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.
Department of Pharmacology, School of Medicine, East Carolina University, Greenville, North Carolina 27858-4354, USA.
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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