Hirsh J K, Wu C H
Department of Molecular Pharmacology and Biological Chemistry, Northwestern University Medical School, IL 60611-3008, USA.
Toxicon. 1997 Feb;35(2):169-76. doi: 10.1016/s0041-0101(96)00136-5.
Palytoxin, the most potent animal toxin, is proposed to convert Na+/K(+)-ATPase into a cation-selective ion channel. Because of the ubiquity of pumps and channels in the living tissues used to study its mechanism of action, it is difficult to rule out that another site may be involved. In order to show that palytoxin selectively acts on Na+/K(+)-ATPase, two entirely in vitro methods were employed: (1) a cell-free expression system to synthesize the rat alpha 3 and beta 1 subunit proteins, and (2) single-channel recording of the synthetic Na+/K(+)-ATPase reconstituted in a planar lipid bilayer. Upon addition of palytoxin, single-channel currents were induced which had a conductance of 10 pS, in agreement with previous studies. In control experiments, when the cDNAs for Na+/K(+)-ATPase subunits were omitted, no single-channel currents were induced with palytoxin. Thus, the results show unambiguously that the Na+/ K(+)-ATPase is the site of action for palytoxin. Because palytoxin turns the Na+/K(+)-ATPase into a channel which can be detected by the exquisitely sensitive single-channel recording technique, the present results are the first to demonstrate the activity of in vitro synthesized Na+/K(+)-ATPase.
岩沙海葵毒素是最具毒性的动物毒素，它被认为可将钠钾ATP酶转化为阳离子选择性离子通道。由于用于研究其作用机制的活组织中普遍存在泵和通道，因此很难排除可能涉及其他位点。为了证明岩沙海葵毒素选择性作用于钠钾ATP酶，采用了两种完全体外的方法：（1）一种无细胞表达系统来合成大鼠α3和β1亚基蛋白，（2）对重构于平面脂质双分子层中的合成钠钾ATP酶进行单通道记录。加入岩沙海葵毒素后，诱导出了单通道电流，其电导为10皮安，这与之前的研究一致。在对照实验中，当省略钠钾ATP酶亚基的cDNA时，加入岩沙海葵毒素未诱导出单通道电流。因此，结果明确表明钠钾ATP酶是岩沙海葵毒素的作用位点。由于岩沙海葵毒素将钠钾ATP酶转化为一个可通过极其灵敏的单通道记录技术检测到的通道，目前的结果首次证明了体外合成的钠钾ATP酶的活性。
