Factor Xa is highly protected from antithrombin-fondaparinux and antithrombin-enoxaparin when incorporated into the prothrombinase complex
Factor Xa is highly protected from antithrombin-fondaparinux and antithrombin-enoxaparin when incorporated into the prothrombinase complex
复制标题
DOI：
10.1046/j.1538-7836.2003.00254.x
复制
发表时间：
2003-06-01
影响因子：
10.4
通讯作者：
Nesheim, ME
中科院分区：
文献类型：
--
作者：
Brufatto, N;Ward, A;Nesheim, ME
抗凝血酶及其辅助因子肝素既针对凝血酶原（凝血酶）激活凝血酶原的产物，也针对负责该反应的酶(F)Xa因子。这些研究是为了量化在抗凝血酶和低分子肝素（enoxaparin, Aventis, Laval, Quebec, Canada）或肝素五糖（fondaparinux, Organon Sanofi-Synthelabo, Cypress， TX， USA）存在下，每种凝血酶原组分对FXa半衰期的影响。实验使用重组形式的凝血酶原进行，其中活性位点丝氨酸已突变为半胱氨酸，随后用荧光素标记。这个突变允许计算抗凝血酶抑制FXa的二阶速率常数，这样就消除了抗凝血酶对抗凝血酶的竞争，而考虑了凝血酶原对FXa的竞争。计算了在各种凝血酶原组分存在的情况下，抗凝血素-依诺肝素和抗凝血素-fondaparinux对FXa抑制的内在速率常数。添加磷脂对抗凝血酶抑制FXa的二级速率常数无显著影响，而添加FVa则表现出轻度保护作用。然而，进一步添加凝血酶原对FXa产生了深远的保护作用，使FXa的半衰期从1.1 s增加到353 s，在氟达肝素的情况下从0.4 s增加到42 s。未分离肝素先前也报道了类似的结果。因此，在未分离肝素、氟达肝素或依诺肝素存在的情况下，凝血酶原受到抗凝血酶的保护。
Antithrombin and its cofactor, heparin, target both the product of prothrombin activation by prothrombinase, thrombin, as well as the enzyme responsible for the reaction, factor (F)Xa. These studies were carried out to quantify the effects of each of the prothrombinase components on the half-life of FXa in the presence of antithrombin and the low-molecular-weight heparins (enoxaparin, Aventis, Laval, Quebec, Canada) or the heparin pentasaccharide (fondaparinux, Organon Sanofi-Synthelabo, Cypress, TX, USA). Experiments were carried out using a recombinant form of prothrombin in which the active site serine has been mutated to cysteine and subsequently labeled with fluorescein. This mutant allowed calculation of the second order rate constant for inhibition of FXa by antithrombin in such a way that competition for antithrombin by thrombin is eliminated and competition for FXa by prothrombin is accounted for. Intrinsic rate constants for the inhibition of FXa by anti thrombin-enoxaparin and antithrombin-fondaparinux, in the presence of the various prothrombinase components, were calculated. Addition of phospholipid had no significant effect on the second order rate constant for inhibition of FXa by antithrombin, while addition of FVa appeared to be mildly protective. Further addition of prothrombin however, caused profound protection of FXa, increasing its half-life from 1.1 to 353 s in the case of fondaparinux, and from 0.4 to 42 s in the case of enoxaparin. Similar results were reported for unfractionated heparin previously [1]. Therefore, in the presence of unfractionated heparin, fondaparinux, or enoxaparin, prothrombinase is profoundly protected from antithrombin.