Department of Immunology, Allergy and Infectious Diseases, University of New South Wales, St George Hospital, Sydney, Australia.
J Thromb Haemost. 2010 Aug;8(8):1754-62. doi: 10.1111/j.1538-7836.2010.03944.x.
β(2) -Glycoprotein I (β(2) GPI) is an abundant plasma protein that is closely linked to blood clotting, as it interacts with various protein and cellular components of the coagulation system. However, the role of β(2) GPI in thrombus formation is unknown. We have recently shown that β(2) GPI is susceptible to reduction by the thiol oxidoreductases thioredoxin-1 and protein disulfide isomerase, and that reduction of β(2) GPI can take place on the platelet surface.
β(2) GPI, reduced by thioredoxin-1, was labeled with the selective sulfhydryl probe N(a)-(3-maleimidylpropionyl)biocytin and subjected to mass spectrometry to identify the specific cysteines involved in the thiol exchange reaction. Binding assays were used to examine the affinity of reduced β(2) GPI for von Willebrand factor (VWF) and the effect of reduced β2GPI on glycoprotein (GP)Ibα binding to VWF. Platelet adhesion to ristocetin-activated VWF was studied in the presence of reduced β(2) GPI.
We demonstrate that the Cys288-Cys326 disulfide in domain V of β(2) GPI is the predominant disulfide reduced by thioredoxin-1. Reduced β(2) GPI in vitro displays increased binding to VWF that is dependent on disulfide bond formation. β(2) GPI reduced by thioredoxin-1, in comparison with non-reduced β(2) GPI, leads to increased binding of GPIbα to VWF and increased platelet adhesion to activated VWF.
Given the importance of thiol oxidoreductases in thrombus formation, we provide preliminary evidence that the thiol-dependent interaction of β(2) GPI with VWF may contribute to the redox regulation of platelet adhesion.
β(2)-糖蛋白 I（β(2)GPI）是一种丰富的血浆蛋白，与血液凝固密切相关，因为它与凝血系统的各种蛋白质和细胞成分相互作用。然而，β(2)GPI 在血栓形成中的作用尚不清楚。我们最近表明，β(2)GPI 易受硫氧还蛋白-1 和蛋白二硫键异构酶等硫醇氧化还原酶的还原，并且β(2)GPI 的还原可以在血小板表面发生。
用选择性巯基探针 N(a)-(3-马来酰亚丙基)生物胞素标记由硫氧还蛋白-1还原的β(2)GPI，并进行质谱分析以鉴定参与巯基交换反应的特定半胱氨酸。结合测定法用于研究还原型β(2)GPI 与血管性血友病因子（VWF）的亲和力，以及还原型β2GPI 对糖蛋白（GP）Ibα与 VWF 结合的影响。在存在还原型β(2)GPI 的情况下研究血小板对瑞斯托菌素激活的 VWF 的粘附。
我们证明β(2)GPI 结构域 V 中的 Cys288-Cys326 二硫键是由硫氧还蛋白-1还原的主要二硫键。体外还原的β(2)GPI 显示与 VWF 的结合增加，这依赖于二硫键的形成。与非还原的β(2)GPI 相比，由硫氧还蛋白-1还原的β(2)GPI 导致 GPIbα与 VWF 的结合增加，以及激活的 VWF 上血小板的粘附增加。
鉴于硫醇氧化还原酶在血栓形成中的重要性，我们提供了初步证据表明，β(2)GPI 与 VWF 的硫醇依赖性相互作用可能有助于血小板粘附的氧化还原调节。
