PDF(Pubmed)
Abstract：
Antithrombin (AT) is a critical regulator of the coagulation cascade by inhibiting multiple coagulation factors including thrombin and FXa. Binding of heparinoids to this serpin enhances the inhibition considerably. Mutations located in the heparin binding site of AT result in thrombophilia in affected individuals. Our aim was to study 10 antithrombin mutations known to affect their heparin binding in a heparin pentasaccharide bound state using two molecular dynamics (MD) based methods providing enhanced sampling, GaMD and LiGaMD2. The latter provides an additional boost to the ligand and the most important binding site residues. From our GaMD simulations we were able to identify four variants (three affecting amino acid Arg47 and one affecting Lys114) that have a particularly large effect on binding. The additional acceleration provided by LiGaMD2 allowed us to study the consequences of several other mutants including those affecting Arg13 and Arg129. We were able to identify several conformational types by cluster analysis. Analysis of the simulation trajectories revealed the causes of the impaired pentasaccharide binding including pentasaccharide subunit conformational changes and altered allosteric pathways in the AT protein. Our results provide insights into the effects of AT mutations interfering with heparin binding at an atomic level and can facilitate the design or interpretation of in vitro experiments.
摘要：
抗凝血酶(AT)是通过抑制包括凝血酶和FXa在内的多种凝血因子的凝血级联的关键调节剂。类肝素与该serpin的结合大大增强了抑制作用。位于AT的肝素结合位点中的突变导致受影响个体的血栓形成倾向。我们的目的是使用两种基于分子动力学（MD）的方法来研究10种已知影响肝素五糖结合状态的肝素结合的抗凝血酶突变，提供增强的采样，GaMD和LiGaMD2。后者为配体和最重要的结合位点残基提供了额外的加强。从我们的GaMD模拟中,我们能够鉴定对结合具有特别大的影响的四种变体(三种影响氨基酸Arg47,一种影响Lys114)。LiGaMD2提供的额外加速使我们能够研究其他几种突变体的后果，包括影响Arg13和Arg129的突变体。我们能够通过聚类分析确定几种构象类型。模拟轨迹的分析揭示了五糖结合受损的原因，包括AT蛋白中的五糖亚基构象变化和变构途径改变。我们的结果提供了对AT突变在原子水平上干扰肝素结合的影响的见解，并且可以促进体外实验的设计或解释。
