STRUCTURES OF CATALYTIC DOMAINS OF BLOOD PROTEINS

长期计划的总主旨是追求 凝血分子的结晶学结构 和纤溶作用，目的是将它们与 分子水平。我们的研究项目一直致力于 在过去的15年里，这一领域取得了重大进展， 确定除一个以外的所有自治域的结构 这些分子(γ-羧基谷氨酸、Kringle、表皮生长因子、 催化)，以及涉及它们的许多信息丰富的衍生研究 或其组合。我们现在将扩大工作范围，以包括其他 催化活性多域结构，这也将解决 以更一般的方式表示域-域关联。因为这些分子 与大分子底物、抑制剂和辅因子相互作用， 这种相互作用的方式将通过结构来揭示 分子络合物的测定，包括合理设计 模拟分子。有针对性的具体研究包括：(L)不活跃 α-凝血酶前体(凝血酶前2)，(2)Ser195Ala脱水 凝血酶和水合脱水凝血酶捕捉活性部位 伴随纤维蛋白原识别胞外结合的构象变化， (3)一种构象受限的非肽模拟物，基于不寻常的 凝血酶活性部位N-末端水飞蓟素相互作用，(4) 钙离子存在下Xa，(A)因子的结构测定 (B)与组织因子途径的第二个kunitz结构域形成复合体 抑制剂和(5)有效因子Xa的结构测定 抑制剂扁虱抗凝蛋白。结晶搜索也是 正在与因子VIIa和IX以及蛋白C和激活的蛋白C一起 将工作范围扩大到凝血的其他领域 卡斯卡德。上述许多具有衍射性的单晶 并对它们的X射线衍射谱进行了初步研究 已经测量了三个强度数据集，这些数据集表明 成功的概率。

The general thrust of the long range plans is to pursue crystallographically structural aspects of molecules of blood coagulation and fibrinolysis with the aim of relating them to functionality at the molecular level. Our research program has been exclusively devoted to the area for about the past 15 years and has made significant progress by determining the structures of all but one of the autonomous domains of these molecules (gamma-carboxyglutamic, kringle, epidermal growth factor, catalytic), along with many informative derivative studies involving them or combinations thereof. We will now extend the work to include additional catalytically active multidomain structures, which will also address domain-domain associations in a more general way. Since these molecules interact with macromolecular substrates, inhibitors and cofactors, the manner of such interactions will be revealed through the structure determinations of molecular complexes, including rationally designed mimetic molecules. Specific studies targeted include: (l) the inactive precursor of alpha-thrombin (prethrombin 2), (2) Serl95Ala anhydro thrombin and hirugen-anhydro thrombin to capture the active site conformational change accompanying fibrinogen recognition exosite binding, (3) a conformationally restricted non-peptide mimetic based on the unusual N-terminal hirudin interaction in the active site of thrombin, (4) the structure determination of Factor Xa, (a) in the presence of Ca+2 ions and (b) in complex with the second Kunitz domain of tissue factor pathway inhibitor and (5) the structure determination of the potent Factor Xa inhibitor tick anticoagulant protein. Crystallization searches are also underway with Factors VIIa and IX and protein C and activated protein C to extend the scope of the work to other areas of the blood coagulation cascade. Diffraction quality single crystals of many of the above have been grown and their X-ray diffraction patterns examined in a preliminary way and three intensity data sets have been measured that suggest a high probability of success.