A Sequential Mechanism for Exosite-mediated Factor IX Activation by Factor XIa
A Sequential Mechanism for Exosite-mediated Factor IX Activation by Factor XIa
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DOI：
10.1074/jbc.m112.376343
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发表时间：
2012-11-02
影响因子：
4.8
通讯作者：
Gailani, David
中科院分区：
文献类型：
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作者：
Geng, Yipeng;Verhamme, Ingrid M.;Gailani, David
在血液凝固过程中，蛋白酶因子 XIa (fXIa) 激活因子 IX (fIX)。我们描述了这个过程的新机制。 FIX 首先在 Arg(145) 之后被切割，形成 fIX α，然后在 Arg(180) 之后被切割，形成蛋白酶 fIXa beta。 FIX α 从 fXIa 中释放，并且必须在 Arg(180) 发生后重新结合以进行裂解。 Arg(180) 后裂解的催化效率比 Arg(145) 后裂解的催化效率高 7 倍，从而限制了 fIX α 积累。 FXIa 包含四个苹果结构域 (A1-A4) 和一个催化结构域。 fXIa 上的外部位点是 fIX 结合所必需的，但是，它们的位置缺乏共识，其中描述了 A2、A3 和催化结构域上的位点。用前激肽释放酶 A3 结构域替换 A3 结构域可使 Arg(145) 和 Arg(180) 后 fIX 裂解的 K-m 分别增加 25 倍和 >= 90 倍，并显着降低 Arg(180) 后裂解的 k(cat)。使用分离的 fXIa 催化结构域或在不存在 Ca2+ 的情况下的 fXIa 获得了类似的结果。缺乏 A3 结构域的 fXIa 形式在 Arg(180) 后的裂解效率比在 Arg(145) 后的裂解效率低 15 倍，导致 fIX α 积累。替换 A2 域不会影响 fIX 激活。结果表明，fXIa 通过外部位点和 Ca2+ 介导的释放重新结合机制激活 fIX，其中第一次裂解引起的构象变化增强了第二次裂解的效率。 fIX 和 fIX α 的初始结合需要 fXIa A3 结构域上的外部位点，但不需要 A2 或催化结构域上的外部位点。
During blood coagulation, the protease factor XIa (fXIa) activates factor IX (fIX). We describe a new mechanism for this process. FIX is cleaved initially after Arg(145) to form fIX alpha, and then after Arg(180) to form the protease fIXa beta. FIX alpha is released from fXIa, and must rebind for cleavage after Arg(180) to occur. Catalytic efficiency of cleavage after Arg(180) is 7-fold greater than for cleavage after Arg(145), limiting fIX alpha accumulation. FXIa contains four apple domains (A1-A4) and a catalytic domain. Exosite(s) on fXIa are required for fIX binding, however, there is lack of consensus on their location(s), with sites on the A2, A3, and catalytic domains described. Replacing the A3 domain with the prekallikrein A3 domain increases K-m for fIX cleavage after Arg(145) and Arg(180) 25- and >= 90-fold, respectively, and markedly decreases k(cat) for cleavage after Arg(180). Similar results were obtained with the isolated fXIa catalytic domain, or fXIa in the absence of Ca2+. Forms of fXIa lacking the A3 domain exhibit 15-fold lower catalytic efficiency for cleavage after Arg(180) than for cleavage after Arg(145), resulting in fIX alpha accumulation. Replacing the A2 domain does not affect fIX activation. The results demonstrate that fXIa activates fIX by an exosite- and Ca2+-mediated release-rebind mechanism in which efficiency of the second cleavage is enhanced by conformational changes resulting from the first cleavage. Initial binding of fIX and fIX alpha requires an exosite on the fXIa A3 domain, but not the A2 or catalytic domain.