STRUCTURAL STUDIES ON PROTEINS INVOLVED IN HEMOSTASIS

凝血酶是止血的中枢调节蛋白，定义为 遏制循环的血液。凝血酶激活不当是一种 过早死亡的主要原因，因为它是大多数 心脏病发作和中风。这个项目的一个长期目标是 通过确定凝血酶的许多不同功能来使其合理化 具有和不具有底物类似物、抑制剂、 和效应器。具体目标是：(1)完成建设 并在2.4A分辨率下对牛α-凝血酶模型进行了改进。 (2)比较其衍生物β-150-凝血酶的结构 α-凝血酶在第150位切割，使用和不使用共价抑制剂 在活动站点中。(3)通过分子置换确定 α-凝血酶或β-150-凝血酶的结构，与重要的 抑制剂。(4)结晶与凝血酶相关的蛋白质，例如 重组凝血酶原，并用分子方法解析其结构 取代或重原子衍生品。 血小板因子4(PF4)，它是由激活的血小板在 凝血过程包括促凝血、抗肝素、趋化作用和 免疫调节活动。它也是一个新的 同源蛋白超家族参与止血、炎症、 和细胞生长。这一分项目的具体目标是：(1) 完成了天然PF4的结构提纯和分析。(2) 通过建模和开发PF4/肝素复合体的结构 结晶学研究(3)确定重组蛋白的结构 现在使用晶体进行分子置换的人PF4及其变异体 在手中。(4)结晶并确定其同系物的结构 作为生长因子、有丝分裂原和伤口愈合剂的PF4。 (5)结晶和测定选定的肝素结合体的结构 与PF4不同源的蛋白质。

Thrombin is the central regulatory protein in hemostasis, defined as the containment of circulating blood. Improper activation of thrombin is a leading cause of premature death in that it is the immediate cause of most heart attacks and strokes. A long term goal of this project is to rationalize the many diverse functions of thrombin by determining its three-dimensional structure with and without substrate analogs, inhibitors, and effectors. The specific goals are: (1) To complete the construction and refinement at 2.4 A resolution of the model of bovine alpha-thrombin. (2) To compare the structure of beta150-thrombin, which is a derivative of alpha-thrombin cut at position 150, with and without a covalent inhibitor in the active site. (3) To determine by molecular replacement the structures of alpha-thrombin or beta150-thrombin, complexed with important inhibitors. (4.) To crystallize proteins related to thrombin, such as recombinant prothrombin, and solve their structures by molecular replacement or heavy atom derivatives. Platelet factor 4 (PF4), which is released by activated platelets during the clotting process has procoagulation, antiheparin, chemotactic, and immunoregulatory activities. It is also an architypical member of a new superfamily of homologous proteins involved in hemostasis, inflammation, and cell growth. The specific aims of this subproject are: (1) To complete the refinement and analysis of the structure of native PF4. (2) To develop a structure for the PF4 / heparin complex from modeling and crystallographic studies. (3) To determine the structure of recombinant human PF4 and its variants by molecular replacement using the crystals now in hand. (4) To crystallize and determine the structures of homologs of PF4 which function as growth factors, mitogens, and wound healing agents. (5) To crystallize and determine the structures of selected heparin-binding proteins not homologous to PF4.