Remarkable stabilization of plasminogen activator inhibitor 1 in a "molecular sandwich" complex.
Remarkable stabilization of plasminogen activator inhibitor 1 in a "molecular sandwich" complex.
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DOI：
10.1021/bi400470s
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发表时间：
2013-07-09
期刊：
影响因子：
2.9
通讯作者：
Komissarov, Andrey A.
中科院分区：
文献类型：
--
作者：
Florova, Galina;Karandashova, Sophia;Declerck, Paul J.;Idell, Steven;Komissarov, Andrey A.
纤溶酶原激活物抑制剂1 （PAI-1）水平在许多危及生命的情况下升高，通常与不利的结果相关。活性到潜伏转化的自发失活限制了PAI-1在体内的活性。虽然内源性玻璃体连接蛋白（Vn）能使PAI-1稳定1.5-2.0倍，但进一步的稳定发生在“分子三明治”复合物（MSC）中，其中限制暴露的反应性中心环的配体与PAI-1/Vn结合。研究了S195A双链尿激酶（tcuPA）和Vn对野生型（wt）糖基化（Gl-PAI-1）、非糖基化（rPAI-1）和非糖基化Q123K PAI-1（缺乏Vn结合）失活的影响。S195A tcuPA使rPAI-1、Q123K和Gl-PAI-1在37°C下的自发失活速率常数（kL）分别降低6.7、3.4和7.8倍，S195A tcuPA和Vn分别降低66.7、5.5和103.3倍。kL的温度依赖性分析表明，由于活化焓的增加和活化熵的降低，wgl - pai -1和rPAI-1在MSC中自发失活的吉布斯自由活化能分别从99.8和96.1增加到111.3和107.0 kJ/mol。与蛋白酶竞争的抗PAI-1单抗也能稳定PAI-1/Vn。MSCs对目标蛋白酶的抑制率受到S195A tcuPA或mAb的解离（k=10−4-10−3 s−1）的限制，其化学计量接近统一。体内间充质干细胞中PAI-1的稳定可能会增强不受控制的血栓形成或血管外纤维蛋白沉积，这提示了使用PAI-1抑制剂的新范式和新的潜在治疗靶点。
Plasminogen activator inhibitor 1 (PAI-1) levels are elevated in a number of life threatening conditions and often correlate with unfavorable outcomes. Spontaneous inactivation due to active to latent transition limits PAI-1 activity in vivo. While endogenous vitronectin (Vn) stabilizes PAI-1 by 1.5–2.0 fold, further stabilization occurs in a “Molecular Sandwich” Complex (MSC) where a ligand that restricts the exposed reactive center loop is bound to PAI-1/Vn. The effects of S195A two chain urokinase (tcuPA) and Vn on inactivation of wild type (wt) glycosylated (Gl-PAI-1), non-glycosylated (rPAI-1) and non- glycosylated Q123K PAI-1 (lacks Vn binding) were studied. S195A tcuPA decreased the rate constant (kL) for spontaneous inactivation at 37°C for rPAI-1, Q123K, and Gl-PAI-1 by 6.7, 3.4, and 7.8 fold, respectively, and with both S195A tcuPA and Vn by 66.7, 5.5, and 103.3 fold. Analysis of the temperature dependences of kL revealed a synergistic increase in the Gibbs free activation energy for spontaneous inactivation of wt Gl-PAI-1 and rPAI-1 in MSC from 99.8 and 96.1 to 111.3 and 107.0 kJ/mol, respectively, due to an increase in the activation enthalpy and a decrease in the activation entropy. Anti-PAI-1 mAbs competing with proteinase also stabilize PAI-1/Vn. The rate of inhibition of target proteinases by MSCs, with stoichiometry close to unity, was limited by the dissociation (k=10−4–10−3 s−1) of S195A tcuPA or mAb. The stabilization of PAI-1 in MSCs in vivo may potentiate uncontrolled thrombosis or extravascular fibrin deposition, suggesting a new paradigm for using PAI-1 inhibitors and novel potential targets for therapy.
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发表时间：
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影响因子：
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