Department of Chemistry, University of Science and Technology of China, Hefei, 230026, People's Republic of China.
Biopolymers. 2012 Oct;97(10):818-24. doi: 10.1002/bip.22078.
Anticoagulation factor II (ACF II), a coagulation factor X- binding protein from the venom of Agkistrodon acutus has both anticoagulant and hypotensive activities. Previous studies show that ACF II binds specifically with activated factor X (FXa) in a Ca(2+) -dependent manner and inhibits intrinsic coagulation pathway. In this study, the inhibition of extrinsic coagulation pathway by ACF II was measured in vivo by prothrombin time assay and the binding of ACF II to factor IX (FIX) was investigated by native polyacrylamide gel electrophoresis and surface plasmon resonance (SPR). The results indicate that ACF II also inhibits extrinsic coagulation pathway, but does not inhibit thrombin activity. ACF II also binds with FIX with high binding affinity in a Ca(2+) -dependent manner and their maximal binding occurs at about 0.1 mM Ca(2+) . ACF II has similar binding affinity to FIX and FX as determined by SPR. Ca(2+) has a slight effect on the secondary structure of FIX as determined by circular dichroism spectroscopy. Ca(2+) ions are required to maintain in vivo function of FIX Gla domain for its recognition of ACF II. However, Ca(2+) at high concentrations (>0.1 mM) inhibits the binding of ACF II to FIX. Ca(2+) functions as a switch for the binding between ACF II and FIX. ACF II extends activated partial thromboplastin time more strongly than prothrombin time, suggesting that the binding of ACF II with FIX may play a dominant role in the anticoagulation of ACF II in vivo.
抗凝血因子 II（ACF II）是尖吻蝮蛇毒液中的一种凝血因子 X 结合蛋白，具有抗凝和降压活性。先前的研究表明，ACF II 以 Ca2+依赖性方式特异性结合激活的因子 X（FXa）并抑制内源性凝血途径。在本研究中，通过凝血酶原时间测定法在体内测定 ACF II 对外源性凝血途径的抑制作用，并通过天然聚丙烯酰胺凝胶电泳和表面等离子体共振（SPR）研究 ACF II 与因子 IX（FIX）的结合。结果表明，ACF II 也抑制外源性凝血途径，但不抑制凝血酶活性。ACF II 还以 Ca2+依赖性方式与 FIX 具有高结合亲和力，其最大结合发生在约 0.1 mM Ca2+。通过 SPR 确定，ACF II 与 FIX 和 FX 的结合亲和力相似。Ca2+对 FIX 二级结构的影响很小，如圆二色性光谱测定。Ca2+离子对于 FIX Gla 结构域在体内识别 ACF II 的功能是必需的。然而，高浓度的 Ca2+（>0.1 mM）抑制 ACF II 与 FIX 的结合。Ca2+ 作为 ACF II 和 FIX 之间结合的开关。ACF II 延长活化部分凝血活酶时间比延长凝血酶原时间更强烈，这表明 ACF II 与 FIX 的结合可能在外源性凝血途径中发挥主导作用。
