Gly74Ser mutation in protein C causes thrombosis due to a defect in protein S-dependent anticoagulant function.
Gly74Ser mutation in protein C causes thrombosis due to a defect in protein S-dependent anticoagulant function.
复制标题
蛋白 C 中的 Gly74Ser 突变会因蛋白 S 依赖性抗凝功能缺陷而导致血栓形成。
DOI：
10.1160/th17-01-0043
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发表时间：
2017-06-28
影响因子：
6.7
通讯作者：
Rezaie AR
中科院分区：
文献类型：
--
作者：
Chen C;Yang L;Villoutreix BO;Wang X;Ding Q;Rezaie AR
蛋白C是血浆中的维生素K依赖性丝氨酸蛋白酶酶原，其在被与血栓调节蛋白（TM）复合的凝血酶激活后通过反馈环抑制机制下调凝血级联。活化蛋白C（APC）通过蛋白S依赖性的因子Va和VIIIa的降解发挥其抗凝功能。我们最近发现了一个静脉血栓患者，其血浆蛋白C抗原水平正常，但其抗凝活性仅为正常水平的34%。遗传学分析显示先证者及其弟弟均携带一种新的杂合突变c.346G>A，p.Gly74Ser（G74 S）。凝血酶生成试验表明先证者血浆中依赖TM的抗凝活性已明显受损。我们在哺乳动物细胞中表达蛋白C-G74 S，并在已建立的凝血试验中表征其特性。我们证明，蛋白C变体可以被凝血酶-TM复合物正常激活，并且所得APC突变体也表现出对FVa和FVIIIa的正常酰胺水解和蛋白水解活性。然而，发现APC变体对两种促凝血辅因子的蛋白S依赖性催化活性已显著受损。FVa降解的蛋白S浓度依赖性揭示APC变体与辅因子相互作用的能力已显著受损。对于FVa-Leiden的失活获得了相同的结果，表明APC变体对Arg-306位点的切割的蛋白S依赖性活性受到了不利影响。这些结果提供了深入了解APC中G74 S取代导致携带这种突变的先证者血栓形成的机制。
Protein C is a vitamin K-dependent serine protease zymogen in plasma which upon activation by thrombin in complex with thrombomodulin (TM) down-regulates the clotting cascade by a feedback loop inhibition mechanism. Activated protein C (APC) exerts its anticoagulant function through protein S-dependent degradation of factors Va and VIIIa. We recently identified a venous thrombosis patient whose plasma level of protein C antigen is normal, but its anticoagulant activity is only 34% of the normal level. Genetic analysis revealed that the proband and her younger brother carry a novel heterozygous mutation c.346G>A, p.Gly74Ser (G74S) in PROC. Thrombin generation assay indicated that the TM-dependent anticoagulant activity of the proband’s plasma has been significantly impaired. We expressed protein C-G74S in mammalian cells and characterized its properties in established coagulation assays. We demonstrate that the protein C variant can be normally activated by the thrombin-TM complex and the resulting APC mutant also exhibits normal amidolytic and proteolytic activities toward both FVa and FVIIIa. However, it was discovered the protein S-dependent catalytic activity of APC variant toward both procoagulant cofactors has been significantly impaired. Protein S concentration-dependence of FVa degradation revealed that the capacity of APC variant to interact with the cofactor has been markedly impaired. The same results were obtained for inactivation of FVa-Leiden suggesting that the protein S-dependent activity of APC variant toward cleavage of Arg-306 site has been adversely affected. These results provide insight into the mechanism through which G74S substitution in APC causes thrombosis in the proband carrying this mutation.