Characterization of myocardial extracellular ATP receptors by photoaffinity labelling and functional assays

FEBS Letters

Powers, Kit¹  Scarpa, Antonio¹  Giannattasio, Bartolo¹ 

[1] Department of Physiology and Biophysics, Case Western Reserve University, 2119 Abington Rd., Cleveland, OH 44116, USA

关键词: ATP receptor;    Photolabelling;    Myocyte;    8-Azido-ATP;    ATPγS;    adenosine-5′-O-(3-thiotriphosphate);    2Az-ATP;    2-azido-adenosine-5′-triphosphate;    8Az-ATP;    8-azido-adenosine-5′-triphosphate;    Bz-ATP;    3′-O-(4-benzoyl)benzoyl-[α-32P]adenosine-5′-triphosphate;    BSA;    bovine serum albumin;    2-meSATP;    2-methylthio- adenosine-5′-triphosphate;    fura2/AM;    fura2-acetoxymethyl ester;    GTP;    guanosine-5′-triphosphate;    SDS;    sodium dodecyl sulfate;    UTP;    uridine-5′-triphosphate;   

DOI  :  10.1016/0014-5793(92)80582-2

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

Extracellular ATP receptors in rat ventricular myocytes were investigated through intact cell photolabelling followed by protein isolation. 8-Azido-ATP (8Az-ATP) was used for labelling under specific conditions determined by parallel functional studies. In those studies ATP-induced cytosolic Ca²⁺ transients were irreversibly and specifically inhibited by UV-photolyzed 8Az-ATP, but not by 2-azido-ATP (2Az-ATP), even in the presence of high concentrations of phosphonucleotides not affecting myocardial ATP receptors. Under those conditions background labelling is minimized and radioactive 8Az-ATP specifically labels a band of 45–48 kDa on a SDS gel. Labelling under the above conditions in the presence of ATPγS or 2-methylthio-ATP (2-meSATP), which are distinct for two functionally different cardiac ATP receptors, shows two different proteins within the same band consistent with the possible labelling of these two receptors.

【 授权许可】

Unknown