Campbell K P, Leung A T, Imagawa T
Department of Physiology and Biophysics, University of Iowa College of Medicine, Iowa City 52242.
J Cardiovasc Pharmacol. 1988;12 Suppl 4:S86-90. doi: 10.1097/00005344-198806124-00017.
The nitrendipine receptor of the voltage-dependent Ca2+ channel purified from rabbit skeletal muscle has been shown to contain four polypeptide components of 175,000, 170,000, 52,000, and 32,000 daltons. Despite the existence of a substantial amount of data on the composition of the nitrendipine receptor, little is known about the relationship between the 175,000 and 170,000 dalton subunits of the receptor and the lower molecular weight components of the receptor. A monoclonal antibody specific to the 52,000 dalton component of the receptor has now been produced. The monoclonal antibody is capable of specifically immunoprecipitating the [3H]dihydropyridine-labeled nitrendipine receptor from detergent-solubilized membranes. Immunoprecipitation experiments with 32P-labeled nitrendipine receptor have demonstrated a tight association between the 170,000 dalton nitrendipine binding subunit and the 52,000 dalton polypeptide of the receptor. Immunoblotting experiments have shown that the 52,000 dalton polypeptide copurifies with the 175,000 and 170,000 dalton subunits of the nitrendipine receptor at all stages of the purification. In addition, the higher molecular weight subunits of the receptor were not labeled by the antibody. Densitometric scanning of Coomassie blue stained SDS-polyacrylamide gels of the purified nitrendipine receptor has shown that the 175,000, 170,000, 52,000, and 32,000 dalton subunits of the nitrendipine receptor exists in a 1:1:1:1 stoichiometric ratio. In conclusion, we have demonstrated that the 52,000 dalton polypeptide is an integral and distinct subunit of the purified nitrendipine receptor of the voltage-dependent Ca2+ channel.
从兔骨骼肌中纯化得到的电压依赖性钙离子通道的尼群地平受体已被证明含有分子量分别为175,000、170,000、52,000和32,000道尔顿的四种多肽成分。尽管关于尼群地平受体的组成已有大量数据，但对于该受体分子量为175,000和170,000道尔顿的亚基与分子量较低的成分之间的关系却知之甚少。现已制备出一种针对该受体52,000道尔顿成分的单克隆抗体。该单克隆抗体能够从去污剂溶解的膜中特异性免疫沉淀[³H]二氢吡啶标记的尼群地平受体。用³²P标记的尼群地平受体进行的免疫沉淀实验表明，分子量为170,000道尔顿的尼群地平结合亚基与该受体52,000道尔顿的多肽之间存在紧密关联。免疫印迹实验表明，在纯化的各个阶段，52,000道尔顿的多肽都与尼群地平受体分子量为175,000和170,000道尔顿的亚基共同纯化。此外，该受体分子量较高的亚基未被该抗体标记。对纯化的尼群地平受体经考马斯亮蓝染色的SDS -聚丙烯酰胺凝胶进行光密度扫描显示，尼群地平受体分子量为175,000、170,000、52,000和32,000道尔顿的亚基以1:1:1:1的化学计量比存在。总之，我们已证明52,000道尔顿的多肽是纯化的电压依赖性钙离子通道尼群地平受体的一个完整且独特的亚基。
