Department of Chemistry and Biochemistry, University of California at San Diego, La Jolla, CA 92093, USA.
Proc Natl Acad Sci U S A. 2012 Dec 26;109(52):21216-22. doi: 10.1073/pnas.1218414109. Epub 2012 Nov 28.
The serine protease α-thrombin is a dual-action protein that mediates the blood-clotting cascade. Thrombin alone is a procoagulant, cleaving fibrinogen to make the fibrin clot, but the thrombin-thrombomodulin (TM) complex initiates the anticoagulant pathway by cleaving protein C. A TM fragment consisting of only the fifth and sixth EGF-like domains (TM56) is sufficient to bind thrombin, but the presence of the fourth EGF-like domain (TM456) is critical to induce the anticoagulant activity of thrombin. Crystallography of the thrombin-TM456 complex revealed no significant structural changes in thrombin, suggesting that TM4 may only provide a scaffold for optimal alignment of protein C for its cleavage by thrombin. However, a variety of experimental data have suggested that the presence of TM4 may affect the dynamic properties of the active site loops. In the present work, we have used both conventional and accelerated molecular dynamics simulation to study the structural dynamic properties of thrombin, thrombin:TM56, and thrombin:TM456 across a broad range of time scales. Two distinct yet interrelated allosteric pathways are identified that mediate both the pro- and anticoagulant activities of thrombin. One allosteric pathway, which is present in both thrombin:TM56 and thrombin:TM456, directly links the TM5 domain to the thrombin active site. The other allosteric pathway, which is only present on slow time scales in the presence of the TM4 domain, involves an extended network of correlated motions linking the TM4 and TM5 domains and the active site loops of thrombin.
丝氨酸蛋白酶α-凝血酶是一种双重作用的蛋白质，介导血液凝固级联反应。凝血酶本身是促凝剂，可裂解纤维蛋白原形成纤维蛋白凝块，但凝血酶-血栓调节蛋白（TM）复合物通过裂解蛋白 C 启动抗凝途径。仅由第五和第六个表皮生长因子样结构域（TM56）组成的 TM 片段足以与凝血酶结合，但第四个表皮生长因子样结构域（TM456）的存在对于诱导凝血酶的抗凝活性至关重要。凝血酶-TM456 复合物的晶体学研究表明凝血酶没有明显的结构变化，这表明 TM4 可能仅提供一个支架，用于最佳对齐蛋白 C 以使其被凝血酶裂解。然而，各种实验数据表明 TM4 的存在可能会影响活性位点环的动态特性。在本工作中，我们使用传统和加速分子动力学模拟来研究凝血酶、凝血酶：TM56 和凝血酶：TM456 在广泛的时间尺度上的结构动态特性。确定了两种独特但相互关联的变构途径，介导凝血酶的促凝和抗凝活性。一种变构途径存在于凝血酶：TM56 和凝血酶：TM456 中，直接将 TM5 结构域与凝血酶活性位点连接起来。另一种变构途径仅在存在 TM4 结构域的慢时间尺度上存在，涉及连接 TM4 和 TM5 结构域以及凝血酶活性位点环的扩展相关运动网络。
