Soluble fibrin preparations inhibit the reaction of plasmin with alpha 2-macroglobulin. Comparison with alpha 2-antiplasmin and leupeptin.
Soluble fibrin preparations inhibit the reaction of plasmin with alpha 2-macroglobulin. Comparison with alpha 2-antiplasmin and leupeptin.
复制标题
可溶性纤维蛋白制剂抑制纤溶酶与α2-巨球蛋白的反应。
DOI：
10.1042/bj2750053
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发表时间：
1991
期刊：
影响因子：
--
通讯作者：
Gonias,SL
中科院分区：
文献类型：
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作者：
Anonick,PK;Gonias,SL
在纤维蛋白单体（Fn）和纤维蛋白原的CNBr片段（Fg-CNBr）存在下，研究了α 2-抗纤溶酶（α 2AP）、α 2-巨球蛋白（α 2 M）和亮抑酶肽（leupeptin）抑制纤溶酶的动力学。使用显色底物D-Val-L-Leu-L-Lys对硝基苯胺盐酸盐（S-2251）在连续和不连续试验中检测活性纤溶酶。这两种“纤维蛋白样”制剂均为S-2251水解的双曲线混合型抑制剂。纤溶酶与Fn和Fg-CNBr结合的解离常数（KF）分别为22 nM和17 nM。Fn和Fg-CNBr抑制纤溶酶与α 2AP的反应：抑制程度依赖于纤维蛋白浓度。在800 nM-Fn或800 nM-Fg-CNBr存在下，实验二级速率常数K″app.分别从2.4 × 10（7）M-1.s-1降至1.2 × 10（6）和5.3 × 10（5）M-1.s-1。Fn和Fg-CNBr对α 2 M抑制纤溶酶速率的影响甚至更大。“K”APP在800 nM-Fn和-Fg-CNBr存在下，值分别从4.0 × 10（5）M-1.s-1降至8.0 × 10（2）和1.3 × 10（3）M-1.s-1。相比之下，纤维蛋白制剂仅引起亮抑酶肽对纤溶酶抑制率的微小变化。k“app的最大变化。是3倍。所有纤溶酶抑制曲线均为线性，表明游离和纤维蛋白结合形式的纤溶酶在与蛋白酶抑制剂反应的过程中保持平衡。Fn和Fg-CNBr对miniplasmin与S-2251、α 2AP和α 2 M的反应无影响。当125 I-纤溶酶与Fg-CNBr孵育，然后与α 2AP和α 2 M的预混溶液反应时，Fg-CNBr没有显著改变与α 2AP结合的纤溶酶的百分比。这些实验证明纤溶酶与α 2 M的反应被纤溶酶与纤维蛋白的非共价结合抑制。我们建议，纤溶酶结合到凝块的表面是保护免受抑制的α 2 M以及α 2AP。
The kinetics of plasmin inhibition by alpha 2-antiplasmin (alpha 2AP), alpha 2-macroglobulin (alpha 2M) and leupeptin were studied in the presence of fibrin monomer (Fn) and CNBr fragments of fibrinogen (Fg-CNBr). Active plasmin was detected in continuous and discontinuous assays using the chromogenic substrate D-Val-L-Leu-L-Lys p-nitroanilide hydrochloride (S-2251). The two ‘fibrin-like’ preparations functioned as hyperbolic mixed-type inhibitors of S-2251 hydrolysis. The dissociation constants (KF) for the binding of plasmin to Fn and Fg-CNBr were 22 nM and 17 nM respectively. Fn and Fg-CNBr inhibited the reaction of plasmin with alpha 2AP: the extent of inhibition depended on the fibrin concentration. In the presence of 800 nM-Fn or 800 nM-Fg-CNBr, the experimental second-order rate constant (K″app.) was decreased from 2.4 x 10(7) M-1.s-1 to 1.2 x 10(6) and 5.3 x 10(5) M-1.s-1 respectively. The effect of Fn and Fg-CNBr on the rate of plasmin inhibition by alpha 2M was even greater. The k″app. value was decreased from 4.0 x 10(5) M-1.s-1 to 8.0 x 10(2) and 1.3 x 10(3) M-1.s-1 in the presence of 800 nM-Fn and -Fg-CNBr respectively. By contrast, the fibrin preparations caused only a small change in the rate of plasmin inhibition by leupeptin. The maximum change in k″app. was 3-fold. All plasmin inhibition curves were linear, suggesting that free and fibrin-bound forms of plasmin remained in equilibrium during the course of reaction with proteinase inhibitors. Fn and Fg-CNBr had no effect on the reaction of miniplasmin with S-2251, alpha 2AP or alpha 2M. When 125I-plasmin was incubated with Fg-CNBr and then allowed to react with a premixed solution of alpha 2AP and alpha 2M, the Fg-CNBr did not significantly change the percentage of plasmin bound to alpha 2AP. These experiments demonstrate that the reaction of plasmin with alpha 2M is inhibited by the non-covalent binding of plasmin to fibrin. We propose that plasmin bound to the surface of a clot is protected from inhibition by alpha 2M as well as by alpha 2AP.