Tissue plasminogen activator binding to the annexin II tail domain - Direct modulation by homocysteine
Tissue plasminogen activator binding to the annexin II tail domain - Direct modulation by homocysteine
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DOI：
10.1074/jbc.273.16.9987
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发表时间：
1998-04-17
影响因子：
4.8
通讯作者：
Chait, BT
中科院分区：
文献类型：
--
作者：
Hajjar, KA;Mauri, L;Chait, BT
组织纤溶酶原激活剂通过钙调节的磷脂结合蛋白Annexin II与内皮细胞结合，这种相互作用被血栓前氨基酸同型半胱氨酸抑制。我们试图确定膜联蛋白II的组织纤溶酶原激活物结合结构域，并确定同型半胱氨酸对其调节的机制。组织型纤溶酶原激活剂与固定化膜联蛋白II的结合可被完整液相膜联蛋白II抑制，但不受其核心片段(残基25-339)的抑制。两个重叠的“尾部”多肽特异性地阻止了65%-75%的结合。组织纤溶酶原激活物结合域的定位通过六肽LCKLSL(残基7-12)的特异性抑制得到证实。表达的C9G膜联蛋白II不支持组织型纤溶酶原激活剂结合，而与C133G、C262G和C335G的结合相当于野生型膜联蛋白II。同型半胱氨酸作用后，膜联蛋白II与Cys(9)特异性结合的质量增加135+/-4-Da，组织纤溶酶原激活剂结合能力下降60-66%(I-50=11 mU M)。用[S-35]同型半胱氨酸处理培养的内皮细胞，用抗Annexin II的免疫沉淀回收二硫苏糖醇敏感的标记。这些数据通过直接阻断膜联蛋白II的组织纤溶酶原激活物结合域，为同型半胱氨酸的血栓前作用提供了一个潜在的机制。
Tissue plasminogen activator binds to endothelial cells via the calcium-regulated phospholipid-binding protein annexin II, an interaction that is inhibited by the prothrombotic amino acid homocysteine. We sought to identify the tissue plasminogen activator binding domain of annexin II and to determine the mechanism of its modulation by homocysteine. Tissue plasminogen activator binding to immobilized annexin II was inhibited by intact fluid phase annexin II but not by its "core" fragment (residues 25-339). Two overlapping "tail" peptides specifically blocked 65-75% of binding. Localization of the tissue plasminogen activator binding domain was confirmed upon specific inhibition by the hexapeptide LCKLSL (residues 7-12). Expressed C9G annexin II protein failed to support tissue plasminogen activator binding, while binding to C133G, C262G, and C335G was equivalent to that of wild type annexin II. Upon exposure to homocysteine, annexin II underwent a 135 +/- 4-Da increase in mass localizing specifically to Cys(9) and a 60-66% loss in tissue plasminogen activator-binding capacity (I-50 = 11 mu M). Upon treatment of cultured endothelial cells with [S-35]homocysteine, the dithiothreitol-sensitive label was recovered by immunoprecipitation with anti-annexin II IgG. These data provide a potential mechanism for the prothrombotic effect of homocysteine by demonstrating direct blockade of the tissue plasminogen activator binding domain of annexin II.