Hamlyn J M, Harris D W, Ludens J H
Department of Physiology, University of Maryland School of Medicine, Baltimore 21201.
J Biol Chem. 1989 May 5;264(13):7395-404.
A factor having digitalis-like characteristics has been isolated from human plasma and its mechanism of action compared with the commonly used cardenolide, ouabain. The purification scheme involved dialysis of human plasma, lyophilization of dialysate, extraction of methanol-soluble components, and flash evaporation, followed by preparative, semipreparative, and analytical scale reverse-phase chromatography. One peak of biologically active material was obtained and shown to possess digitalis-like activity in assays of sodium pump activity, receptor binding, and Na,K-ATPase activity. Results from (i) the determination of the ligand conditions supporting binding, (ii) kinetics of association and dissociation from the Na,K-ATPase, (iii) affinity titration, (iv) selectivity, and (v) competition studies, when taken together, show that the endogenous digitalis-like factor is a specific inhibitor of the sodium pump that stabilizes the E2P form of the enzyme in a manner analogous to ouabain. The endogenous digitalis-like factor binds competitively in or near the receptor site for cardiac glycosides with an apparent affinity 8-20-fold greater than any known cardioactive steroid. The presence of digitalis-like activity in the circulation of individuals with no known intake of these compounds suggests that the material characterized here is an endogenous counterpart to the cardenolides. This factor may regulate sodium pump activity and provide a rationale for the existence of gene and tissue-specific forms of the Na,K-ATPase having distinct sensitivity to the cardenolides.
一种具有洋地黄样特性的因子已从人血浆中分离出来，并将其作用机制与常用的强心甾类药物哇巴因进行了比较。纯化方案包括对人血浆进行透析、将透析液冻干、提取甲醇可溶成分并进行快速蒸发，随后进行制备级、半制备级和分析级反相色谱。获得了一个具有生物活性物质的峰，并在钠泵活性、受体结合和钠钾ATP酶活性测定中显示出具有洋地黄样活性。来自（i）支持结合的配体条件的测定、（ii）与钠钾ATP酶结合和解离的动力学、（iii）亲和滴定、（iv）选择性以及（v）竞争研究的结果综合起来表明，内源性洋地黄样因子是钠泵的特异性抑制剂，其以类似于哇巴因的方式稳定酶的E2P形式。内源性洋地黄样因子在强心苷受体位点内或附近竞争性结合，其表观亲和力比任何已知的心脏活性甾体大8至20倍。在没有已知摄入这些化合物的个体循环中存在洋地黄样活性，这表明此处表征的物质是强心甾类的内源性对应物。该因子可能调节钠泵活性，并为存在对强心甾类具有不同敏感性的钠钾ATP酶的基因和组织特异性形式提供了理论依据。
