INVOLVEMENT OF (NA++K+)-ATPASE IN BINDING AND ACTIONS OF PALYTOXIN ON HUMAN-ERYTHROCYTES
INVOLVEMENT OF (NA++K+)-ATPASE IN BINDING AND ACTIONS OF PALYTOXIN ON HUMAN-ERYTHROCYTES
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DOI：
10.1016/0005-2736(86)90415-3
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发表时间：
1986-09-25
期刊：
影响因子：
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通讯作者：
HABERMANN, E
中科院分区：
文献类型：
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作者：
BOTTINGER, H;BERESS, L;HABERMANN, E
孢子虫毒素(约下午1点)可增加人红细胞的通透性。我们现在报道它用125I标记，然后在猪肾膜上进行亲和纯化。得到的配体与完整的红细胞快速且可逆地结合。由速度和平衡测量得到的Kd为2·10−11M，结合位点数约为2 0 0个/细胞。二价阳离子(Ca2+和GT；Sr2+和GT；Ba2+)和硼酸盐促进了结合。K+(IC50 2 MM)、哇巴因(IC50 3·10−9M)和哇巴豆胶(IC50 6·10−6M)对其有抑制作用。相反，[H]哇巴因被上述物质和浓度取代，也被孢子毒素(KI3·10−11M)取代。众所周知，狗的红细胞具有很低的(Na++K+)-ATPase活性，对孢子虫毒素具有抵抗力，并且缺乏特异的结合位点。与[~3H]哇巴因一样，~(125)I-孢霉毒素的结合也依赖于(Na~++K~+)-ATPase的状态。ATP耗尽会减少两种配体与红细胞的结合。ATP可部分恢复~(125)I-孢子虫毒素和[~3H]哇巴因与红细胞基质的结合。与[~3H]哇巴因不同，镁离子和P~(2+)不能促进125I-孢子虫毒素与红细胞基质的结合。结果表明：(A)所有已知的促进或抑制人红细胞孢子虫毒素作用的促进剂和抑制剂都是通过增强或降低其结合来实现的，(B)(Na++K+)-ATPase是孢子虫毒素的受体，哇巴因的拮抗作用在受体水平上是竞争性的。它们支持我们之前的假设，即孢霉毒素通过(Na++K+)-ATPase或其附近形成毛孔来增加人红细胞的通透性。
Palytoxin (about 1 pM) increases the permeability of human erythrocytes. We now report its radiolabeling with 125 I, followed by affinity purification on porcine kidney membranes. The resulting ligand binds fast and reversibly to intact erythrocytes. The K d from velocity and equilibrium measurements is 2· 10− 11 M, and the number of binding sites about 200 per cell. Binding is promoted by divalent cations (Ca 2+> Sr 2+> Ba 2+) and by borate. It is inhibited by K+(IC 50 2 mM), ouabain (IC 50 3· 10− 9 M) and ouabagenin (IC 50 6· 10− 6 M). Conversely,[3 H] ouabain is displaced by the substances and concentrations mentioned, and also by palytoxin (K i 3· 10− 11 M). Dog erythrocytes, which are known to possess a very low (Na++ K+)-ATPase activity, are resistant to and lack specific binding sites for palytoxin. Binding of 125 I-palytoxin, like that of [3 H] ouabain, depends on the state of (Na++ K+)-ATPase. ATP depletion decreases binding of both ligands to erythrocytes. Binding of 125 I-palytoxin and [3 H] ouabain to red cell stroma is partially restored by ATP. In contrast to [3 H] ouabain, binding of 125 I-palytoxin to red cell stroma is not promoted by Mg 2+ and P i. The data show that (a) all known promoters and inhibitors of palytoxin action on human red cells do so by enhancing or decreasing its binding,(b)(Na++ K+)-ATPase serves as a receptor for palytoxin, and (c) the antagonism by ouabain is competitive at the receptor level. They support our previous hypothesis that palytoxin increases human erythrocyte permeability by formation of pores through (Na++ K+)-ATPase or its close vicinity.