presence of A2A adenosine receptor in ventricle and atria in both human and porcine.
Marala
R B
RB
Department of Pharmacology, School of Medicine, East Carolina University, Greenville, North Carolina 27858-4354, USA.
Mustafa
S J
SJ
D000293:Adolescent; D000595:Amino Acid Sequence; D000818:Animals; D001345:Autoradiography; D001792:Blood Platelets / Q000378:metabolism; D015153:Blotting, Western; D002319:Cardiovascular System / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D003331:Coronary Vessels / Q000378:metabolism; D004285:Dogs; D005260:Female; D006801:Humans; D007120:Immunochemistry; D066298:In Vitro Techniques; D008566:Membranes / Q000378:metabolism; D008969:Molecular Sequence Data; D019920:Photoaffinity Labels; D011817:Rabbits; D051381:Rats; D018047:Receptors, Purinergic P1 / Q000378:metabolism*; D013552:Swine
D019920:Photoaffinity Labels; D018047:Receptors, Purinergic P1