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首页> 外文期刊>FEBS Open Bio >Maturation of coagulation factor IX during Xase formation as deduced using factor VIII‐derived peptides
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Maturation of coagulation factor IX during Xase formation as deduced using factor VIII‐derived peptides

机译：X酶形成过程中凝血因子IX的成熟度（使用因子VIII衍生的肽推断）

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Blood coagulation involves extrinsic and intrinsic pathways, which merge at the activation step of blood coagulation factor X to factor Xa. This step is catalysed by the extrinsic or intrinsic Xase, which consists of a complex of factor VIIa and its cofactor tissue factor or factor IXa (FIXa) and its cofactor coagulation factor VIIIa (FVIIIa). Upon complex formation with FVIIIa, FIXa is conformationally activated to the Xase complex. However, the mechanistic understanding of this molecular recognition is limited. Here, we examined FVIIIa‐FIXa binding in the context of FIXa's activation status. Given the complexity and the labile nature of FVIIIa, we decided to employ two FVIII‐derived peptides (558‐loop, a2 peptide) to model the cofactor binding of FIX(a) using biosensor chip technology. These two FVIII peptides are known to mediate the key interactions between FVIIIa and FIXa. We found both of these cofactor mimetics as well as full‐length FVIIIa bind more tightly to zymogenic FIX than to proteolytically activated FIXa. Consequently and surprisingly, we observed that the catalytically inactive FIX zymogen can outcompete the activated FIXa from the complex with FVIIIa, resulting in an inactive, zymogenic Xase complex. By contrast, the thrombophilic Padua mutant FIXa‐R170 in complex with the protein–substrate analogue BPTI bound tighter to FVIIIa than to the zymogen form FIX‐R170L, suggesting that the active Xase complex preferentially forms in the Padua variant. Together, these results provide a mechanistic basis for the thrombophilic nature of the FIX‐R170L mutant and suggest the existence of a newly discovered safety measure within the coagulation cascade.
机译：凝血涉及外在和内在的途径，它们在凝血因子X的活化步骤合并为因子Xa。此步骤由外部或固有Xase催化，该酶由因子VIIa及其辅因子组织因子或因子IXa（FIXa）和辅因子凝血因子VIIIa（FVIIIa）的复合物组成。与FVIIIa形成复合物后，FIXa被构象活化为Xase复合物。但是，这种分子识别的机理理解是有限的。在这里，我们在FIXa激活状态的背景下检查了FVIIIa-FIXa绑定。鉴于FVIIIa的复杂性和不稳定性质，我们决定使用生物传感器芯片技术，使用两种FVIII衍生肽（558-loop，a2肽）来模拟FIX（a）的辅因子结合。已知这两个FVIII肽介导FVIIIa和FIXa之间的关键相互作用。我们发现，这些辅因子模拟物以及全长FVIIIa与酶原性FIX的结合比与蛋白水解激活的FIXa的结合更紧密。因此，令人惊讶的是，我们观察到无催化活性的FIX酶原可以从具有FVIIIa的复合物中竞争出活化的FIXa，从而导致无活性的产酶Xase复合物。相比之下，与蛋白质底物类似物BPTI结合的亲血性帕多瓦突变体FIXa-R170与FVIIIa的结合比与酶原形式FIX-R170L结合更紧密，这表明活性Xase复合物优先在帕多瓦变体中形成。总之，这些结果为FIX-R170L突变体的血栓形成性质提供了机械基础，并表明在凝血级联中存在新发现的安全措施。

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