High affinity interaction between a synthetic, highly negatively charged pentasaccharide and α- or β-antithrombin is predominantly due to nonionic interactions
High affinity interaction between a synthetic, highly negatively charged pentasaccharide and α- or β-antithrombin is predominantly due to nonionic interactions
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DOI：
10.1021/bi6024929
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发表时间：
2007-03-20
期刊：
影响因子：
2.9
通讯作者：
Schedin-Weiss, Sophia
中科院分区：
文献类型：
--
作者：
Hjelm, Rebecka;Schedin-Weiss, Sophia
Idraparinux是一种合成的o -硫酸化，o -甲基化的五糖，与抗凝血酶（AT）紧密结合，从而特异性和有效地诱导促凝蛋白酶Xa因子失活。在本研究中，我们比较了这种高亲和力的五糖与α - at和β - at相互作用的亲和力和动力学，并与含有肝素天然at结合序列的合成五糖进行了比较。Idraparinux与α - at和β - at相互作用的解离平衡常数K-d分别约为0.4和0.1 nM，与普通五糖相比，亲和性提高了100倍以上。这种巨大的增强是由于在第二步结合中形成了类似400倍的构象激活复合物，而在第一步建立的相遇复合物则弱了4倍。高亲和糖和正常五糖都与AT发生了4次离子相互作用，高亲和糖在第一步只与AT建立了1次离子相互作用，在第二步补偿了3次离子相互作用，而正常五糖在每一步建立了2次离子相互作用。相比之下，非离子相互作用的亲和力（与α - at和β - at结合的K-d分别为450 nM和90 nM）明显高于正常的五糖，并且是迄今为止报道的所有at糖相互作用中最高的。因此，高亲和五糖与AT结合的总自由能中，非离子的贡献接近70%。这些发现表明，非离子相互作用可以在高电荷糖配体与蛋白质的结合中发挥主导作用，并且可以成功地利用这种生物活性配体的设计。
Idraparinux is a synthetic O-sulfated, O-methylated pentasaccharide that binds tightly to antithrombin (AT) and thereby specifically and efficiently induces the inactivation of the procoagulant protease, factor Xa. In this study, the affinity and kinetics of the interaction of this high-affinity pentasaccharide with alpha- and beta-AT were compared with those of a synthetic pentasaccharide comprising the natural AT-binding sequence of heparin. Dissociation equilibrium constants, K-d, for the interactions of Idraparinux with alpha- and beta-AT were approximately 0.4 and 0.1 nM, respectively, corresponding to an over 100-fold enhancement in affinity compared with that of the normal pentasaccharide. This large enhancement was due to a similar to 400-fold tighter conformationally activated complex formed in the second binding step, whereas the encounter complex established in the first step was similar to 4-fold weaker. The high-affinity and normal pentasaccharides both made a total of four ionic interactions with AT, although the high-affinity saccharide only established one ionic interaction in the first binding step and was compensated by three in the second step, whereas the normal pentasaccharide established two ionic interactions in each step. In contrast, the affinities of the nonionic interactions (K-d similar to 450 and 90 nM for the binding to alpha- and beta-AT, respectively) were considerably higher than those for the normal pentasaccharide and the highest of all AT-saccharide interactions reported so far. The nonionic contribution to the total free energy of the high-affinity pentasaccharide binding to AT thus amounted to similar to 70%. These findings show that nonionic interactions can play a predominant role in the binding of highly charged saccharide ligands to proteins and can be successfully exploited in the design of such biologically active ligands.