Yatani A, Hamm H, Codina J, Mazzoni M R, Birnbaumer L, Brown A M
Department of Physiology and Molecular Biophysics, Baylor College of Medicine, Houston, TX 77030.
Science. 1988 Aug 12;241(4867):828-31. doi: 10.1126/science.2457252.
The activated heterotrimeric guanine nucleotide binding (G) protein Gk, at subpicomolar concentrations, mimics muscarinic stimulation of a specific atrial potassium current. Reconstitution studies have implicated the alpha and beta gamma subunits as mediators, but subunit coupling by the endogenous G protein has not been analyzed. To study this process, a monoclonal antibody (4A) that binds to alpha k but not to beta gamma was applied to the solution bathing an inside-out patch of atrial membrane; the antibody blocked carbachol-activated currents irreversibly. The state of the endogenous Gk determined its susceptibility to block by the antibody. When agonist was absent or when activation by muscarinic stimulation was interrupted by withdrawal of guanosine triphosphate (GTP) in the presence or absence of guanosine diphosphate (GDP), the effects of the antibody did not persist. Thus, monoclonal antibody 4A blocked muscarinic activation of potassium channels by binding to the activated G protein in its holomeric form or by binding to the dissociated alpha subunit.
活化的异三聚体鸟嘌呤核苷酸结合（G）蛋白Gk在亚皮摩尔浓度下可模拟毒蕈碱对特定心房钾电流的刺激作用。重组研究表明α和βγ亚基是介导因子，但内源性G蛋白的亚基偶联尚未得到分析。为了研究这一过程，将一种与αk结合但不与βγ结合的单克隆抗体（4A）应用于浸泡着外向内式心房膜片的溶液中；该抗体不可逆地阻断了卡巴胆碱激活的电流。内源性Gk的状态决定了其被抗体阻断的敏感性。当不存在激动剂时，或者当在存在或不存在二磷酸鸟苷（GDP）的情况下通过撤去三磷酸鸟苷（GTP）中断毒蕈碱刺激的激活时，抗体的作用不会持续。因此，单克隆抗体4A通过以其全聚体形式结合活化的G蛋白或通过结合解离的α亚基来阻断毒蕈碱对钾通道的激活。
