Whiting P J, Lindstrom J M
Biochemistry. 1986 Apr 22;25(8):2082-93. doi: 10.1021/bi00356a037.
Immunohistochemical studies have previously shown that both the chick brain and chick ciliary ganglion neurons contain a component which shares antigenic determinants with the main immunogenic region of the nicotinic acetylcholine receptor from electric organ and skeletal muscle. Here we describe the purification and initial characterization of this putative neuronal acetylcholine receptor. The component was purified by monoclonal antibody affinity chromatography. The solubilized component sediments on sucrose gradients as a species slightly larger than Torpedo acetylcholine receptor monomers. It was affinity labeled with bromo[3H]acetylcholine. Labeling was prevented by carbachol, but not by alpha-bungarotoxin. Two subunits could be detected in the affinity-purified component, apparent molecular weights 48 000 and 59 000. The 48 000 molecular weight subunit was bound both by a monoclonal antibody directed against the main immunogenic region of electric organ and skeletal muscle acetylcholine receptor and by antisera raised against the alpha subunit of Torpedo receptor. Evidence suggests that there are two alpha subunits in the brain component. Antisera from rats immunized with the purified brain component exhibited little or no cross-reactivity with Torpedo electric organ or chick muscle acetylcholine receptor. One antiserum did, however, specifically bind to all four subunits of Torpedo receptor. Experiments to be described elsewhere (J. Stollberg et al., unpublished results) show that antisera to the purified brain component specifically inhibit the electrophysiological function of acetylcholine receptors in chick ciliary ganglion neurons without inhibiting the function of acetylcholine receptors in chick muscle cells. All of these properties suggest that this component is a neuronal nicotinic acetylcholine receptor with limited structural homology to muscle nicotinic acetylcholine receptor.
免疫组织化学研究先前已表明，鸡脑和鸡睫状神经节神经元均含有一种成分，该成分与来自电器官和骨骼肌的烟碱型乙酰胆碱受体的主要免疫原性区域具有共同的抗原决定簇。在此，我们描述了这种假定的神经元乙酰胆碱受体的纯化及初步特性。该成分通过单克隆抗体亲和层析进行纯化。溶解的成分在蔗糖梯度上沉降，其大小略大于电鳐乙酰胆碱受体单体。它用溴化[³H]乙酰胆碱进行亲和标记。标记可被卡巴胆碱阻止，但不能被α-银环蛇毒素阻止。在亲和纯化的成分中可检测到两个亚基，表观分子量分别为48000和59000。分子量为48000的亚基既被针对电器官和骨骼肌乙酰胆碱受体主要免疫原性区域的单克隆抗体所结合，也被针对电鳐受体α亚基产生的抗血清所结合。有证据表明脑成分中有两个α亚基。用纯化的脑成分免疫的大鼠抗血清与电鳐电器官或鸡肌肉乙酰胆碱受体几乎没有或没有交叉反应。然而，一种抗血清确实能特异性结合电鳐受体的所有四个亚基。将在其他地方描述的实验（J. Stollberg等人，未发表的结果）表明，针对纯化脑成分的抗血清能特异性抑制鸡睫状神经节神经元中乙酰胆碱受体的电生理功能，而不抑制鸡肌肉细胞中乙酰胆碱受体的功能。所有这些特性表明该成分是一种神经元烟碱型乙酰胆碱受体，与肌肉烟碱型乙酰胆碱受体具有有限的结构同源性。
