Modulation of thrombin-induced platelet aggregation by inhibition of calpain by a synthetic peptide derived from the thiol-protease inhibitory sequence of kininogens and S-(3-nitro-2-pyridinesulfenyl)-cysteine.
Modulation of thrombin-induced platelet aggregation by inhibition of calpain by a synthetic peptide derived from the thiol-protease inhibitory sequence of kininogens and S-(3-nitro-2-pyridinesulfenyl)-cysteine.
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通过由激肽原和 S-(3-硝基-2-吡啶硫基)-半胱氨酸的硫醇蛋白酶抑制序列衍生的合成肽抑制钙蛋白酶来调节凝血酶诱导的血小板聚集。
DOI：
10.1111/j.1432-1033.1993.tb17916.x
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
Colman,RW
中科院分区：
文献类型：
--
作者：
Puri,RN;Matsueda,R;Umeyama,H;Bradford,HN;Colman,RW
凝血酶诱导的血小板聚集已被认为在心肌梗死血管成形术溶栓治疗后再闭塞中发挥重要作用。我们以前证明，凝血酶洗涤血小板的聚集伴随着凝集素，一个假定的ADP受体的裂解，这些事件是间接介导的钙蛋白酶，外膜表面上表达。高分子量激肽原（HK）在其重链中含有结构域2，该结构域2负责其作为血小板钙蛋白酶的强效抑制剂的作用。HK重链的结构域3直接抑制凝血酶与血小板的结合，混淆了使用整个分子的机制研究。此外，HK是一种120 kDa的蛋白酶，不适合作为潜在的药理学试剂。存在于HK及其进化前体半胱氨酸蛋白酶抑制剂中的高度保守序列Gln-瓦尔-瓦尔-Ala-Gly被认为参与半胱氨酸蛋白酶的结合，但其本身不具有抑制性。合成了一种亲和类似物Phe-Gln-瓦尔-瓦尔-Cys（Npys）-Gly-NH 2（Npys，3-硝基-2-亚硫基吡啶），P1，对应于HK中的巯基蛋白酶结合序列，并含有一个配体Npys，可以与钙蛋白酶活性位点的游离巯基反应。P1是血小板钙蛋白酶的不可逆抑制剂。P1选择性抑制凝血酶诱导的洗涤血小板和血浆中血小板的聚集，但不抑制其他血小板激动剂的聚集作用。P1不抑制凝血酶的酰胺水解活性和凝血活性。与HK不同，P1不抑制凝血酶与洗涤血小板的结合。P1不抑制凝血酶诱导的血小板形状变化。P1既不升高细胞内cAMP水平，也不干扰凝血酶拮抗前列腺素I2类似物伊洛前列素诱导的细胞内cAMP水平升高的能力。P1的设计和合成可以开发一类新的抑制剂，其选择性地阻断凝血酶诱导的血小板聚集，同时保留这种病理生理蛋白酶的其他功能，并且不抑制其他血小板激动剂的作用。
Thrombin‐induced platelet aggregation has been suggested to play an important role in reocclusion following thrombolytic therapy of angioplasty for treatment of myocardial infarction. We previously demonstrated that aggregation of washed platelets by thrombin is accompanied by cleavage of aggregin, a putative ADP receptor, and that these events are indirectly mediated by calpain, expressed on the surface of the external membrane. High‐molecular‐mass kininogen (HK) contains, in its heavy chain, domain 2, which is responsible for its action as a potent inhibitor of platelet calpain. Domain 3 of the heavy chain of HK directly inhibits binding of thrombin to platelets, confounding mechanistic studies using the entire molecule. Moreover, HK, a protease of 120 kDa, is unsuitable as a potential pharmacological agent. The highly conserved sequence Gln‐Val‐Val‐Ala‐Gly, present in HK and its evolutionary precursors, the cystatins, is thought to be involved in the binding of cysteine proteases but is, itself, not inhibitory. An affinity analog, Phe‐Gln‐Val‐Val‐Cys(Npys)‐Gly‐NH2(Npys, 3‐nitro‐2‐sulfenylpyridine), P1, corresponding to the thiol‐protease‐binding sequence in HK and containing a ligand, Npys, that can react with the free sulfhydryl group in the active site of calpain, was synthesized. P1 was an irreversible inhibitor of platelet calpain. P1 selectively inhibited thrombin‐induced aggregation of washed platelets and platelets in plasma, but did not inhibit the aggregatory effects of other platelet agonists. P1 did not inhibit the amidolytic activity and coagulant activity of thrombin. Unlike HK, P1 did not inhibit binding of thrombin to washed platelets. P1 did not inhibit thrombin‐induced platelet‐shape change. P1 neither raised intracellular levels of cAMP nor did it interfere with the ability of thrombin to antagonize the rise in intracellular levels of cAMP induced by iloprost, an analog of prostaglandin I2. The design and synthesis of P1 could leave to the development of a new class of inhibitors that selectively block thrombin‐induced platelet aggregation while sparing other functions of this pathophysiological protease and without inhibiting the action of other platelet agonists.