Regulation of von Willebrand Factor - platelet binding by Force and Interdomain I

项目摘要 动脉血栓形成发生在高流体流动的条件下，并且通过结合 血小板通过其受体糖蛋白Ib（GPIb）转化为血浆蛋白，称为von Willebrand 因子（VWF）通过A1结构域。这一过程通常被下调，以防止 自发粘附和血栓形成，但被高剪切应力激活。拟议 这项工作将检验GPIb与A1的结合被邻近的蛋白质下调的假设。 结构域或VWF内的侧翼区域，使得VWF显示结构域间自抑制。它将 还检验了与高剪切应力相关的机械力激活 通过将这些调节区域从A1结构域拉离，以减轻血小板的自动结合， 抑制作用最后一个有待检验的假设是， 血小板与VWF的结合，包括2B型血管性血友病，是通过破坏 正常的域间自抑制。VWF中的监管领域和要素将 通过克隆VWF的不同区域并确定两者的粘附行为来鉴定 可溶的和表面固定的分子。机械力和剪切应力对 这种调节将使用原子力显微镜和微流体来确定， 以定向方式固定分子。

Project Summary Arterial thrombosis occurs in conditions of high fluid flow and is mediated by the binding of platelets via their receptor Glycoprotein Ib (GPIb) to the plasma protein called von Willebrand Factor (VWF) via the A1 domain. This process is normally down-regulated to prevent spontaneous adhesion and thrombosis, but is activated by high shear stress. The proposed work will test the hypothesis that binding of GPIb to A1 is down-regulated by the neighboring domains or flanking regions within VWF, so that VWF displays interdomain auto-inhibition. It will also test the hypothesis that mechanical force associated with high shear stress activates platelet binding by pulling these regulatory regions away from the A1 domain to relieve the auto- inhibition. The final hypothesis to be tested is that conditions that increase the spontaneous binding of platelets to VWF, including type 2B von Willebrand disease, do so by damaging the normal interdomain auto-inhibition. Regulatory domains and elements within VWF will be identified by cloning different regions of VWF and determining the adhesive behavior of both soluble and surface-immoblized molecules. The effect of mechanical force and shear stress on this regulation will be determined using an atomic force microscope and microfluidics after immobilizing the molecules in an oriented fashion.