Venter P A, Naudé R J, Oelofsen W, Swan G E
Department of Biochemistry, University of Port Elizabeth, South Africa.
Int J Biochem Cell Biol. 1997 Aug-Sep;29(8-9):1103-12. doi: 10.1016/s1357-2725(97)00046-0.
The inhibition of cardiac Na,K-ATPase by 1 alpha,2 alpha-epoxyscillirosidin is the principal cause of poisoning of cattle by the tulip, Homeria pallida. The ultimate goals of this study were to study the interaction between 1 alpha,2 alpha-epoxyscillirosidin and ovine Na,K-ATPase by means of inhibition and displacement binding studies. Ovine cardiac Na,K-ATPase was isolated in membrane-bound form by means of deoxycholate treatment, high-speed ultracentrifugation, NaI treatment and selective solubilization in Lubrol. The inhibition of ovine cardiac and commercial porcine cerebral cortex Na,K-ATPase by 1 alpha,2 alpha-epoxyscilirosidin and ouabain was studied using a discontinuous Na,K-ATPase assay. The binding of 1 alpha,2 alpha-epoxyscillirosidin, ouabain and digoxin to the above enzymes was compared using a displacement binding assay with [3H] oubain. The Lubrol-solubilized ovine cardiac Na,K-ATPase showed a specific activity of 0.3 U/mg with no ouabain insensitive activity. I50 values of 2.1 x 10(-8) and 2.7 x 10(-8) were obtained for the inhibition of this enzyme by 1 alpha,2 alpha-epoxyscillirosidin and ouabain, respectively. 1 alpha,2 alpha-Epoxyscillirosidin has a much higher KD value (1.5 x 10(-7) M), however, than ouabain (9.5 x 10(-9) M) and digoxin (1.7 x 10(-8) M) in displacement binding studies with [3H]ouabain. 1 alpha,2 alpha-Epoxyscillirosidin is a potent inhibitor of ovine cardiac Na,K-ATPase and is a slightly stronger inhibitor of the enzyme than ouabain. The anomalous result for the displacement of 1 alpha,2 alpha-epoxyscillirosidin from its receptor is either a result of different affinities that K+ has for the enzyme ouabain and enzyme-1 alpha,2 alpha-epoxyscillirosidin complexes or because of different complex stabilities of these complexes.
1α,2α -环氧异黄水仙苷对心脏钠钾 - ATP酶的抑制作用是郁金香（苍白荷马芥）致使牛中毒的主要原因。本研究的最终目的是通过抑制和置换结合研究来探究1α,2α -环氧异黄水仙苷与绵羊钠钾 - ATP酶之间的相互作用。通过脱氧胆酸盐处理、高速超速离心、碘化钠处理以及在卢勃罗尔中选择性增溶的方法，以膜结合形式分离出绵羊心脏钠钾 - ATP酶。使用不连续的钠钾 - ATP酶测定法研究了1α,2α -环氧异黄水仙苷和哇巴因对绵羊心脏及商业猪脑皮质钠钾 - ATP酶 的抑制作用。使用[³H]哇巴因置换结合测定法比较了1α,2α -环氧异黄水仙苷、哇巴因和地高辛与上述酶的结合情况。经卢勃罗尔增溶的绵羊心脏钠钾 - ATP酶的比活性为0.3 U/mg，不存在对哇巴因不敏感的活性。1α,2α -环氧异黄水仙苷和哇巴因对该酶抑制作用的半数抑制浓度（I50）值分别为2.1×10⁻⁸和2.7×10⁻⁸。然而，在使用[³H]哇巴因进行的置换结合研究中，1α,2α -环氧异黄水仙苷的解离常数（KD）值（1.5×10⁻⁷ M）比哇巴因（9.5×10⁻⁹ M）和地高辛（1.7×10⁻⁸ M）高得多。1α,2α -环氧异黄水仙苷是绵羊心脏钠钾 - ATP酶的强效抑制剂，且对该酶的抑制作用略强于哇巴因。1α,2α -环氧异黄水仙苷从其受体上被置换的异常结果，要么是由于钾离子对哇巴因酶和1α,2α -环氧异黄水仙苷 - 酶复合物具有不同亲和力，要么是由于这些复合物具有不同的复合物稳定性。
