Laboratório de Tecido Conjuntivo, Hospital Universitário Clementino Fraga Filho, Universidade Federal do Rio de Janeiro, Cidade Universitária, Ilha do Fundão, Rio de Janeiro, Brazil.
Thromb Haemost. 2009 Dec;102(6):1183-93. doi: 10.1160/TH09-04-0273.
An algal sulfated galactan has high anticoagulant and antithrombotic activities. Its serpin-dependent anticoagulant action is due to promoting thrombin and factor (F)Xa inhibition by antithrombin and heparin cofactor II. Here, we evaluated the anticoagulant effect of the algal sulfated galactan using serpin-free plasma. In contrast to heparin, the sulfated galactan is still able to prolong coagulation time and delay thrombin and FXa generation in serpin-free plasma. We further investigated this effect using purified blood coagulation proteins, discovering that sulfated galactan inhibits the intrinsic tenase and prothrombinase complexes, which are critical for FXa and thrombin generation, respectively. We also investigated the mechanism by which sulfated galactan promotes FXa inhibition by antithrombin using specific recombinant mutants of the protease. We show that sulfated galactan interacts with the heparin-binding exosite of FXa and Arg-236 and Lys-240 of this site are critical residues for this interaction, as observed for heparin. Thus, sulfated galactan and heparin have similar high-affinity and specificity for interaction with FXa, though they have differences in their chemical structures. Similar to heparin, the ability of sulfated galactan to potentiate FXa inhibition by antithrombin is calcium-dependent. However, in contrast to heparin, this effect is not entirely dependent on the conformation of the gamma-carboxyglutamic acid-rich domain of the protease. In conclusion, sulfated galactan and heparin have some similar effects on blood coagulation, but also differ significantly at the molecular level. This sulfated galactan opens new perspective for the development of antithrombotic drugs.
一种藻类硫酸化半乳聚糖具有很高的抗凝和抗血栓活性。其丝氨酸蛋白酶抑制剂依赖性抗凝作用是由于抗凝血酶和肝素辅因子 II 促进凝血酶和因子 (F)Xa 的抑制。在这里，我们使用无丝氨酸蛋白酶血浆评估了藻类硫酸化半乳聚糖的抗凝作用。与肝素不同，硫酸化半乳聚糖仍然能够延长无丝氨酸蛋白酶血浆的凝血时间，并延迟凝血酶和 FXa 的产生。我们使用纯化的血液凝血蛋白进一步研究了这种作用，发现硫酸化半乳聚糖抑制内在的 tenase 和 prothrombinase 复合物，这分别对 FXa 和凝血酶的产生至关重要。我们还研究了硫酸化半乳聚糖通过特定的重组蛋白酶突变体促进抗凝血酶抑制 FXa 的机制。我们表明，硫酸化半乳聚糖与 FXa 的肝素结合外位相互作用，Arg-236 和该位点的 Lys-240 是该相互作用的关键残基，如肝素一样。因此，硫酸化半乳聚糖和肝素与 FXa 的相互作用具有相似的高亲和力和特异性，尽管它们在化学结构上存在差异。与肝素相似，硫酸化半乳聚糖增强抗凝血酶抑制 FXa 的能力依赖于钙。然而，与肝素不同的是，这种作用不完全依赖于蛋白酶的 γ-羧基谷氨酸丰富结构域的构象。总之，硫酸化半乳聚糖和肝素对血液凝固有一些相似的作用，但在分子水平上也有很大的差异。这种硫酸化半乳聚糖为开发抗血栓药物开辟了新的前景。
