Schneider T, Hofmann F
Institut für Physiologische Chemie, Medizinische Fakultät der Saarlandes, Homburg, Federal Republic of Germany.
Eur J Biochem. 1988 Jun 1;174(2):369-75. doi: 10.1111/j.1432-1033.1988.tb14107.x.
The cardiac receptor for calcium channel blockers was purified from bovine microsomal membranes which contained 235 +/- 33 fmol nimodipine-binding sites/mg protein (mean +/- SEM of nine preparations). To identify the receptor during the purification 20% of its binding sites were prelabeled with (+)[3H]PN200-110. The receptor was solubilized with 0.6% digitonin and was purified to a specific density of 157 pmol/mg using a combination of ion-exchange, wheat-germ-agglutinin-Sepharose chromatography and sucrose density gradient centrifugation. In the last sucrose gradient bound (+)[3H]PN200-110 comigrated with a 195-kDa protein. ( +/-)[3H]Azidopine and [3H]ludopamil, the photoaffinity ligands for the dihydropyridine and phenylalkylamine-binding site of the calcium channel, were incorporated specifically into the 195-kDa protein. These data indicate that the bovine cardiac receptor for calcium channel blockers is a 195-kDa protein. Its molecular mass suggests that the bovine cardiac receptor differs considerably from the rabbit skeletal muscle receptor protein for calcium channel blockers.
从牛微粒体膜中纯化出钙通道阻滞剂的心脏受体，该牛微粒体膜含有235±33 fmol尼莫地平结合位点/毫克蛋白质（九份制剂的平均值±标准误）。为了在纯化过程中鉴定该受体，其20%的结合位点用(+)[3H]PN200 - 110进行预标记。该受体用0.6%的洋地黄皂苷溶解，并通过离子交换、麦胚凝集素 - 琼脂糖凝胶层析和蔗糖密度梯度离心相结合的方法纯化至比活性为157 pmol/毫克。在最后的蔗糖梯度中，结合的(+)[3H]PN200 - 110与一种195 kDa的蛋白质共同迁移。钙通道二氢吡啶和苯烷基胺结合位点的光亲和配体(±)[3H]叠氮平与[3H]卢帕米，特异性地掺入到195 kDa的蛋白质中。这些数据表明，牛心脏钙通道阻滞剂受体是一种195 kDa的蛋白质。其分子量表明，牛心脏受体与兔骨骼肌钙通道阻滞剂受体蛋白有很大差异。
