Complex formation of platelet thrombospondin with plasminogen. Modulation of activation by tissue activator.
Complex formation of platelet thrombospondin with plasminogen. Modulation of activation by tissue activator.
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血小板血小板反应蛋白与纤溶酶原的复合形成。
DOI：
10.1172/jci111578
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发表时间：
1984
期刊：
影响因子：
--
通讯作者：
Nachman,RL
中科院分区：
文献类型：
--
作者：
Silverstein,RL;Leung,LL;Harpel,PC;Nachman,RL
血小板反应蛋白（TSP）是一种多功能的血小板α-颗粒糖蛋白，可结合纤维蛋白原、纤连蛋白、肝素和富含组氨酸的糖蛋白，因此可能在调节血管表面血栓形成方面发挥重要作用。在这项研究中，我们已经证明，纯化的人血小板TSP与纯化的人纤溶酶原（Plg）形成复合物。复合物的形成，通过火箭免疫电泳的混合物的纯化的放射性标记的蛋白质进行检测。酶联免疫吸附试验（ELISA）也证明了液相Plg与吸附TSP的显着复合物的形成。复合物的形成是特定的，饱和的，并抑制过量的液相TSP，具有约35 nM的表观KD。在ELISA和火箭免疫电泳系统中，复合物的形成被抑制10 mM ε-氨基-正己酸，这意味着有一个作用的赖氨酸结合位点的Plg介导的相互作用。TSP也形成了一个复合物与纤溶酶通过ELISA检测，但没有直接抑制纤溶酶活性与合成的荧光底物或125 I-纤维蛋白平板测定。TSP，当与Plg孵育前加入到125 I-纤维蛋白板显着抑制组织纤溶酶原激活剂（TPA）的纤溶酶活性的产生的方式是钙依赖性的。在TSP存在下，TPA激活Plg的动力学研究表明，遵循Michaelis-Menten动力学，TSP作为非竞争性抑制剂。这些研究支持了TSP作为主动止血局灶性区域的多功能调节剂的假设，可以作为促血栓形成的影响，导致纤维蛋白沉积增加。图片
Thrombospondin (TSP), a multifunctional alpha-granule glycoprotein of platelets, binds fibrinogen, fibronectin, heparin, and histidine-rich glycoprotein and thus may play an important role in regulating thrombotic influences at vessel surfaces. In this study we have demonstrated that purified human platelet TSP formed a complex with purified human plasminogen (Plg). Complex formation was detected by rocket immunoelectrophoresis of mixtures of the purified radiolabeled proteins. Significant complex formation of fluid-phase Plg with adsorbed TSP was also demonstrated by enzyme-linked immunosorbent assay (ELISA). The complex formation was specific, saturable, and inhibited by excess fluid-phase TSP, with an apparent KD of approximately 35 nM. In both ELISA and rocket immunoelectrophoresis systems, complex formation was inhibited by 10 mM epsilon-amino-n-caproic acid, implying that there is a role for the lysine binding sites of Plg in mediating the interaction. TSP also formed a complex with plasmin as detected by ELISA but did not directly inhibit plasmin activity measured with a synthetic fluorometric substrate or with a 125I-fibrin plate assay. TSP, when incubated with Plg before addition to 125I-fibrin plates significantly inhibited the generation of plasmin activity by tissue plasminogen activator (TPA) in a manner that was calcium dependent. A kinetic study of Plg activation by TPA in the presence of TSP demonstrated that Michaelis-Menten kinetics were followed and that TSP acted as a noncompetitive inhibitor. These studies support the hypothesis that TSP, acting as a multifunctional regulator in focal areas of active hemostasis, could serve as a prothrombotic influence, leading to increased deposition of fibrin.Images